Treatment Heterogeneity of Water, Sanitation, Hygiene, and Nutrition Interventions on Child Growth by Environmental Enteric Dysfunction and Pathogen Status for Young Children in Bangladesh.

Background: Water, sanitation, hygiene (WSH), nutrition (N), and combined (N+WSH) interventions are often implemented by global health organizations, but WSH interventions may insufficiently reduce pathogen exposure, and nutrition interventions may be modified by environmental enteric dysfunction (EED), a condition of increased intestinal permeability and inflammation. This study investigated the heterogeneity of these treatments' effects based on individual pathogen and EED biomarker status with respect to child linear growth. Methods: We applied cross-validated targeted maximum likelihood estimation and super learner ensemble machine learning to assess the conditional treatment effects in subgroups defined by biomarker and pathogen status. We analyzed treatment (N+WSH, WSH, N, or control) randomly assigned in-utero, child pathogen and EED data at 14 months of age, and child LAZ at 28 months of age. We estimated the difference in mean child length for age Z-score (LAZ) under the treatment rule and the difference in stratified treatment effect (treatment effect difference) comparing children with high versus low pathogen/biomarker status while controlling for baseline covariates. Results: We analyzed data from 1,522 children, who had median LAZ of -1.56. We found that myeloperoxidase (N+WSH treatment effect difference 0.0007 LAZ, WSH treatment effect difference 0.1032 LAZ, N treatment effect difference 0.0037 LAZ) and Campylobacter infection (N+WSH treatment effect difference 0.0011 LAZ, WSH difference 0.0119 LAZ, N difference 0.0255 LAZ) were associated with greater effect of all interventions on growth. In other words, children with high myeloperoxidase or Campylobacter infection experienced a greater impact of the interventions on growth. We found that a treatment rule that assigned the N+WSH (LAZ difference 0.23, 95% CI (0.05, 0.41)) and WSH (LAZ difference 0.17, 95% CI (0.04, 0.30)) interventions based on EED biomarkers and pathogens increased predicted child growth compared to the randomly allocated intervention. Conclusions: These findings indicate that EED biomarker and pathogen status, particularly Campylobacter and myeloperoxidase (a measure of gut inflammation), may be related to impact of N+WSH, WSH, and N interventions on child linear growth.

Early-Life Enteric Pathogen Exposure, Socioeconomic Status, and School-Age Cognitive Outcomes.

Early-life experiences of enteric infections and diarrheal illness are common in low-resource settings and are hypothesized to affect child development. However, longer-term associations of enteric infections with school-age cognitive outcomes are difficult to estimate due to lack of long-term studies. The objective of this study was to examine the relationship between enteropathogen exposure in the first 2 years of life with school-age cognitive skills in a cohort of children followed from birth until 6 to 8 years in low-resource settings in Brazil, Tanzania, and South Africa. The study included participants from three sites from the Etiology, Risk Factors, and Interactions of Enteric Infections and Malnutrition and the Consequences for Child Health Study who were enrolled just after birth and followed for enteric infections, diarrheal illness, and cognitive development until 2 years of age. When the children were school-age, further data were collected on reasoning skills and semantic/phonemic fluency. We estimated associations between the burden of specific enteric pathogens and etiology-specific diarrhea from 0 to 2 years with cognitive test scores at 6 to 8 years using linear regression and adjusting for confounding variables. In this study, children who carried more enteric pathogens in the first 2 years of life showed overall decreases in school-age cognitive abilities, particularly children who carried protozoa, although this was not statistically significant in this sample. Socioeconomic factors such as maternal education and income were more closely associated with school-age cognitive abilities. Early-life enteric pathogens may have a small, lasting influence on school-age cognitive outcomes, although other socioeconomic factors likely contribute more significantly.

Giardia hinders growth by disrupting nutrient metabolism independent of inflammatory enteropathy.

