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Abstract The composition and function of the intestinal microbiota are major determinants of human health and are strongly influenced by diet, antibiotic treatment, lifestyle and geography. Nevertheless, we currently have only little data on microbiomes of non-westernized communities. We assess the stool microbiota composition in 59 children aged 2–5 years from the Adadle district of Ethiopia, Somali Regional State. Here, milk and starch-rich food are predominant components of the local diet, where the inhabitants live a remote, traditional agropastoral lifestyle. Microbiota composition, function and the resistome were characterized by both 16S rRNA gene amplicon and shotgun metagenomic sequencing and compared to 1471 publicly available datasets from children living in traditional, transitional, and industrial communities with different subsistence strategies. Samples from the Adadle district are low in Bacteroidaceae, and Prevotellaceae, the main bacterial representatives in the feces of children living in industrialized and non-industrialized communities, respectively. In contrast, they had a higher relative abundance in Streptococcaceae, Bifidobacteriaceae and Erysipelatoclostridiaceae. Further, genes involved in degradation pathways of lactose, d-galactose and simple carbohydrates were enriched. Overall, our study revealed a unique composition of the fecal microbiota of these agropastoral children, highlighting the need to further characterize the fecal bacterial composition of human populations living different lifestyles.

BackgroundChildren in Lao People’s Democratic Republic (Lao PDR) receive suboptimal nutrition because of low breastfeeding rates, undermining their developmental potential. While major public health campaigns have attempted to increase breastfeeding rates, they have been largely unsuccessful. One explanation for these unsuccessful interventions is the economic and financial constraints faced by mothers. A potential solution for alleviating these pressures is providing social transfers to support breastfeeding; defined as a cash or in-kind transfer. Capitalizing on key strategies used in previous social transfer programs, we will assess the effectiveness of social transfer intervention for increasing exclusive breastfeeding rates in Vientiane, Lao PDR. ObjectiveThis study aims to conduct a randomized controlled trial (RCT) designed to assess whether social transfers can increase exclusive breastfeeding rates in Vientiane Capital, Lao PDR. MethodsA prospective, parallel cluster-RCT was conducted among 300 mothers who recently gave birth and initiated breastfeeding. Enrolling 100 participants for each intervention arm provided us with 80% power to detect an increase in exclusive breastfeeding from the anticipated 21% in the control arm to 40% in either of the 2 intervention arms. Mother-infant dyads were enrolled at approximately 1 month post partum. Follow-up visits will occur at 6 months, 1 year, 2 years, and 3 years post partum; with the ambition to extend the follow-up period. Mother-infant dyads were enrolled between August 2022 and April 2023 with follow-up until 3 years post partum (2026). A local study team comprised of 2 nurses and 2 laboratory technicians is responsible for enrollment and follow-up of participants. Participants were randomly assigned to one of three groups during the baseline, 1-month visit: (1) control group, no social transfer; (2) intervention group 1, an unconditional social transfer at 6 months post partum; and (3) intervention group 2, a social transfer at 6 months post partum conditional upon mothers exclusively breastfeeding. All groups received educational materials supporting mothers to exclusively breastfeed. The primary end point will be exclusive breastfeeding at 6 months post partum. Secondary end points will include exclusive and complementary breastfeeding duration, childhood wasting and stunting, child growth, maternal and infant stress, predictors of early breastfeeding cessation, intestinal inflammation, anemia, maternal weight loss, maternal blood pressure, maternal anxiety, and GRIT personality score. Questionnaires and physical examinations were used to collect information. ResultsAs of November 2023, the study has enrolled 300 participants. Study participation is ongoing until December 2026 at minimum. Over the study lifetime, 93% have completed all visits. ConclusionsWe see potential for a long-term program that may be implemented in other low- or lower-middle-income countries with only minor modifications. The RCT will be used as a basis for observational studies and to investigate the impact of human milk on child fecal microbiota and growth. Trial RegistrationClinicalTrials.gov NCT05665049; https://clinicaltrials.gov/study/NCT05665049 International Registered Report Identifier (IRRID)DERR1-10.2196/54768

There is an urgent need for interventions in addressing the rapid and disproportionate impact of antimicrobial resistance (AMR) and climate change (CC) on low- and middle-income countries. Within this context, it is important to understand indigenous knowledge in rural communities, which are highly affected. This study examined knowledge, attitude, and practices (KAP) regarding AMR and CC in the Adadle district, Somali region, Ethiopia, utilizing mixed methods, including 362 surveys and 12 focus group discussions among rural communities. Findings showed that 39% and 63% of participants were familiar with AMR and CC, respectively. Of those surveyed, 57% attributed AMR to inappropriate antimicrobial use in animals and humans, while CC was often associated with Allah/God. Multivariable analysis indicated that males exhibited superior knowledge and a positive attitude towards AMR and CC. Additionally, individuals aged 26–35 and 36–45 years showed heightened awareness of AMR and CC, respectively. Moreover, participants who were government employees, pastoralists, and business owners showed better knowledge on CC compared to family caretaker. Religious education and households with more than six members were linked to lower AMR knowledge. This study underlines a greater awareness of CC than AMR and highlights gender-based disparities, recommending integrated educational AMR programs targeting different demographics through a One Health lens, actively involving females, and incorporating local beliefs and practices.

BackgroundIntestinal parasitic infections (IPIs) can cause illness, morbidity, and occasional mortality in children. Agro-pastoralist and pastoralist children in the Somali Regional State of Ethiopia (ESRS) are especially at risk for IPIs, as access to safe water, sanitation, and health services is lacking. Minimal data on the prevalence of IPIs and associated risk factors exists in this region.MethodologyWe assessed the prevalence of IPIs and associated risk factors during the wet season from May-June 2021 in 366 children aged 2 to 5 years in four agro-pastoralist and four pastoralist kebeles (wards) in Adadle woreda (district) of the Shebelle zone, ESRS. Household information, anthropometric measurements, and stool samples were obtained from included children. Parasites were identified microscopically using Kato-Katz and direct smear methods. Risk factors were assessed using general estimating equation models accounting for clustering.Principal findingsOverall prevalence of IPIs was 35%: 30.6% for single infections and 4.4% for poly-parasitic infections. Intestinal protozoan prevalence was 24.9%: 21.9% Giardia intestinalis, and 3.0% Entamoeba spp.. Intestinal helminth prevalence was 14.5%: 12.8% Ascaris lumbricoides, 1.4% hookworm (Ancylostoma duodenale /Necator americanus.), and 0.3% Hymenolepis nana. G. intestinalis infection was associated with drinking water sourced from the river (aOR 15.6, 95%CI 6.84, 35.4) and from collected rainwater (aOR 9.48, 95%CI 3.39, 26.5), with toilet sharing (aOR 2.93, 95%CI 1.36, 6.31) and with household ownership of cattle (1-5 cattle: aOR 1.65, 95%CI 1.13, 2.41; 6+ cattle: aOR 2.07, 95%CI 1.33, 3.21) and chickens (aOR 3.80, 95%CI 1.77, 8.17). A. lumbricoides infection was associated with children 36 to 47 months old (aOR 1.92, 95%CI 1.03, 3.58).Conclusions/significanceImproving access to safe water, sanitation, and hygiene services in Adadle and employing a One Health approach would likely improve the health of children living in (agro-) pastoralist communities in Adadle and the ESRS; however, further studies are required.

Abstract Background Anaemia occurs in children when the haemoglobin level in the blood is less than the normal (11 g/dL), the consequence is the decrease of oxygen quantity in the tissues. It is a prevalent public health problem in many low-income countries, including Madagascar, and data on risk factors are lacking. We used existing data collected within the pathophysiology of environmental enteric dysfunction (EED) in Madagascar and the Central African Republic project (AFRIBIOTA project) conducted in underprivileged neighbourhoods of Antananarivo to investigate the factors associated with anaemia in children 24 to 59 months of age. Methods Children included in the AFRIBIOTA project in Antananarivo for whom data on haemoglobin and ferritin concentrations were available were included in the study. Logistic regression modelling was performed to identify factors associated with anaemia. Results Of the 414 children included in this data analysis, 24.4% were found to suffer from anaemia. We found that older children (adjusted OR: 0.95; 95% CI: 0.93–0.98) were less likely to have anaemia. Those with iron deficiency (adjusted OR: 6.1; 95% CI: 3.4–11.1) and those with a high level of faecal calprotectin (adjusted OR: 2.5; 95% CI: 1.4–4.4) were more likely to have anaemia than controls. Conclusions To reduce anaemia in the children in this underprivileged area, more emphasis should be given to national strategies that improve children’s dietary quality and micronutrient intake. Furthermore, existing measures should be broadened to include measures to reduce infectious disease burden.

