Dietary aflatoxins exposure, environmental enteropathy, and their relation with childhood stunting.

Childhood stunting is a global phenomenon affecting more than 149 million children under the age of 5 worldwide. Exposure to aflatoxins (AFs) in utero, during breastfeeding, and consumption of contaminated food affect the gut microbiome, resulting in intestinal dysfunction and potentially contributing to stunting. This review explores the potential relationship between AF exposure, environmental enteropathy and childhood stunting. AFs bind to DNA, disrupt protein synthesis and elicit environmental enteropathy (EE). An EE alters the structure of intestinal epithelial cells, impairs nutrient uptake and leads to malabsorption. This article proposes possible intervention strategies for researchers and policymakers to reduce AF exposure, EE and childhood stunting, such as exposure reduction, the implementation of good agricultural practices, dietary diversification and improving environmental water sanitation and hygiene.

Intestinal Biopsies for the Evaluation of Environmental Enteropathy and Environmental Enteric Dysfunction.

Environmental enteric dysfunction (EED) is a syndrome characterized by impairments of digestion and absorption and intestinal barrier failure in people living in insanitary or tropical environments. There is substantial evidence that it contributes to impaired linear growth of millions of children in low- and middle-income countries, to slowed neurocognitive development, and to diminished responses to oral vaccines. It represents the functional consequences of environmental enteropathy, an asymptomatic inflammatory disorder of the mucosa, and there is considerable overlap with the enteropathy observed in severe clinical malnutrition. The majority of studies of EED have employed functional tests based on lactulose permeation to define the presence of abnormal leak in the gut. However, where intestinal biopsies can safely be collected the opportunity then arises to study the underlying enteropathy in cellular and molecular detail, as well as to measure important functional elements such as enzyme expression. The purpose of this narrative review is to summarize the current understanding of environmental enteropathy obtained from small intestinal biopsies, and prospects for future work. We review histology, electron microscopy, transcription and protein expression, physiological measures, and the microbiome. We conclude that while noninvasive biomarkers of enteropathy and intestinal dysfunction permit large-scale studies of unquestionable value, intestinal biopsies are still required to investigate pathophysiology in depth.

Effects of Water, Sanitation, Handwashing, and Nutritional Interventions on Environmental Enteric Dysfunction in Young Children: A Cluster-randomized, Controlled Trial in Rural Bangladesh.

BACKGROUND: We hypothesized that drinking water, sanitation, handwashing (WSH), and nutritional interventions would improve environmental enteric dysfunction (EED), a potential contributor to stunting. METHODS: Within a subsample of a cluster-randomized, controlled trial in rural Bangladesh, we enrolled pregnant women in 4 arms: control, WSH, child nutrition counseling plus lipid-based nutrient supplements (N), and nutrition plus WSH (N+WSH). Among the birth cohort, we measured biomarkers of gut inflammation (myeloperoxidase, neopterin), permeability (alpha-1-antitrypsin, lactulose, mannitol), and repair (regenerating gene 1ß) at median ages 3, 14, and 28 months. Analysis was intention-to-treat. RESULTS: We assessed 1512 children. At age 3 months, compared to controls, neopterin was reduced by nutrition (-0.21 log nmol/L; 95% confidence interval [CI], -.37, -.05) and N+WSH (-0.20 log nmol/L; 95% CI, -.34, -.06) interventions; similar reductions were observed at 14 months. At 3 months, all interventions reduced lactulose and mannitol (-0.60 to -0.69 log mmol/L). At 28 months, myeloperoxidase was elevated in the WSH and nutrition arms (0.23-0.27 log ng/mL) and lactulose was higher in the WSH arm (0.30 log mmol/L; 95% CI, .07, .53). CONCLUSIONS: Reductions in permeability and inflammation at ages 3 and 14 months suggest that the interventions promoted healthy intestinal maturation; however, by 28 months, the WSH and nutrition arms showed elevated EED biomarkers. These results underscore the importance of developing a better understanding of EED pathophysiology and targeting interventions early in childhood, when they are likely to have the largest benefit to intestinal health. CLINICAL TRIALS REGISTRATION: NCT01590095.

Ingestion of Fecal Bacteria along Multiple Pathways by Young Children in Rural Bangladesh Participating in a Cluster-Randomized Trial of Water, Sanitation, and Hygiene Interventions (WASH Benefits).