Giardia lamblia (Giardia) is among the most common intestinal pathogens in children in low- and middle-income countries (LMICs). Although Giardia associates with early-life linear growth restriction, mechanistic explanations for Giardia-associated growth impairments remain elusive. Unlike other intestinal pathogens associated with constrained linear growth that cause intestinal or systemic inflammation or both, Giardia seldom associates with chronic inflammation in these children. Here we leverage the MAL-ED longitudinal birth cohort and a model of Giardia mono-association in gnotobiotic and immunodeficient mice to propose an alternative pathogenesis of this parasite. In children, Giardia results in linear growth deficits and gut permeability that are dose-dependent and independent of intestinal markers of inflammation. The estimates of these findings vary between children in different MAL-ED sites. In a representative site, where Giardia associates with growth restriction, infected children demonstrate broad amino acid deficiencies, and overproduction of specific phenolic acids, byproducts of intestinal bacterial amino acid metabolism. Gnotobiotic mice require specific nutritional and environmental conditions to recapitulate these findings, and immunodeficient mice confirm a pathway independent of chronic T/B cell inflammation. Taken together, we propose a new paradigm that Giardia-mediated growth faltering is contingent upon a convergence of this intestinal protozoa with nutritional and intestinal bacterial factors.

The positive effect of malaria IPTp-SP on birthweight is mediated by gestational weight gain but modifiable by maternal carriage of enteric pathogens.

BACKGROUND: Poor pregnancy and birth outcomes are common in sub-Saharan Africa and have complex aetiologies. Sulfadoxine-pyrimethamine (SP), given for intermittent preventive therapy of malaria in pregnancy (IPTp), is one of few existing interventions that improves outcomes of both mother and baby despite widespread SP-resistant malaria. Compelling evidence exists that malaria-independent pathways contribute to this protective effect, but the exact sources of non anti-malarial protection remained unknown. We hypothesized that the beneficial effect of SP on birthweight is mediated by SP activity on maternal factors, including increased gestational weight gain and antibiotic activity on pathogens in the maternal gut. METHODS: Expectant mothers from a larger randomized control trial comparing the efficacy of IPTp-SP to IPTp with dihydroartemisinin-piperaquine (DP) were also enrolled in this sub-study study at their first antenatal care visit before commencement of IPTp (n = 105). Participants were followed monthly until delivery. Weights and mid-to-upper-arm circumferences (MUAC) were recorded. Monthly stool samples were collected and screened for five Escherichia coli pathotypes, Shigella spp., Vibrio cholerae, Salmonella, Campylobacter coli/jejuni, and three protozoa (Giardia spp., Entameba histolytica, and Cryptosporidium spp.) using previously validated molecular assays. FINDINGS: IPTp-SP vs. IPTP-DP was associated with higher maternal gestational weight gain (GWG) and nutritional indicators (MUAC and body-mass index, BMI). GWG was found to be a mediator of the birthweight and IPTp-SP relationship, as the birthweight of SP infants, but not DP infants, varied according to maternal GWG. The burden of maternal enteric infections was high. The three most commonly observed pathogens were enteroaggregative E. coli (EAEC), atypical enteropathogenic E.coli/enterohaemorrhagic E. coli (aEPEC/EHEC), and typical enteropathogenic E.coli (tEPEC). We found that SP reduced the prevalence of EAEC in a dose-dependent manner. After 3 or more doses, SP-recipients were 90% less likely to be infected with EAEC compared to DP-recipients (ORadj = 0.07, CI95 = 0.12, 0.39, p = 0.002). Compared to DP, this coincided with higher maternal gestational weight gain (GWG) and nutritional indicators (MUAC and body-mass index, BMI). The beneficial effect of SP on maternal GWG, MUAC and BMI, was lower if SP mothers had detectable EAEC, aEPEC/EHEC, tEPEC, and LT-ETEC at baseline. Maternal EAEC and tEPEC at baseline associated with lower birthweight for babies of both SP mothers and DP mothers. When comparing IPTp regimens, the positive effect of SP on birthweight compared to DP was only observed for infants of women who did not test positive for EAEC at baseline (adjusted mean birthweight difference SP vs. DP = 156.0 g, CI95 = -18.0 g, 336.9 g, p = 0.087), though confidence intervals crossed the null. INTERPRETATION: Our findings indicate that in pregnant Malawian women, IPTp-SP vs. IPTp-DP is consistently associated with higher MUAC, BMI, and GWG following the WHO-recommended regimen of at least 3 doses, but carriage of maternal gut pathogens before initiation of IPTp lessens this effect. Because GWG was a mediator of the association between birthweight and SP, we show that SP's previously proven positive effect on birthweight is by promoting maternal weight gain. Overall, our results present one plausible pathway SP exerts malaria-independent protection against poor birth outcomes in the context of its waning antimalarial activity and warrants further investigation. FUNDING: A full list of funding bodies that contributed to this study can be found in the Acknowledgements section.