IntroductionEarly life under- and overnutrition (jointly termed malnutrition) is increasingly recognized as an important risk factor for adult obesity and metabolic syndrome, a diet-related cluster of conditions including high blood sugar, fat and cholesterol. Nevertheless, the exact factors linking early life malnutrition with metabolic syndrome remain poorly characterized. We hypothesize that the microbiota plays a crucial role in this trajectory and that the pathophysiological mechanisms underlying under- and overnutrition are, to some extent, shared. We further hypothesize that a “dysbiotic seed microbiota” is transmitted to children during the birth process, altering the children’s microbiota composition and metabolic health. The overall objective of this project is to understand the precise causes and biological mechanisms linking prenatal or early life under- or overnutrition with the predisposition to develop overnutrition and/or metabolic disease in later life, as well as to investigate the possibility of a dysbiotic seed microbiota inheritance in the context of maternal malnutrition.Methods/designVITERBI GUT is a prospective birth cohort allowing to study the link between early life malnutrition, the microbiota and metabolic health. VITERBI GUT will include 100 undernourished, 100 normally nourished and 100 overnourished pregnant women living in Vientiane, Lao People’s Democratic Republic (PDR). Women will be recruited during their third trimester of pregnancy and followed with their child until its second birthday. Anthropometric, clinical, metabolic and nutritional data are collected from both the mother and the child. The microbiota composition of maternal and child’s fecal and oral samples as well as maternal vaginal and breast milk samples will be determined using amplicon and shotgun metagenomic sequencing. Epigenetic modifications and lipid profiles will be assessed in the child’s blood at 2 years of age. We will investigate for possible associations between metabolic health, epigenetics, and microbial changes.DiscussionWe expect the VITERBI GUT project to contribute to the emerging literature linking the early life microbiota, epigenetic changes and growth/metabolic health. We also expect this project to give new (molecular) insights into the mechanisms linking malnutrition-induced early life dysbiosis and metabolic health in later life, opening new avenues for microbiota-engineering using microbiota-targeted interventions.

IntroductionMicrobial isolates from culture can be identified using 16S or whole-genome sequencing which generates substantial costs and requires time and expertise. Protein fingerprinting via Matrix-assisted Laser Desorption Ionization–time of flight mass spectrometry (MALDI-TOF MS) is widely used for rapid bacterial identification in routine diagnostics but shows a poor performance and resolution on commensal bacteria due to currently limited database entries. The aim of this study was to develop a MALDI-TOF MS plugin database (CLOSTRI-TOF) allowing for rapid identification of non-pathogenic human commensal gastrointestinal bacteria.MethodsWe constructed a database containing mass spectral profiles (MSP) from 142 bacterial strains representing 47 species and 21 genera within the class Clostridia. Each strain-specific MSP was constructed using >20 raw spectra measured on a microflex Biotyper system (Bruker-Daltonics) from two independent cultures.ResultsFor validation, we used 58 sequence-confirmed strains and the CLOSTRI-TOF database successfully identified 98 and 93% of the strains, respectively, in two independent laboratories. Next, we applied the database to 326 isolates from stool of healthy Swiss volunteers and identified 264 (82%) of all isolates (compared to 170 (52.1%) with the Bruker-Daltonics library alone), thus classifying 60% of the formerly unknown isolates.DiscussionWe describe a new open-source MSP database for fast and accurate identification of the Clostridia class from the human gut microbiota. CLOSTRI-TOF expands the number of species which can be rapidly identified by MALDI-TOF MS.

Shigellosis is a leading cause of diarrhea and dysentery in young children from low to middle-income countries and adults experiencing traveler’s diarrhea worldwide. In addition to acute illness, infection by Shigella bacteria is associated with stunted growth among children, which has been linked to detrimental long-term health, developmental, and economic outcomes. On March 24 and 29, 2021, PATH convened an expert panel to discuss the potential impact of Shigella vaccines on these long-term outcomes. Based on current empirical evidence, this discussion focused on whether Shigella vaccines could potentially alleviate the long-term burden associated with Shigella infections. Also, the experts provided recommendations about how to best model the burden, health and vaccine impact, and economic consequences of Shigella infections. This international multidisciplinary panel included 13 scientists, physicians, and economists from multiple relevant specialties.According to the panel, while the relationship between Shigella infections and childhood growth deficits is complex, this relationship likely exists. Vaccine probe studies are the crucial next step to determine whether vaccination could ameliorate Shigella infection-related long-term impacts. Infants should be vaccinated during their first year of life to maximize their protection from severe acute health outcomes and ideally reduce stunting risk and subsequent negative long-term developmental and health impacts. With vaccine schedule crowding, targeted or combination vaccination approaches would likely increase vaccine uptake in high-burden areas. Shigella impact and economic assessment models should include a wider range of linear growth outcomes. Also, these models should produce a spectrum of results—ones addressing immediate benefits for usual health care decision-makers and others that include broader health impacts, providing a more comprehensive picture of vaccination benefits. While many of the underlying mechanisms of this relationship need better characterization, the remaining gaps can be best addressed by collecting data post-vaccine introduction or through large trials.

Malnourished children experience a high burden of intestinal pathogens that exacerbate growth stunting, and preventing this pathogen overgrowth has proved challenging. Here the authors show that diet-specific bacterial crossfeeding contributes to the overgrowth of intestinal pathogens during child malnutrition.

Environmental Enteric Dysfunction (EED) is an associate driver of stunting in poor settings, and intestinal infections indirectly contribute to the pathophysiology underlying EED. Our work aimed at assessing whether enteric viral carriage is determinant to stunting. A total of 464 healthy and asymptomatic children, aged 2 to 5 years, were recruited, and classified as non-stunted, moderately stunted, or severely stunted. Among the recruited children, 329 stool samples were obtained and screened for enteric and non-enteric viruses by real-time polymerase chain reaction. We statistically tested for the associations between enteric viral and potential risk factors. Approximately 51.7% of the stool samples were positive for at least one virus and 40.7% were positive for non-enteric adenoviruses. No statistical difference was observed between virus prevalence and the growth status of the children. We did not find any statistically significant association between viral infection and most of the socio-demographic risk factors studied, except for having an inadequate food quality score or an over-nourished mother. In addition, being positive for Ascaris lumbricoides was identified as a protective factor against viral infection. In conclusion, we did not find evidence of a direct link between stunting and enteropathogenic viral carriage in our population.

Stunting and environmental enteric dysfunction (EED) may be responsible for altered gut and systemic immune responses. However, their impact on circulating immune cell populations remains poorly characterized during early life. A detailed flow cytometry analysis of major systemic immune cell populations in 53 stunted and 52 non-stunted (2 to 5 years old) children living in Antananarivo (Madagascar) was performed. Compared to age-matched non-stunted controls, stunted children aged 2-3 years old had a significantly lower relative proportion of classical monocytes. No significant associations were found between stunting and the percentages of effector T helper cell populations (Th1, Th2, Th17, Th1Th17, and cTfh). However, we found that HLA-DR expression (MFI) on all memory CD4+ or CD8+ T cell subsets was significantly lower in stunted children compared to non-stunted controls. Interestingly, in stunted children compared to the same age-matched non-stunted controls, we observed statistically significant age-specific differences in regulatory T cells (Treg) subsets. Indeed, in 2- to 3-year-old stunted children, a significantly higher percentage of memory Treg, whilst a significantly lower percentage of naive Treg, was found. Our results revealed that both innate and adaptive systemic cell percentages, as well as activation status, were impacted in an age-related manner during stunting. Our study provides valuable insights into the understanding of systemic immune system changes in stunted children.