Quantifying the contribution of individual exposure pathways to a child's total ingestion of fecal matter could help prioritize interventions to reduce environmental enteropathy and diarrhea. This study used data on fecal contamination of drinking water, food, soil, hands, and objects and second-by-second data on children's contacts with these environmental reservoirs in rural Bangladesh to assess the relative contribution of different pathways to children's ingestion of fecal indicator bacteria and if ingestion decreased with the water, sanitation, and hygiene interventions implemented in the WASH Benefits Trial. Our model estimated that rural Bangladeshi children <36 months old consume 3.6-4.9 log10 most probable number E. coli/day. Among children <6 months, placing objects in the mouth accounted for 60% of E. coli ingested. For children 6-35 months old, mouthing their own hands, direct soil ingestion, and ingestion of contaminated food were the primary pathways of E. coli ingestion. The amount of E. coli ingested by children and the predominant pathways of E. coli ingestion were unchanged by the water, sanitation, and hygiene interventions. These results highlight contaminated soil, children's hands, food, and objects as primary pathways of E. coli ingestion and emphasize the value of intervening along these pathways.

Study of Environmental Enteropathy and Malnutrition (SEEM) in Pakistan: protocols for biopsy based biomarker discovery and validation.

BACKGROUND: Environmental Enteropathy (EE), characterized by alterations in intestinal structure, function, and immune activation, is believed to be an important contributor to childhood undernutrition and its associated morbidities, including stunting. Half of all global deaths in children < 5 years are attributable to under-nutrition, making the study of EE an area of critical priority. METHODS: Community based intervention study, divided into two sub-studies, 1) Longitudinal analyses and 2) Biopsy studies for identification of EE features via omics analyses. Birth cohorts in Matiari, Pakistan established: moderately or severely malnourished (weight for height Z score (WHZ) < - 2) children, and well-nourished (WHZ > 0) children. Blood, urine, and fecal samples, for evaluation of potential biomarkers, will be collected at various time points from all participants (longitudinal analyses). Participants will receive appropriate educational and nutritional interventions; non-responders will undergo further evaluation to determine eligibility for further workup, including upper gastrointestinal endoscopy. Histopathological changes in duodenal biopsies will be compared with duodenal biopsies obtained from USA controls who have celiac disease, Crohn's disease, or who were found to have normal histopathology. RNA-Seq will be employed to characterize mucosal gene expression across groups. Duodenal biopsies, luminal aspirates from the duodenum, and fecal samples will be analyzed to define microbial community composition (omic analyses). The relationship between histopathology, mucosal gene expression, and community configuration will be assessed using a variety of bioinformatic tools to gain better understanding of disease pathogenesis and to identify mechanism-based biomarkers. Ethical review committees at all collaborating institutions have approved this study. All results will be made available to the scientific community. DISCUSSION: Operational and ethical constraints for safely obtaining intestinal biopsies from children in resource-poor settings have led to a paucity of human tissue-based investigations to understand and reverse EE in vulnerable populations. Furthermore, EE biomarkers have rarely been correlated with gold standard histopathological confirmation. The Study of Environmental Enteropathy and Malnutrition (SEEM) is designed to better understand the pathophysiology, predictors, biomarkers, and potential management strategies of EE to inform strategies to eradicate this debilitating pathology and accelerate progress towards the 2030 Sustainable Development Goals. TRIAL REGISTRATION: Retrospectively registered; clinicaltrials.gov ID NCT03588013 .

Amixicile Reduces Severity of Cryptosporidiosis but Does Not Have In Vitro Activity against Cryptosporidium.

Cryptosporidium species cause significant morbidity in malnourished children. Nitazoxanide (NTZ) is the only approved treatment for cryptosporidiosis, but NTZ has diminished effectiveness during malnutrition. Here, we show that amixicile, a highly selective water-soluble derivative of NTZ diminishes Cryptosporidium infection severity in a malnourished mouse model despite a lack of direct anticryptosporidial activity. We suggest that amixicile, by tamping down anaerobes associated with intestinal inflammation, reverses weight loss and indirectly mitigates infection-associated pathology.

Enteric Infections in Young Children are Associated with Environmental Enteropathy and Impaired Growth.