Impact of Biannual Mass Azithromycin Treatment on Enteropathogen Carriage in Children <5 Years Old in Niger.

We analyzed samples obtained at baseline and 24 months in a mass azithromycin administration trial in Niger using quantitative polymerase chain reaction. In villages randomized to azithromycin, Shigella was the only pathogen reduced at 24 months (prevalence ratio, 0.36 [95% confidence interval: .17-.79]; difference in log quantity, -.42 [-.75 to -.10]).

Duration of Postdiarrheal Enteric Pathogen Carriage in Young Children in Low-resource Settings.

BACKGROUND: Prolonged enteropathogen shedding after diarrhea complicates the identification of etiology in subsequent episodes and is an important driver of pathogen transmission. A standardized approach has not been applied to estimate the duration of shedding for a wide range of pathogens. METHODS: We used a multisite birth cohort of children 0-24 months of age from whom diarrheal and monthly nondiarrheal stools were previously tested by quantitative polymerase chain reaction for 29 enteropathogens. We modeled the probability of detection of the etiologic pathogen before and after diarrhea using a log-normal accelerated failure time survival model and estimated the median duration of pathogen carriage as well as differences in subclinical pathogen carriage 60 days after diarrhea onset in comparison to a prediarrhea baseline. RESULTS: We analyzed 3247 etiologic episodes of diarrhea for the 9 pathogens with the highest attributable burdens of diarrhea. The median duration of postdiarrheal carriage varied widely by pathogen, from about 1 week for rotavirus (median, 8.1 days [95% confidence interval {CI}, 6.2-9.6]) to >1 month for Cryptosporidium (39.5 days [95% CI, 30.6-49.0]). The largest increases in subclinical pathogen carriage before and after diarrhea were seen for Cryptosporidium (prevalence difference between 30 days prior and 60 days after diarrhea onset, 0.30 [95% CI, .23-.39]) and Shigella (prevalence difference, 0.21 [95% CI, .16-.27]). CONCLUSIONS: Postdiarrheal shedding was widely variable between pathogens, with strikingly prolonged shedding seen for Cryptosporidium and Shigella. Targeted antimicrobial therapy and vaccination for these pathogens may have a relatively large impact on transmission.

Effect of water, sanitation, handwashing and nutrition interventions on enteropathogens in children 14 months old: a cluster-randomized controlled trial in rural Bangladesh.

BACKGROUND: We evaluated the impact of low-cost water, sanitation, handwashing (WSH) and child nutrition interventions on enteropathogen carriage in the WASH Benefits cluster-randomized controlled trial in rural Bangladesh. METHODS: We analyzed 1411 routine fecal samples from children 14±2 months old in the WSH (n = 369), nutrition counseling plus lipid-based nutrient supplement (n = 353), nutrition plus WSH (n = 360), and control (n = 329) arms for 34 enteropathogens using quantitative PCR. Outcomes included the number of co-occurring pathogens; cumulative quantity of four stunting-associated pathogens; and prevalence and quantity of individual pathogens. Masked analysis was by intention-to-treat. RESULTS: 326 (99.1%) control children had one or more enteropathogens detected (mean 3.8±1.8). Children receiving WSH interventions had lower prevalence and quantity of individual viruses than controls (prevalence difference for norovirus: -11% [95% confidence interval [CI], -5 to -17%]; sapovirus: -9% [95%CI, -3 to -15%]; and adenovirus 40/41: -9% [95%CI, -2 to - 15%]). There was no difference in bacteria, parasites, or cumulative quantity of stunting-associated pathogens between controls and any intervention arm. CONCLUSIONS: WSH interventions were associated with fewer enteric viruses in children aged 14 months. Different strategies are needed to reduce enteric bacteria and parasites at this critical young age.