Environmental Enteric Dysfunction (EED) refers to an incompletely defined syndrome of inflammation, reduced absorptive capacity, and reduced barrier function in the small intestine. It is widespread among children and adults in low- and middle-income countries and is also associated with poor sanitation and certain gut infections possibly resulting in an abnormal gut microbiota, small intestinal bacterial overgrowth (SIBO) and stunting. We investigated bacterial pathogen exposure in stunted and non-stunted children in Antananarivo, Madagascar by collecting fecal samples from 464 children (96 severely stunted, 104 moderately stunted and 264 non-stunted) and the prevalence of SIBO in 109 duodenal aspirates from stunted children (61 from severely stunted and 48 from moderately stunted children). SIBO assessed by both aerobic and anaerobic plating techniques was very high: 85.3% when selecting a threshold of ≥105 CFU/ml of bacteria in the upper intestinal aspirates. Moreover, 58.7% of the children showed more than 106 bacteria/ml in these aspirates. The most prevalent cultivated genera recovered were Streptococcus, Neisseria, Staphylococcus, Rothia, Haemophilus, Pantoea and Branhamella. Feces screening by qPCR showed a high prevalence of bacterial enteropathogens, especially those categorized as being enteroinvasive or causing mucosal disruption, such as Shigella spp., enterotoxigenic Escherichia coli, enteropathogenic E. coli and enteroaggregative E. coli. These pathogens were detected at a similar rate in stunted children and controls, all showing no sign of severe diarrhea the day of inclusion but both living in a highly contaminated environment (slum-dwelling). Interestingly Shigella spp. was the most prevalent enteropathogen found in this study (83.3%) without overrepresentation in stunted children.

Abstract Background Child undernutrition is a global health issue that is associated with poor sanitation and an altered intestinal microbiota. Immunoglobulin (Ig) A mediates host-microbial homeostasis in the intestine, and acutely undernourished children have been shown to have altered IgA recognition of the fecal microbiota. We sought to determine whether chronic undernutrition (stunting) or intestinal inflammation were associated with antibody recognition of the microbiota using two geographically distinct populations from the Afribiota project. Fecal bacteria from 200 children between 2 and 5 years old in Antananarivo, Madagascar, and Bangui, Central African Republic (CAR), were sorted into IgA-positive (IgA+) and IgA-negative (IgA−) populations by flow cytometry and subsequently characterized by 16S rRNA gene sequencing to determine IgA-bacterial targeting. We additionally measured IgG+ fecal bacteria by flow cytometry in a subset of 75 children. Results Stunted children (height-for-age z-score ≤ −2) had a greater proportion of IgA+ bacteria in the fecal microbiota compared to non-stunted controls. This trend was consistent in both countries, despite the higher overall IgA-targeting of the microbiota in Madagascar, but lost significance in each country individually. Two of the most highly IgA-recognized bacteria regardless of nutritional status were Campylobacter (in CAR) and Haemophilus (in both countries), both of which were previously shown to be more abundant in stunted children; however, there was no association between IgA-targeting of these bacteria and either stunting or inflammatory markers. IgG-bound intestinal bacteria were rare in both stunted and non-stunted children, similar to levels observed in healthy populations. Conclusions Undernourished children carry a high load of intestinal pathogens and pathobionts. Our data suggest that stunted children have a greater proportion of IgA-recognized fecal bacteria. We moreover identify two putative pathobionts, Haemophilus and Campylobacter, that are broadly targeted by intestinal IgA. This study furthers our understanding of host-microbiota interactions in undernutrition and identifies immune-recognized microbes for future study.

Environmental enteric dysfunction (EED) is an elusive, inflammatory syndrome of the small intestine thought to be associated with enterocyte loss and gut leakiness and lead to stunted child growth. To date, the gold standard for diagnosis is small intestine biopsy followed by histology. Several putative biomarkers for EED have been proposed and are widely used in the field. Here, we assessed in a cross-sectional study of children aged 2–5 years for a large set of biomarkers including markers of protein exudation (duodenal and fecal alpha-1-antitrypsin (AAT)), inflammation (duodenal and fecal calprotectin, duodenal, fecal and blood immunoglobulins, blood cytokines, C-reactive protein (CRP)), gut permeability (endocab, lactulose-mannitol ratio), enterocyte mass (citrulline) and general nutritional status (branched-chain amino acids (BCAA), insulin-like growth factor) in a group of 804 children in two Sub-Saharan countries. We correlated these markers with each other and with anemia in stunted and non-stunted children. AAT and calprotectin, CRP and citrulline and citrulline and BCAA correlated with each other. Furthermore, BCAA, citrulline, ferritin, fecal calprotectin and CRP levels were correlated with hemoglobin levels. Our results show that while several of the biomarkers are associated with anemia, there is little correlation between the different biomarkers. Better biomarkers and a better definition of EED are thus urgently needed.

BackgroundThis study aimed to compare the prevalence of intestinal parasite infestations (IPIs) in stunted children, compared to control children, in Ankasina and Andranomanalina Isotry (two disadvantaged neighborhoods of Antananarivo, Madagascar), to characterize associated risk factors and to compare IPI detection by real-time PCR and standard microscopy techniques.Methodology/principal findingsFecal samples were collected from a total of 410 children (171 stunted and 239 control) aged 2-5 years. A single stool sample per subject was examined by simple merthiolate-iodine-formaldehyde (MIF), Kato-Katz smear and real-time PCR techniques. A total of 96.3% of the children were infested with at least one intestinal parasite. The most prevalent parasites were Giardia intestinalis (79.5%), Ascaris lumbricoides (68.3%) and Trichuris trichiura (68.0%). For all parasites studied, real-time PCR showed higher detection rates compared to microscopy (G. intestinalis [77.6% (n = 318) versus 20.9% (n = 86)], Entamoeba histolytica [15.8% (n = 65) versus 1.9% (n = 8)] and A. lumbricoides [64.1% (n = 263) versus 50.7% (n = 208)]). Among the different variables assessed in the study, age of 4 to 5 years (AOR = 4.61; 95% CI, (1.35-15.77)) and primary and secondary educational level of the mother (AOR = 12.59; 95% CI, (2.76-57.47); AOR = 9.17; 95% CI, (2.12-39.71), respectively) were significantly associated with IPIs. Children drinking untreated water was associated with infestation with G. intestinalis (AOR = 1.85; 95% CI, (1.1-3.09)) and E. histolytica (AOR = 1.9; 95% CI, (1.07-3.38)). E. histolytica was also associated with moderately stunted children (AOR = 0.37; 95% CI, 0.2-0.71). Similarly, children aged between 4 and 5 years (AOR = 3.2; 95% CI (2.04-5.01)) and living on noncemented soil types (AOR = 1.85; 95% CI, (1.18-2.09)) were associated with T. trichiura infestation.Conclusions/significanceThe prevalence of IPIs is substantial in the studied areas in both stunted and control children, despite the large-scale drug administration of antiparasitic drugs in the country. This high prevalence of IPIs warrants further investigation. Improved health education, environmental sanitation and quality of water sources should be provided.