OBJECTIVE: To investigate the relationship between faecal contamination in child play spaces, enteric infections, environmental enteropathy (EE) and impaired growth among young children. METHODS: A prospective cohort study was conducted of 203 children 6-30 months of age in rural Bangladesh. Stool samples were analysed by quantitative PCR for Shigella, Enterotoxigenic Escherichia coli (ETEC), Campylobacter jejuni, Giardia intestinalis and Cryptosporidium spp. Four faecal markers of intestinal inflammation were also measured: alpha-1-antitrypsin, myeloperoxidase, neopterin and calprotectin. Child growth was measured at baseline and 9 months after enrolment. E. coli was measured in soil in child play spaces. RESULTS: Forty-seven percent of study children had three or more enteric pathogens in their stool. Thirty five percent (71/203) of children had Shigella, 30% (61/203) had ETEC, 73% (148/203) had C. jejuni, 79% (160/203) had Giardia intestinalis and none had Cryptosporidium. Children with ETEC had significantly higher calprotectin concentrations (Coefficient: 1.35, 95% Confidence Interval [CI]: 1.005, 1.82). Children with Shigella had a significantly higher odds of being stunted at our 9-month follow-up (OR: 2.01, 95% CI: 1.02, 3.93). Children with Giardia intestinalis had significantly higher E.coli counts in the soil collected from their play spaces (OR: 1.23, 95% CI: 1.02, 1.48). CONCLUSION: Enteric infections were significantly associated with EE and impaired growth in rural Bangladesh. These findings provide further evidence to support the hypothesis that contaminated soil in child play spaces can lead to enteric infections, many of which are likely subclinical, resulting in EE and impaired growth in young children.

Rotavirus epidemiology and vaccine demand: considering Bangladesh chapter through the book of global disease burden.

BACKGROUND: Rotavirus is the major cause of gastroenteritis in children throughout the world. Every year, a large number of children aged < 5 years die from rotavirus-related diarrhoeal diseases. Though these infections are vaccine-preventable, the vast majority of children in low-income countries suffer from the infection. The situation leads to severe economic loss and constitutes a major public health problem. METHODS: We searched electronic databases including PubMed and Google scholar using the following words: "features of rotavirus," "epidemiology of rotavirus," "rotavirus serotypes," "rotavirus in Bangladesh," "disease burden of rotavirus," "rotavirus vaccine," "low efficacy of rotavirus vaccine," "inactivated rotavirus vaccine". Publications until July 2017 have been considered for this work. RESULTS AND CONCLUSION: Currently, two live attenuated vaccines are available throughout the world. Many countries have included rotavirus vaccines in national immunization program to reduce the disease burden. However, due to low efficacy of the available vaccines, satisfactory outcome has not yet been achieved in developing countries such as Bangladesh. Poor economic, public health, treatment, and sanitation status of the low-income countries necessitate the need for the most effective rotavirus vaccines. Therefore, the present scenario demands the development of a highly effective rotavirus vaccine. In this regard, inactivated rotavirus vaccine concept holds much promise for reducing the current disease burden. Recent advancements in developing an inactivated rotavirus vaccine indicate a significant progress towards disease prophylaxis and control.

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.

A Combined Intervention of Zinc, Multiple Micronutrients, and Albendazole Does Not Ameliorate Environmental Enteric Dysfunction or Stunting in Rural Malawian Children in a Double-Blind Randomized Controlled Trial.

BACKGROUND: Environmental enteric dysfunction (EED) and linear growth stunting affect many rural agrarian children in the developing world and contribute to the persistently high rates of stunting that are observed worldwide. Effective interventions to consistently ameliorate EED are lacking. OBJECTIVE: We tested whether a bundle of safe and affordable interventions would decrease EED and stunting over 12-24 wk in a cohort of rural Malawian children 12-35 mo old. METHODS: This was a randomized, double-blind, placebo-controlled clinical trial in which the intervention group received a single dose of albendazole and 14 d of zinc at enrollment and after 20 wk. The intervention group also received a daily multiple micronutrient powder throughout the 24 wk of study. The primary outcomes were improvements in EED, as measured by the urinary lactulose-to-mannitol ratio (L:M ratio) from dual-sugar absorption testing, and linear growth. Urinary L:M ratios and anthropometric measurements were evaluated after 12 and 24 wk of intervention and compared with a placebo group that did not receive any of these interventions. RESULTS: A total of 254 children were enrolled at a mean age of 24 mo; 55% were female. Their mean weight-for-age z score was -1.5, and their mean length-for-age z score was -0.9. After 12 and 24 wk of study, increases in the L:M ratio did not differ between the intervention group (0.071 and 0.088 units, respectively) and the placebo group (0.073 and 0.080 units, respectively) (P = 0.87 and 0.19, respectively). Relative changes in length and weight also did not differ significantly between groups at any time point. CONCLUSION: The combined usage of albendazole, zinc, and a daily multiple micronutrient powder did not decrease EED or stunting in this population of agrarian children 12-35 mo old in rural Malawi. Alternative interventions to improve these diseases should be investigated. This trial was registered at clinicaltrials.gov as NCT02253095.

