Stunted children display ectopic small intestinal colonization by oral bacteria, which cause lipid malabsorption in experimental models.

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.

Metabolic outcomes in obese mice undergoing one-anastomosis gastric bypass (OAGB) with a long or a short biliopancreatic limb.

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.

Whole-genome sequencing for One Health surveillance of antimicrobial resistance in conflict zones: a case study of Salmonella spp. and Campylobacter spp. in the West Bank, Palestine.

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.

Factors associated with anaemia among preschool- age children in underprivileged neighbourhoods in Antananarivo, Madagascar.

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.

Systematic review and meta-analysis of integrated studies on antimicrobial resistance genes in Africa-A One Health perspective.

BACKGROUND: Increasing antimicrobial resistance (AMR) raises serious health and financial concerns. However, the main drivers of the emergence, spread and subsequent colonisation of resistant bacterial strains between humans, animals and the environment are still poorly understood. OBJECTIVE: The aim of this review was to identify molecular studies on AMR in One Health settings in Africa and to determine the prevalence of antimicrobial resistance genes in humans, animals and the environment. Due to the very low number of studies including environmental samples, the meta-analysis only includes data obtained from animals and humans. METHODS: The PubMed, Web of Science and Scopus databases were searched, identifying 10 464 publications on AMR in Africa from January 1st, 2000 until June 1st, 2020. Inclusion criteria were: (i) Integrated studies assessing AMR simultaneously in an animal-human, animal-environment, human-environment or animal-human-environment context, (ii) Genotypic characterisation of AMR and (iii) temporal and spatial relationship between samples from humans and animals. Statistical random-effects model meta-analysis was performed. RESULTS: Overall, 18 studies met our eligibility criteria and were included in this review. Six studies investigated Escherichia coli and Salmonella spp. (N = 6). The most prevalent AMR genes in animals included sul1 (36.2%), sul2 (32.0%), tetA (31.5%), strB (30.8%) and blaTEM (30.0%), whereas sul2 (42.4%), tetA (42.0%), strB (34.9%), blaTEM (28.8%) and sul1 (27.8%) were most prevalent in humans. We observed no clear pattern for a higher prevalence in either the animal or the human reservoir. CONCLUSION: To date, data on AMR in a One Health perspective in Africa are scarce. Prospective and longitudinal studies using an integrated One Health approach assessing the environment, animals and humans at the same time are needed to better understand the main drivers of AMR sharing in Africa.

Factors Associated with Stunted Growth in Children Under Five Years in Antananarivo, Madagascar and Bangui, Central African Republic.

OBJECTIVES: With a fourth of all under-five children affected, stunting remains one of the biggest health challenges worldwide. Even though the main underlying factors are known, the exact pathways to stunting varying in affected regions, and interventions thus need to be tailored to the local contexts. This study aimed assessing and comparing factors associated with stunting in two understudied sub-Saharan urban contexts with some of the highest stunting prevalence globally: Bangui, Central African Republic (~ 36%) and Antananarivo, Madagascar (42%). METHODS: We performed a case-control study on 175 + 194 stunted and 237 + 230 non-stunted control children aged 2-5 years and matched for age, gender and district of residency. Factors associated with stunting were identified using a standardized, paper questionnaire delivered by trained interviewers. Statistical analysis was done using logistic regression modelling. RESULTS: In both sites, formal maternal education lowered the risk of being stunted and restricted access to soap, suffering of anaemia and low birth weight were associated with higher risk of stunting. Short maternal stature, household head different from parents, diarrhoea and coughing were associated with an increased risk and continuing breastfeeding was associated with a lower risk of stunting in Antananarivo. Previous severe undernutrition and dermatitis/ fungal skin infections were associated with higher and changes in diet during pregnancy with lower risk of stunting in Bangui. CONCLUSIONS: Our results suggest maternal education, antenatal care, iron supplementation and simple WASH interventions such as using soap and infection control as general and breastfeeding (Antananarivo) or better nutrition (Bangui) as area-specified interventions.

Stunted childhood growth is associated with decompartmentalization of the gastrointestinal tract and overgrowth of oropharyngeal taxa.

Linear growth delay (stunting) affects roughly 155 million children under the age of 5 years worldwide. Treatment has been limited by a lack of understanding of the underlying pathophysiological mechanisms. Stunting is most likely associated with changes in the microbial community of the small intestine, a compartment vital for digestion and nutrient absorption. Efforts to better understand the pathophysiology have been hampered by difficulty of access to small intestinal fluids. Here, we describe the microbial community found in the upper gastrointestinal tract of stunted children aged 2-5 y living in sub-Saharan Africa. We studied 46 duodenal and 57 gastric samples from stunted children, as well as 404 fecal samples from stunted and nonstunted children living in Bangui, Central African Republic, and in Antananarivo, Madagascar, using 16S Illumina Amplicon sequencing and semiquantitative culture methods. The vast majority of the stunted children showed small intestinal bacterial overgrowth dominated by bacteria that normally reside in the oropharyngeal cavity. There was an overrepresentation of oral bacteria in fecal samples of stunted children, opening the way for developing noninvasive diagnostic markers. In addition, Escherichia coli/Shigella sp. and Campylobacter sp. were found to be more prevalent in stunted children, while Clostridia, well-known butyrate producers, were reduced. Our data suggest that stunting is associated with a microbiome "decompartmentalization" of the gastrointestinal tract characterized by an increased presence of oropharyngeal bacteria from the stomach to the colon, hence challenging the current view of stunting arising solely as a consequence of small intestine overstimulation through recurrent infections by enteric pathogens.