Background:Campylobacter and Salmonella are the leading causes of foodborne diseases worldwide. Recently, antimicrobial resistance (AMR) has become one of the most critical challenges for public health and food safety. To investigate and detect infections commonly transmitted from animals, food, and the environment to humans, a surveillance–response system integrating human and animal health, the environment, and food production components (iSRS), called a One Health approach, would be optimal. Objective: We aimed to identify existing integrated One Health studies on foodborne illnesses in the Middle East and to determine the prevalence, serovars, and antimicrobial resistance phenotypes and genotypes of Salmonella and Campylobacter strains among humans and food-producing animals. Methods: The databases Web of Science, Scopus, and PubMed were searched for literature published from January 2010 until September 2021. Studies meeting inclusion criteria were included and assessed for risk of bias. To assess the temporal and spatial relationship between resistant strains from humans and animals, a statistical random-effects model meta-analysis was performed. Results: 41 out of 1610 studies that investigated Campylobacter and non-typhoid Salmonella (NTS) in the Middle East were included. The NTS prevalence rates among human and food-producing animals were 9% and 13%, respectively. The Campylobacter prevalence rates were 22% in humans and 30% in food-producing animals. The most-reported NTS serovars were Salmonella Enteritidis and Salmonella Typhimurium, while Campylobacter jejuni and Campylobacter coli were the most prevalent species of Campylobacter. NTS isolates were highly resistant to erythromycin, amoxicillin, tetracycline, and ampicillin. C. jejuni isolates showed high resistance against amoxicillin, trimethoprim–sulfamethoxazole, nalidixic acid, azithromycin, chloramphenicol, ampicillin, tetracycline, and ciprofloxacin. The most prevalent Antimicrobial Resistance Genes (ARGs) in isolates from humans included tetO (85%), Class 1 Integrons (81%), blaOXA-61 (53%), and cmeB (51%), whereas in food-producing animals, the genes were tetO (77%), Class 1 integrons (69%), blaOXA-61 (35%), and cmeB (35%). The One Health approach was not rigorously applied in the Middle East countries. Furthermore, there was an uneven distribution in the reported data between the countries. Conclusion: More studies using a simultaneous approach targeting human, animal health, the environment, and food production components along with a solid epidemiological study design are needed to better understand the drivers for the emergence and spread of foodborne pathogens and AMR in the Middle East.

Abstract Background Globally one out of four children under 5 years is affected by linear growth delay (stunting). This syndrome has severe long-term sequelae including increased risk of illness and mortality and delayed psychomotor development. Stunting is a syndrome that is linked to poor nutrition and repeated infections. To date, the treatment of stunted children is challenging as the underlying etiology and pathophysiological mechanisms remain elusive. We hypothesize that pediatric environmental enteropathy (PEE), a chronic inflammation of the small intestine, plays a major role in the pathophysiology of stunting, failure of nutritional interventions and diminished response to oral vaccines, potentially via changes in the composition of the pro- and eukaryotic intestinal communities. The main objective of AFRIBIOTA is to describe the intestinal dysbiosis observed in the context of stunting and to link it to PEE. Secondary objectives include the identification of the broader socio-economic environment and biological and environmental risk factors for stunting and PEE as well as the testing of a set of easy-to-use candidate biomarkers for PEE. We also assess host outcomes including mucosal and systemic immunity and psychomotor development. This article describes the rationale and study protocol of the AFRIBIOTA project. Methods AFRIBIOTA is a case-control study for stunting recruiting children in Bangui, Central African Republic and in Antananarivo, Madagascar. In each country, 460 children aged 2–5 years with no overt signs of gastrointestinal disease are recruited (260 with no growth delay, 100 moderately stunted and 100 severely stunted). We compare the intestinal microbiota composition (gastric and small intestinal aspirates; feces), the mucosal and systemic immune status and the psychomotor development of children with stunting and/or PEE compared to non-stunted controls. We also perform anthropological and epidemiological investigations of the children’s broader living conditions and assess risk factors using a standardized questionnaire. Discussion To date, the pathophysiology and risk factors of stunting and PEE have been insufficiently investigated. AFRIBIOTA will add new insights into the pathophysiology underlying stunting and PEE and in doing so will enable implementation of new biomarkers and design of evidence-based treatment strategies for these two syndromes.

Stunting remains a major public health concern worldwide. Although its global prevalence is slowly decreasing, the actual number of affected children is still rising in Sub-Saharan Africa. In the Central African Republic (CAR), about one third of all children below the age of five are stunted. Stunting is correlated with many long-term consequences, including poor cognitive development and a higher rate of morbidity and mortality, making stunting a major contributor to poverty. In CAR, little is known about the factors that contribute to stunting. This study aimed at analysing, in a cross-sectional study, the main factors associated with stunting in a group of 414 children recruited between December 2011 and November 2013, aged five years or less and living in Bangui. For all children, demographic, socio-economic and anthropometric data were recorded and asymptomatic enteropathogen carriage was assessed in stool samples using classical microbiological assays. The study group had a mean age of 14.2±10 months. Fifty-eight percent (292/414) were boys, and 36 percent (148/414) exhibited stunted growth. Of the stunted children, 51% (75/148) showed a moderate delay in linear growth for their age group [height-for-age z-score (HAZ) between -2 and -3 SD] while 49% (73/148) presented a severe delay (HAZ < -3). Factors significantly associated with stunting included gender (aOR: 1.67; 95% CI: 1.07; 2.62 for boys compared to girls) and age (aOR of 3.98 (95% CI: 2.45; 6.46) for toddlers and aOR 4.42 (95% CI: 2.36; 8.28) for children compared to infants). Most importantly, we identified being overweight [weight-for-height z-score (WHZ) > 2 SD; aOR: 3.21; 95% CI: 1.50; 6.90 of overweight compared to normal weight] as also being significantly associated with stunting. This is the first study showing that even in the poorest countries of the world there is an association of stunting with being overweight.

Abstract The pathogen Salmonella Typhimurium is a common cause of diarrhea and invades the gut tissue by injecting a cocktail of virulence factors into epithelial cells, triggering actin rearrangements, membrane ruffling and pathogen entry. One of these factors is SopE, a G‐nucleotide exchange factor for the host cellular Rho GTPases Rac1 and Cdc42. How SopE mediates cellular invasion is incompletely understood. Using genome‐scale RNAi screening we identified 72 known and novel host cell proteins affecting SopE‐mediated entry. Follow‐up assays assigned these ‘hits’ to particular steps of the invasion process; i.e., binding, effector injection, membrane ruffling, membrane closure and maturation of the Salmonella‐containing vacuole. Depletion of the COPI complex revealed a unique effect on virulence factor injection and membrane ruffling. Both effects are attributable to mislocalization of cholesterol, sphingolipids, Rac1 and Cdc42 away from the plasma membrane into a large intracellular compartment. Equivalent results were obtained with the vesicular stomatitis virus. Therefore, COPI‐facilitated maintenance of lipids may represent a novel, unifying mechanism essential for a wide range of pathogens, offering opportunities for designing new drugs.

Roux-en-Y gastric bypass (RYGB) results in long-term weight loss and reduced obesity related co-morbidities. However, little is known about how the lengths of the biliopancreatic limb (BPL), the alimentary limb (AL), and the common limb (CL) affect weight loss and glucose metabolism. Our aim was to establish a RYGB obese mouse model with defined proportions of the AL and BPL and a constant CL to assess the effects on weight loss,glucose metabolism, and obesity-related co-morbidities. In vivo mouse study. Six-week-old male C57BL/6J mice fed with a high-fat diet (HFD) underwent bariatric surgery with defined BPL lengths: a very long, long, and short BPL (35%, 25%, and 15% of total bowel length), or sham surgery. The length of the AL was adjusted to achieve the same CL length. Mice were analyzed for weight loss, glycemic control, and obesity-related co-morbidities. Mice undergoing RYGB surgery with a very long BPL had excessive weight loss and mortality and were therefore not further analyzed. Mice with a long BPL showed a significantly increased total weight loss when compared with mice with a short BPL. In addition, a long BPL improved glucose tolerance, particularly early after surgery. A long BPL was also associated with lower triglyceride levels. Resolution of hepatic steatosis and adipose tissue inflammation was, however, not statistically significant. Of note, bariatric surgery dramatically changed gut microbiota, regardless of limb length. In obese mice, a long BPL results in enhanced weight loss and improved glucose tolerance. These findings could potentially be translated to humans by tailoring the BPL length according to body weight, obesity-related co-morbidities, and total bowel length of an individual patient. [ABSTRACT FROM AUTHOR]

Targeting of permissive entry sites is crucial for bacterial infection. The targeting mechanisms are incompletely understood. We have analyzed target-site selection by S. Typhimurium. This enteropathogenic bacterium employs adhesins (e.g. fim) and the type III secretion system 1 (TTSS-1) for host cell binding, the triggering of ruffles and invasion. Typically, S. Typhimurium invasion is focused on a subset of cells and multiple bacteria invade via the same ruffle. It has remained unclear how this is achieved. We have studied target-site selection in tissue culture by time lapse microscopy, movement pattern analysis and modeling. Flagellar motility (but not chemotaxis) was required for reaching the host cell surface in vitro. Subsequently, physical forces trapped the pathogen for ∼1.5-3 s in "near surface swimming". This increased the local pathogen density and facilitated "scanning" of the host surface topology. We observed transient TTSS-1 and fim-independent "stopping" and irreversible TTSS-1-mediated docking, in particular at sites of prominent topology, i.e. the base of rounded-up cells and membrane ruffles. Our data indicate that target site selection and the cooperative infection of membrane ruffles are attributable to near surface swimming. This mechanism might be of general importance for understanding infection by flagellated bacteria.

Summary Shigella is a leading cause of dysentery worldwide, with the majority of infections caused by two subgroups, S. flexneri and S. sonnei . Although S. flexneri has been highly prevalent in low-income countries, global development has brought an increase in S. sonnei at the expense of S. flexneri. However, the mechanisms behind this shift are not understood. Here we report that S. sonnei , but not S. flexneri , encodes a type VI secretion system (T6SS) that provides a competitive advantage in the gut. S . sonnei competes against E . coli and S . flexneri in mixed cultures, but this advantage is reduced in T6SS mutant strains. In addition, S. sonnei can persist as well as outcompete E. coli and S . flexneri in mice in a T6SS-dependent manner. These findings suggest that S. sonnei has a competitive advantage over S. flexneri and potentially explain the increasing global prevalence of S . sonnei . [ABSTRACT FROM AUTHOR]

The small intestinal microbiota (SIM) is essential for gastrointestinal health, influencing digestion, immune modulation, and nutrient metabolism. Unlike the colonic microbiota, the SIM has been poorly characterized due to sampling challenges and ethical considerations. Current evidence suggests that the SIM consists of five core genera and additional segment-specific taxa. These bacteria closely interact with the human host, regulating nutrient absorption and metabolism. Recent work suggests the presence of two forms of small intestinal bacterial overgrowth, one dominated by oral bacteria (SIOBO) and a second dominated by coliform bacteria. Less invasive sampling techniques, omics approaches, and mechanistic studies will allow a more comprehensive understanding of the SIM, paving the way for interventions engineering the SIM towards better health., Competing Interests: Declaration of interests No interests are declared., (Copyright © 2024. Published by Elsevier Ltd.)

One-anastomosis gastric bypass (OAGB) has gained importance as a simple, safe, and effective operation to treat morbid obesity. We previously found that Roux-en-Y gastric bypass surgery with a long compared with a short biliopancreatic limb (BPL) leads to improved weight loss and glucose tolerance in obese mice. However, it is not known whether a long BPL in OAGB surgery also results in beneficial metabolic outcomes. Five-week-old male C57BL/6J mice fed a high-fat diet (HFD) for 8 weeks underwent OAGB surgery with defined BPL lengths (5.5 cm distally of the duodenojejunal junction for short and 9.5 cm for long BPL), or sham surgery combined with caloric restriction. Weight loss, glucose tolerance, obesity-related comorbidities, endocrine effects, gut microbiota, and bile acids were assessed. Total weight loss was independent of the length of the BPL after OAGB surgery. However, a long BPL was associated with lower glucose-stimulated insulin on day 14 , and an improved glucose tolerance on day 35 after surgery. Moreover, a long BPL resulted in reduced total cholesterol, while there were no differences in the resolution of metabolic dysfunction-associated steatotic liver disease (MASLD) and adipose tissue inflammation. Tendencies of an attenuated hypothalamic-pituitary-adrenal (HPA) axis and aldosterone were present in the long BPL group. With both the short and long BPL, we found an increase in primary conjugated bile acids (pronounced in long BPL) along with a loss in bacterial Desulfovibrionaceae and Erysipelotrichaceae and simultaneous increase in Akkermansiaceae , Sutterellaceae , and Enterobacteriaceae . In summary, OAGB surgery with a long compared with a short BPL led to similar weight loss, but improved glucose metabolism, lipid, and endocrine outcomes in obese mice, potentially mediated through changes in gut microbiota and related bile acids. Tailoring the BPL length in humans might help to optimize metabolic outcomes after bariatric surgery. NEW & NOTEWORTHY Weight loss following OAGB surgery in obese mice was not influenced by BPL length, but a longer BPL was associated with improved metabolic outcomes, including glucose and lipid homeostasis. These changes could be mediated by bile acids upon altered gut microbiota. Further validation of these findings is required through a randomized human study.

Background: Children in Lao People's Democratic Republic (Lao PDR) receive suboptimal nutrition because of low breastfeeding rates, undermining their developmental potential. While major public health campaigns have attempted to increase breastfeeding rates, they have been largely unsuccessful. One explanation for these unsuccessful interventions is the economic and financial constraints faced by mothers. A potential solution for alleviating these pressures is providing social transfers to support breastfeeding; defined as a cash or in-kind transfer. Capitalizing on key strategies used in previous social transfer programs, we will assess the effectiveness of social transfer intervention for increasing exclusive breastfeeding rates in Vientiane, Lao PDR., Objective: This study aims to conduct a randomized controlled trial (RCT) designed to assess whether social transfers can increase exclusive breastfeeding rates in Vientiane Capital, Lao PDR., Methods: A prospective, parallel cluster-RCT was conducted among 300 mothers who recently gave birth and initiated breastfeeding. Enrolling 100 participants for each intervention arm provided us with 80% power to detect an increase in exclusive breastfeeding from the anticipated 21% in the control arm to 40% in either of the 2 intervention arms. Mother-infant dyads were enrolled at approximately 1 month post partum. Follow-up visits will occur at 6 months, 1 year, 2 years, and 3 years post partum; with the ambition to extend the follow-up period. Mother-infant dyads were enrolled between August 2022 and April 2023 with follow-up until 3 years post partum (2026). A local study team comprised of 2 nurses and 2 laboratory technicians is responsible for enrollment and follow-up of participants. Participants were randomly assigned to one of three groups during the baseline, 1-month visit: (1) control group, no social transfer; (2) intervention group 1, an unconditional social transfer at 6 months post partum; and (3) intervention group 2, a social transfer at 6 months post partum conditional upon mothers exclusively breastfeeding. All groups received educational materials supporting mothers to exclusively breastfeed. The primary end point will be exclusive breastfeeding at 6 months post partum. Secondary end points will include exclusive and complementary breastfeeding duration, childhood wasting and stunting, child growth, maternal and infant stress, predictors of early breastfeeding cessation, intestinal inflammation, anemia, maternal weight loss, maternal blood pressure, maternal anxiety, and GRIT personality score. Questionnaires and physical examinations were used to collect information., Results: As of November 2023, the study has enrolled 300 participants. Study participation is ongoing until December 2026 at minimum. Over the study lifetime, 93% have completed all visits., Conclusions: We see potential for a long-term program that may be implemented in other low- or lower-middle-income countries with only minor modifications. The RCT will be used as a basis for observational studies and to investigate the impact of human milk on child fecal microbiota and growth., Trial Registration: ClinicalTrials.gov NCT05665049; https://clinicaltrials.gov/study/NCT05665049., International Registered Report Identifier (irrid): DERR1-10.2196/54768., (©Souliviengkham Sonephet, Sengchanh Kounnavong, Lucienne Zinsstag, Pascale Vonaesch, Somphou Sayasone, Latsamy Siengsounthone, Peter Odermatt, Günther Fink, Jordyn Tinka Wallenborn. Originally published in JMIR Research Protocols (https://www.researchprotocols.org), 03.05.2024.)