Identifying the etiology and pathophysiology underlying stunting and environmental enteropathy: study protocol of the AFRIBIOTA project.

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.

Shigella flexneri modulates stress granule composition and inhibits stress granule aggregation.

Invasion and multiplication of the facultative, cytosolic, enteropathogen Shigella flexneri within the colonic epithelial lining leads to an acute inflammatory response, fever and diarrhea. During the inflammatory process, infected cells are subjected to numerous stresses including heat, oxidative stress and genotoxic stress. The evolutionarily conserved pathway of cellular stress management is the formation of stress granules that store translationally inactive cellular mRNAs and interfere with cellular signalling pathways by sequestering signalling components. In this study, we investigated the ability of S. flexneri-infected cells to form stress granules in response to exogenous stresses. We found that S. flexneri infection inhibits movement of the stress granule markers eIF3 and eIF4B into stress granules and prevents the aggregation of G3BP1 and eIF4G-containing stress granules. This inhibition occurred only with invasive, but not with non-invasive bacteria and occurred in response to stresses that induce translational arrest through the phosphorylation of eIF2α and by treating cells with pateamine A, a drug that induces stress granules by inhibiting the eIF4A helicase. The S. flexneri-mediated stress granule inhibition could be largely phenocopied by the microtubule-destabilizing drug nocodazole and while S. flexneri infection did not lead to microtubule depolymerization, infection greatly enhanced acetylation of alpha-tubulin. Our data suggest that qualitative differences in the microtubule network or subversion of the microtubule-transport machinery by S. flexneri may be involved in preventing the full execution of this cellular stress response.

The Salmonella†Typhimurium effector protein SopE transiently localizes to the early SCV and contributes to intracellular replication.

Salmonella enterica serovar Typhimurium (S. Tm) is a facultative intracellular pathogen that induces entry into non-phagocytic cells by a Type III secretion system (TTSS) and cognate effector proteins. Upon host cell entry, S. Tm expresses a second TTSS and subverts intracellular trafficking to create a replicative niche - the Salmonella-containing vacuole (SCV). SopE, a guanidyl exchange factor (GEF) for Rac1 and Cdc42, is translocated by the TTSS-1 upon host cell contact and promotes entry through triggering of actin-dependent ruffles. After host cell entry, the bulk of SopE undergoes proteasomal degradation. Here we show that a subfraction is however detectable on the nascent SCV membrane up to ∼ 6 h post infection. Membrane localization of SopE and the closely related SopE2 differentially depend on the Rho-GTPase-binding GEF domain, and to some extent involves also the unstructured N-terminus. SopE localizes transiently to the early SCV, dependent on continuous synthesis and secretion by the TTSS-1 during the intracellular state. Mutant strains lacking SopE or SopE2 are attenuated in early intracellular replication, while complementation restores this defect. Hence, the present study reveals an unanticipated role for SopE and SopE2 in establishing the Salmonella replicative niche, and further emphasizes the importance of entry effectors in later stages of host-cell manipulation.

Near surface swimming of Salmonella Typhimurium explains target-site selection and cooperative invasion.

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.

Quantitative insights into actin rearrangements and bacterial target site selection from Salmonella†Typhimurium infection of micropatterned cells.

Reorganization of the host cell actin cytoskeleton is crucial during pathogen invasion. We established micropatterned cells as a standardized infection model for cell invasion to quantitatively study actin rearrangements triggered by Salmonella Typhimurium (S. Tm). Micropatterns of extracellular matrix proteins force cells to adopt a reproducible shape avoiding strong cell-to-cell variations, a major limitation in classical cell culture conditions. S. Tm induced F-actin-rich ruffles and invaded micropatterned cells similar to unconstrained cells. Yet, standardized conditions allowed fast and unbiased comparison of cellular changes triggered by the SipA and SopE bacterial effector proteins. Intensity measurements in defined regions revealed that the content of pre-existing F-actin remained unchanged during infection, suggesting that newly polymerized F-actin in bacteria-triggered ruffles originates from the G-actin pool. Analysing bacterial target sites, we found that bacteria did not show any preferences for the local actin cytoskeleton specificities. Rather, invasion was constrained to a specific 'cell height', due to flagella-mediated near-surface swimming. We found that invasion sites were similar to bacterial binding sites, indicating that S. Tm can induce a permissive invasion site wherever it binds. As micropatterned cells can be infected by many different pathogens they represent a valuable new tool for quantitative analysis of host-pathogen interactions.

Small intestinal microbiota: from taxonomic composition to metabolism.

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.

Ectopic colonization by oral bacteria as an emerging theme in health and disease.

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.

Social Transfers for Exclusive Breastfeeding (STEB) Intervention in Lao People's Democratic Republic: Protocol for a Randomized Controlled Trial.

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.

Knowledge, Attitudes, and Practices of Rural Communities Regarding Antimicrobial Resistance and Climate Change in Adadle District, Somali Region, Ethiopia: A Mixed-Methods Study.

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.

Prevalence and associated risk factors of intestinal parasitic infections among children in pastoralist and agro-pastoralist communities in the Adadle woreda of the Somali Regional State of Ethiopia.

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.

Dynamics and consequences of nutrition-related microbial dysbiosis in early life: study protocol of the VITERBI GUT project.

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.

Gut microbiomes of agropastoral children from the Adadle region of Ethiopia reflect their unique dietary habits.

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.

The eukaryome of African children is influenced by geographic location, gut biogeography, and nutritional status.

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.