There is an urgent need for interventions in addressing the rapid and disproportionate impact of antimicrobial resistance (AMR) and climate change (CC) on low- and middle-income countries. Within this context, it is important to understand indigenous knowledge in rural communities, which are highly affected. This study examined knowledge, attitude, and practices (KAP) regarding AMR and CC in the Adadle district, Somali region, Ethiopia, utilizing mixed methods, including 362 surveys and 12 focus group discussions among rural communities. Findings showed that 39% and 63% of participants were familiar with AMR and CC, respectively. Of those surveyed, 57% attributed AMR to inappropriate antimicrobial use in animals and humans, while CC was often associated with Allah/God. Multivariable analysis indicated that males exhibited superior knowledge and a positive attitude towards AMR and CC. Additionally, individuals aged 26-35 and 36-45 years showed heightened awareness of AMR and CC, respectively. Moreover, participants who were government employees, pastoralists, and business owners showed better knowledge on CC compared to family caretaker. Religious education and households with more than six members were linked to lower AMR knowledge. This study underlines a greater awareness of CC than AMR and highlights gender-based disparities, recommending integrated educational AMR programs targeting different demographics through a One Health lens, actively involving females, and incorporating local beliefs and practices.

The number of research papers published on the involvement of the oral microbiota in systemic diseases has grown exponentially over the last 4 years clearly demonstrating the growing interest in this field. Indeed, accumulating evidence highlights the central role of ectopic colonization by oral bacteria in numerous noncommunicable diseases including inflammatory bowel diseases (IBDs), undernutrition, preterm birth, neurological diseases, liver diseases, lung diseases, heart diseases, or colonic cancer. There is thus much interest in understanding the molecular mechanisms that lead to the colonization and maintenance of ectopic oral bacteria. The aim of this review is to summarize and conceptualize the current knowledge about ectopic colonization by oral bacteria, highlight wherever possible the underlying molecular mechanisms and describe its implication in health and disease. The focus lies on the newly discovered molecular mechanisms, showcasing shared pathophysiological mechanisms across different body sites and syndromes and highlighting open questions in the field regarding the pathway from oral microbiota dysbiosis to noncommunicable diseases., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)

SUMMARYCommunities of microorganisms (microbiota) are present in all habitats on Earth and are relevant for agriculture, health, and climate. Deciphering the mechanisms that determine microbiota dynamics and functioning within the context of their respective environments or hosts (the microbiomes) is crucially important. However, the sheer taxonomic, metabolic, functional, and spatial complexity of most microbiomes poses substantial challenges to advancing our knowledge of these mechanisms. While nucleic acid sequencing technologies can chart microbiota composition with high precision, we mostly lack information about the functional roles and interactions of each strain present in a given microbiome. This limits our ability to predict microbiome function in natural habitats and, in the case of dysfunction or dysbiosis, to redirect microbiomes onto stable paths. Here, we will discuss a systematic approach (dubbed the N + 1/N-1 concept) to enable step-by-step dissection of microbiome assembly and functioning, as well as intervention procedures to introduce or eliminate one particular microbial strain at a time. The N+1/N-1 concept is informed by natural invasion events and selects culturable, genetically accessible microbes with well-annotated genomes to chart their proliferation or decline within defined synthetic and/or complex natural microbiota. This approach enables harnessing classical microbiological and diversity approaches, as well as omics tools and mathematical modeling to decipher the mechanisms underlying N+1/N-1 microbiota outcomes. Application of this concept further provides stepping stones and benchmarks for microbiome structure and function analyses and more complex microbiome intervention strategies., Competing Interests: The authors declare no conflict of interest.

Antimicrobial resistance (AMR) is a critical global concern driven by the overuse, misuse, and/or usage of inadequate antibiotics on humans, animals' agriculture, and as a result of contaminated environments. This study is the first One Health survey in the Middle East that incorporated whole-genome sequencing (WGS) to examine the spread of AMR in Campylobacter spp. and Salmonella spp. This cross-sectional study was conducted to examine the role of AMR at the human-animal-environmental interface and was performed in Ramallah/Al-Bireh and Jerusalem governorates of the central West Bank, Palestine. In 2021 and 2022, a total of 592 samples were collected and analyzed. From a total of 65 Campylobacter jejuni and 19 Salmonella spp. isolates, DNA was extracted for WGS using Oxford Nanopore Technologies MinION platform. We found that the dominant serotypes of C. jejuni and Salmonella enterica were present in chicken manure, chicken meat sold in markets, and feces of asymptomatic farm workers, with high genetic similarities between the isolates regardless of origin. Additionally, our results showed rapid strain turnover in C. jejuni from the same sites between 2021 and 2022. Most of the positive Salmonella spp. samples were multidrug-resistant (MDR) S. enterica serovar Muenchen carrying the plasmid of emerging S. infantis (pESI) megaplasmid, conferring resistance to multiple antibiotics. Our findings highlight the spread of MDR foodborne pathogens from animals to humans through the food chain, emphasizing the importance of a One Health approach that considers the interconnections between human, animal, and environmental health. IMPORTANCE Prior to this study, there existed hardly an integrated human-animal-environmental study of Salmonellosis and Campylobacteriosis and related AMR in Middle Eastern countries. The few existing studies lack robust epidemiological study designs, adequate for a One Health approach, and did not use WGS to determine the circulating serotypes and their AMR profiles. Civil unrest and war in Middle Eastern countries drive AMR because of the breakdown of public health and food security services. This study samples simultaneously humans, animals, and the environment to comprehensively investigate foodborne pathogens in the broiler chicken production chain in Palestine using WGS. We show that identical serotypes of C. jejuni and S. enterica can be found in samples from chicken farms, chicken meat sold in markets, and asymptomatic broiler chicken production workers. The most striking feature is the rapid dynamic of change in the genetic profile of the detected species in the same sampling locations. The majority of positive Salmonella spp. samples are MDR S. enterica serovar Muenchen isolates carrying the pESI megaplasmid. The results demonstrate a close relationship between the S. enterica serovar Muenchen isolates found in our sample collection and those responsible for 40% of all clinical Salmonella spp. isolates in Israel as previously reported, with a sequence identity of over 99.9%. These findings suggest the transboundary spread of MDR S. enterica serovar Muenchen strains from animals to humans through the food chain. The study underscores the importance of combining integrated One Health studies with WGS for detecting environmental-animal-human transmission of foodborne pathogens that could not be detected otherwise. This study showcases the benefits of integrated environmental-animal-human sampling and WGS for monitoring AMR. Environmental samples, which may be more accessible in conflict-torn places where monitoring systems are limited and regulations are weak, can provide an effective AMR surveillance solution. WGS of bacterial isolates provides causal inference of the distribution and spread of bacterial serotypes and AMR in complex social-ecological systems. Consequently, our results point toward the expected benefits of operationalizing a One Health approach through closer cooperation of public and animal health and food safety authorities., Competing Interests: The authors declare no conflict of interest.

Environmental Enteric Dysfunction (EED) is an associate driver of stunting in poor settings, and intestinal infections indirectly contribute to the pathophysiology underlying EED. Our work aimed at assessing whether enteric viral carriage is determinant to stunting. A total of 464 healthy and asymptomatic children, aged 2 to 5 years, were recruited, and classified as non-stunted, moderately stunted, or severely stunted. Among the recruited children, 329 stool samples were obtained and screened for enteric and non-enteric viruses by real-time polymerase chain reaction. We statistically tested for the associations between enteric viral and potential risk factors. Approximately 51.7% of the stool samples were positive for at least one virus and 40.7% were positive for non-enteric adenoviruses. No statistical difference was observed between virus prevalence and the growth status of the children. We did not find any statistically significant association between viral infection and most of the socio-demographic risk factors studied, except for having an inadequate food quality score or an over-nourished mother. In addition, being positive for Ascaris lumbricoides was identified as a protective factor against viral infection. In conclusion, we did not find evidence of a direct link between stunting and enteropathogenic viral carriage in our population.

Eukaryotes have historically been studied as parasites, but recent evidence suggests they may be indicators of a healthy gut ecosystem. Here, we describe the eukaryome along the gastrointestinal tract of children aged 2-5 years and test for associations with clinical factors such as anaemia, intestinal inflammation, chronic undernutrition, and age. Children were enrolled from December 2016 to May 2018 in Bangui, Central African Republic and Antananarivo, Madagascar. We analyzed a total of 1104 samples representing 212 gastric, 187 duodenal, and 705 fecal samples using a metabarcoding approach targeting the full ITS2 region for fungi, and the V4 hypervariable region of the 18S rRNA gene for the overall eukaryome. Roughly, half of all fecal samples showed microeukaryotic reads. We find high intersubject variability, only a handful of taxa that are likely residents of the gastrointestinal tract, and frequent co-occurrence of eukaryotes within an individual. We also find that the eukaryome differs between the stomach, duodenum, and feces and is strongly influenced by country of origin. Our data show trends towards higher levels of Fusarium equiseti , a mycotoxin producing fungus, and lower levels of the protist Blastocystis in stunted children compared to nonstunted controls. Overall, the eukaryome is poorly correlated with clinical variables. Our study is of one of the largest cohorts analyzing the human intestinal eukaryome to date and the first to compare the eukaryome across different compartments of the gastrointestinal tract. Our results highlight the importance of studying populations across the world to uncover common features of the eukaryome in health., Competing Interests: None declared., (© The Author(s) 2023. Published by Oxford University Press on behalf of FEMS.)

Background: Intestinal parasitic infections (IPIs) can cause illness, morbidity, and occasional mortality in children. Agro-pastoralist and pastoralist children in the Somali Regional State of Ethiopia (ESRS) are especially at risk for IPIs, as access to safe water, sanitation, and health services is lacking. Minimal data on the prevalence of IPIs and associated risk factors exists in this region., Methodology: We assessed the prevalence of IPIs and associated risk factors during the wet season from May-June 2021 in 366 children aged 2 to 5 years in four agro-pastoralist and four pastoralist kebeles (wards) in Adadle woreda (district) of the Shebelle zone, ESRS. Household information, anthropometric measurements, and stool samples were obtained from included children. Parasites were identified microscopically using Kato-Katz and direct smear methods. Risk factors were assessed using general estimating equation models accounting for clustering., Principal Findings: Overall prevalence of IPIs was 35%: 30.6% for single infections and 4.4% for poly-parasitic infections. Intestinal protozoan prevalence was 24.9%: 21.9% Giardia intestinalis, and 3.0% Entamoeba spp.. Intestinal helminth prevalence was 14.5%: 12.8% Ascaris lumbricoides, 1.4% hookworm (Ancylostoma duodenale /Necator americanus.), and 0.3% Hymenolepis nana. G. intestinalis infection was associated with drinking water sourced from the river (aOR 15.6, 95%CI 6.84, 35.4) and from collected rainwater (aOR 9.48, 95%CI 3.39, 26.5), with toilet sharing (aOR 2.93, 95%CI 1.36, 6.31) and with household ownership of cattle (1-5 cattle: aOR 1.65, 95%CI 1.13, 2.41; 6+ cattle: aOR 2.07, 95%CI 1.33, 3.21) and chickens (aOR 3.80, 95%CI 1.77, 8.17). A. lumbricoides infection was associated with children 36 to 47 months old (aOR 1.92, 95%CI 1.03, 3.58)., Conclusions/significance: Improving access to safe water, sanitation, and hygiene services in Adadle and employing a One Health approach would likely improve the health of children living in (agro-) pastoralist communities in Adadle and the ESRS; however, further studies are required., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Lanker et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Introduction: Early life under- and overnutrition (jointly termed malnutrition) is increasingly recognized as an important risk factor for adult obesity and metabolic syndrome, a diet-related cluster of conditions including high blood sugar, fat and cholesterol. Nevertheless, the exact factors linking early life malnutrition with metabolic syndrome remain poorly characterized. We hypothesize that the microbiota plays a crucial role in this trajectory and that the pathophysiological mechanisms underlying under- and overnutrition are, to some extent, shared. We further hypothesize that a "dysbiotic seed microbiota" is transmitted to children during the birth process, altering the children's microbiota composition and metabolic health. The overall objective of this project is to understand the precise causes and biological mechanisms linking prenatal or early life under- or overnutrition with the predisposition to develop overnutrition and/or metabolic disease in later life, as well as to investigate the possibility of a dysbiotic seed microbiota inheritance in the context of maternal malnutrition., Methods/design: VITERBI GUT is a prospective birth cohort allowing to study the link between early life malnutrition, the microbiota and metabolic health. VITERBI GUT will include 100 undernourished, 100 normally nourished and 100 overnourished pregnant women living in Vientiane, Lao People's Democratic Republic (PDR). Women will be recruited during their third trimester of pregnancy and followed with their child until its second birthday. Anthropometric, clinical, metabolic and nutritional data are collected from both the mother and the child. The microbiota composition of maternal and child's fecal and oral samples as well as maternal vaginal and breast milk samples will be determined using amplicon and shotgun metagenomic sequencing. Epigenetic modifications and lipid profiles will be assessed in the child's blood at 2 years of age. We will investigate for possible associations between metabolic health, epigenetics, and microbial changes., Discussion: We expect the VITERBI GUT project to contribute to the emerging literature linking the early life microbiota, epigenetic changes and growth/metabolic health. We also expect this project to give new (molecular) insights into the mechanisms linking malnutrition-induced early life dysbiosis and metabolic health in later life, opening new avenues for microbiota-engineering using microbiota-targeted interventions., Competing Interests: 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., (Copyright © 2023 Tamarelle, Creze, Savathdy, Phonekeo, Wallenborn, Siengsounthone, Fink, Odermatt, Kounnavong, Sayasone and Vonaesch.)

Introduction: Microbial isolates from culture can be identified using 16S or whole-genome sequencing which generates substantial costs and requires time and expertise. Protein fingerprinting via Matrix-assisted Laser Desorption Ionization-time of flight mass spectrometry (MALDI-TOF MS) is widely used for rapid bacterial identification in routine diagnostics but shows a poor performance and resolution on commensal bacteria due to currently limited database entries. The aim of this study was to develop a MALDI-TOF MS plugin database (CLOSTRI-TOF) allowing for rapid identification of non-pathogenic human commensal gastrointestinal bacteria., Methods: We constructed a database containing mass spectral profiles (MSP) from 142 bacterial strains representing 47 species and 21 genera within the class Clostridia . Each strain-specific MSP was constructed using >20 raw spectra measured on a microflex Biotyper system (Bruker-Daltonics) from two independent cultures., Results: For validation, we used 58 sequence-confirmed strains and the CLOSTRI-TOF database successfully identified 98 and 93% of the strains, respectively, in two independent laboratories. Next, we applied the database to 326 isolates from stool of healthy Swiss volunteers and identified 264 (82%) of all isolates (compared to 170 (52.1%) with the Bruker-Daltonics library alone), thus classifying 60% of the formerly unknown isolates., Discussion: We describe a new open-source MSP database for fast and accurate identification of the Clostridia class from the human gut microbiota. CLOSTRI-TOF expands the number of species which can be rapidly identified by MALDI-TOF MS., Competing Interests: 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., (Copyright © 2023 Asare, Lee, Hürlimann, Teo, Cuénod, Akduman, Gekeler, Afrizal, Corthesy, Kohout, Thomas, de Wouters, Greub, Clavel, Pamer, Egli, Maier and Vonaesch.)

Environmental enteric dysfunction (EED) is an inflammatory syndrome postulated to contribute to stunted child growth and to be associated with intestinal dysbiosis and nutrient malabsorption. However, the small intestinal contributions to EED remain poorly understood. This study aimed to assess changes in the proximal and distal intestinal microbiota in the context of stunting and EED and to test for a causal role of these bacterial isolates in the underlying pathophysiology. We performed a cross-sectional study in two African countries recruiting roughly 1,000 children aged 2 to 5 years and assessed the microbiota in the stomach, duodenum, and feces. Upper gastrointestinal samples were obtained from stunted children and stratified according to stunting severity. Fecal samples were collected. We then investigated the role of clinical isolates in EED pathophysiology using tissue culture and animal models. We find that small intestinal bacterial overgrowth (SIBO) is extremely common (>80%) in stunted children. SIBO is frequently characterized by an overgrowth of oral bacteria, leading to increased permeability and inflammation and to replacement of classical small intestinal strains. These duodenal bacterial isolates decrease lipid absorption in both cultured enterocytes and mice, providing a mechanism by which they may exacerbate EED and stunting. Further, we find a specific fecal signature associated with the EED markers fecal calprotectin and alpha-antitrypsin. Our study shows a causal implication of ectopic colonization of oral bacterial isolated from the small intestine in nutrient malabsorption and gut leakiness in vitro. These findings have important therapeutic implications for modulating the microbiota through microbiota-targeted interventions.

Shigellosis is a leading cause of diarrhea and dysentery in young children from low to middle-income countries and adults experiencing traveler's diarrhea worldwide. In addition to acute illness, infection by Shigella bacteria is associated with stunted growth among children, which has been linked to detrimental long-term health, developmental, and economic outcomes. On March 24 and 29, 2021, PATH convened an expert panel to discuss the potential impact of Shigella vaccines on these long-term outcomes. Based on current empirical evidence, this discussion focused on whether Shigella vaccines could potentially alleviate the long-term burden associated with Shigella infections. Also, the experts provided recommendations about how to best model the burden, health and vaccine impact, and economic consequences of Shigella infections. This international multidisciplinary panel included 13 scientists, physicians, and economists from multiple relevant specialties. According to the panel, while the relationship between Shigella infections and childhood growth deficits is complex, this relationship likely exists. Vaccine probe studies are the crucial next step to determine whether vaccination could ameliorate Shigella infection - related long-term impacts. Infants should be vaccinated during their first year of life to maximize their protection from severe acute health outcomes and ideally reduce stunting risk and subsequent negative long-term developmental and health impacts. With vaccine schedule crowding, targeted or combination vaccination approaches would likely increase vaccine uptake in high-burden areas. Shigella impact and economic assessment models should include a wider range of linear growth outcomes. Also, these models should produce a spectrum of results-ones addressing immediate benefits for usual health care decision-makers and others that include broader health impacts, providing a more comprehensive picture of vaccination benefits. While many of the underlying mechanisms of this relationship need better characterization, the remaining gaps can be best addressed by collecting data post-vaccine introduction or through large trials., Competing Interests: Dr. Kotloff receives funding from Institut Pasteur to conduct Shigella vaccine clinical trials. Dr. Lanata is a member of the World Health Organization (WHO) COVID-19 vaccine effectiveness working group and WHO Product Development Advisory Group. All other authors have no competing interests to declare., (© 2022 The Authors.)

Stunting and environmental enteric dysfunction (EED) may be responsible for altered gut and systemic immune responses. However, their impact on circulating immune cell populations remains poorly characterized during early life. A detailed flow cytometry analysis of major systemic immune cell populations in 53 stunted and 52 non-stunted (2 to 5 years old) children living in Antananarivo (Madagascar) was performed. Compared to age-matched non-stunted controls, stunted children aged 2-3 years old had a significantly lower relative proportion of classical monocytes. No significant associations were found between stunting and the percentages of effector T helper cell populations (Th1, Th2, Th17, Th1Th17, and cTfh). However, we found that HLA-DR expression (MFI) on all memory CD4 + or CD8 + T cell subsets was significantly lower in stunted children compared to non-stunted controls. Interestingly, in stunted children compared to the same age-matched non-stunted controls, we observed statistically significant age-specific differences in regulatory T cells (Treg) subsets. Indeed, in 2- to 3-year-old stunted children, a significantly higher percentage of memory Treg, whilst a significantly lower percentage of naive Treg, was found. Our results revealed that both innate and adaptive systemic cell percentages, as well as activation status, were impacted in an age-related manner during stunting. Our study provides valuable insights into the understanding of systemic immune system changes in stunted children., Competing Interests: 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., (Copyright © 2022 Andriamanantena, Randrianarisaona, Rakotondrainipiana, Andriantsalama, Randriamparany, Randremanana, Randrianirina, Novault, Duffy, Huetz, Hasan, Schoenhals, Sansonetti, Vonaesch, Vigan-Womas and Afribiota Investigators.)

Environmental Enteric Dysfunction (EED) refers to an incompletely defined syndrome of inflammation, reduced absorptive capacity, and reduced barrier function in the small intestine. It is widespread among children and adults in low- and middle-income countries and is also associated with poor sanitation and certain gut infections possibly resulting in an abnormal gut microbiota, small intestinal bacterial overgrowth (SIBO) and stunting. We investigated bacterial pathogen exposure in stunted and non-stunted children in Antananarivo, Madagascar by collecting fecal samples from 464 children (96 severely stunted, 104 moderately stunted and 264 non-stunted) and the prevalence of SIBO in 109 duodenal aspirates from stunted children (61 from severely stunted and 48 from moderately stunted children). SIBO assessed by both aerobic and anaerobic plating techniques was very high: 85.3% when selecting a threshold of ≥105 CFU/ml of bacteria in the upper intestinal aspirates. Moreover, 58.7% of the children showed more than 106 bacteria/ml in these aspirates. The most prevalent cultivated genera recovered were Streptococcus, Neisseria, Staphylococcus, Rothia, Haemophilus, Pantoea and Branhamella. Feces screening by qPCR showed a high prevalence of bacterial enteropathogens, especially those categorized as being enteroinvasive or causing mucosal disruption, such as Shigella spp., enterotoxigenic Escherichia coli, enteropathogenic E. coli and enteroaggregative E. coli. These pathogens were detected at a similar rate in stunted children and controls, all showing no sign of severe diarrhea the day of inclusion but both living in a highly contaminated environment (slum-dwelling). Interestingly Shigella spp. was the most prevalent enteropathogen found in this study (83.3%) without overrepresentation in stunted children., Competing Interests: The authors have declared that no competing interests exist.

Background: Campylobacter and Salmonella are the leading causes of foodborne diseases worldwide. Recently, antimicrobial resistance (AMR) has become one of the most critical challenges for public health and food safety. To investigate and detect infections commonly transmitted from animals, food, and the environment to humans, a surveillance-response system integrating human and animal health, the environment, and food production components (iSRS), called a One Health approach, would be optimal. Objective : We aimed to identify existing integrated One Health studies on foodborne illnesses in the Middle East and to determine the prevalence, serovars, and antimicrobial resistance phenotypes and genotypes of Salmonella and Campylobacter strains among humans and food-producing animals. Methods : The databases Web of Science, Scopus, and PubMed were searched for literature published from January 2010 until September 2021. Studies meeting inclusion criteria were included and assessed for risk of bias. To assess the temporal and spatial relationship between resistant strains from humans and animals, a statistical random-effects model meta-analysis was performed. Results : 41 out of 1610 studies that investigated Campylobacter and non-typhoid Salmonella (NTS) in the Middle East were included. The NTS prevalence rates among human and food-producing animals were 9% and 13%, respectively. The Campylobacter prevalence rates were 22% in humans and 30% in food-producing animals. The most-reported NTS serovars were Salmonella Enteritidis and Salmonella Typhimurium, while Campylobacter jejuni and Campylobacter coli were the most prevalent species of Campylobacter . NTS isolates were highly resistant to erythromycin, amoxicillin, tetracycline, and ampicillin. C. jejuni isolates showed high resistance against amoxicillin, trimethoprim-sulfamethoxazole, nalidixic acid, azithromycin, chloramphenicol, ampicillin, tetracycline, and ciprofloxacin. The most prevalent Antimicrobial Resistance Genes (ARGs) in isolates from humans included tetO (85%), Class 1 Integrons (81%), blaOXA-61 (53%), and cmeB (51%), whereas in food-producing animals, the genes were tetO (77%), Class 1 integrons (69%), blaOXA-61 (35%), and cmeB (35%). The One Health approach was not rigorously applied in the Middle East countries. Furthermore, there was an uneven distribution in the reported data between the countries. Conclusion : More studies using a simultaneous approach targeting human, animal health, the environment, and food production components along with a solid epidemiological study design are needed to better understand the drivers for the emergence and spread of foodborne pathogens and AMR in the Middle East.