Mouthing of Soil Contaminated Objects is Associated with Environmental Enteropathy in Young Children.

OBJECTIVE: To characterise childhood mouthing behaviours and to investigate the association between object-to-mouth and food-to-mouth contacts, diarrhoea prevalence and environmental enteropathy. METHODS: A prospective cohort study was conducted of 216 children ≤30 months of age in rural Bangladesh. Mouthing contacts with soil and food and objects with visible soil were assessed by 5-h structured observation. Stool was analysed for four faecal markers of intestinal inflammation: alpha-1-antitrypsin, myeloperoxidase, neopterin and calprotectin. RESULTS: Overall 82% of children were observed mouthing soil, objects with visible soil, or food with visible soil during the structured observation period. Sixty two percent of children were observed mouthing objects with visible soil, 63% were observed mouthing food with visible soil, and 18% were observed mouthing soil only. Children observed mouthing objects with visible soil had significantly elevated faecal calprotectin concentrations (206.81 µg/g, 95% confidence interval [CI]: 6.27, 407.36). There was also a marginally significant association between Escherichia coli counts in soil from a child's play space and the prevalence rate of diarrhoea (diarrhoea prevalence ratio: 2.03, 95% CI 0.97, 4.25). CONCLUSION: These findings provide further evidence to support the hypothesis that childhood mouthing behaviour in environments with faecal contamination can lead to environmental enteropathy in susceptible paediatric populations. Furthermore, these findings suggest that young children mouthing objects with soil, which occurred more frequently than soil directly (60% vs. 18%), was an important exposure route to faecal pathogens and a risk factor for environmental enteropathy.

Tropical Enteropathies.

PURPOSE OF REVIEW: The term 'tropical enteropathy' originated in observations in the 1960s that small intestinal morphology and function differed in the tropics from the norms found in temperate climates. It was subsequently shown that this enteropathy is more closely related to environmental conditions than latitude, and it was re-labelled 'environmental enteropathy'. It is now recognised that environmental enteropathy (also now called environmental enteric dysfunction) has implications for the health and linear growth of children in low- and middle-income countries, and it may underlie poor responses to oral vaccination in these countries. The purpose of this review is to define and clarify this enteropathy despite the confusing terminology it has attracted and to contrast it with other enteropathic states. RECENT FINDINGS: Recent work has begun to demonstrate the nature of the mucosal lesion and the relationship with microbial translocation which is currently thought to link a failure of mucosal barrier function and the cascade of systemic inflammation which inhibits growth. The evidence is still correlative rather than definitive, but derives some additional support from animal models. There are some common features between environmental enteropathy and other enteropathies, but there are important differences also. The mechanism of the link between enteropathy and vaccine failure is not understood, and neither is it clear how the more severe form of enteropathy, which we refer to as malnutrition enteropathy, is driven by nutrient depletion and intestinal infection. Tropical enteropathies form a group of disorders which include environmental and nutritional enteropathies. The long-term health implications of these disorders for health in low-income countries are just being explored, but the scale of their effects is very large, with millions of people affected.

A fresh look at the hygiene hypothesis: how intestinal microbial exposure drives immune effector responses in atopic disease.

There currently is no consensus on which immunological mechanisms can best explain the rise in atopic disease post industrialization. The hygiene hypothesis lays groundwork for our understanding of how altered microbial exposures can drive atopy; yet since its introduction increasing evidence suggests the exposure of our immune system to the intestinal microbiota plays a key role in development of atopic disease. As societal change shifts our microbial exposure, concordant shifts in the tolerant and effector functions of our immune systems give rise to more hypersensitive responses to external antigens. This is contrasted with the greater immune tolerant capabilities of individuals still living in regions with lifestyles more representative of our evolutionary history. Recent findings, buoyed by technological advances in the field, suggest a direct role for the intestinal microbiota-immune system interplay in the development of atopic disease mechanisms. Overall, harnessing current mechanistic studies for translational research into microbiota composition and function in relation to atopy have potential for the design of therapeutics that could moderate these diseases.

Role of the Gut Microbiota of Children in Diarrhea Due to the Protozoan Parasite Entamoeba histolytica.

BACKGROUND: An estimated 1 million children die each year before their fifth birthday from diarrhea. Previous population-based surveys of pediatric diarrheal diseases have identified the protozoan parasite Entamoeba histolytica, the etiological agent of amebiasis, as one of the causes of moderate-to-severe diarrhea in sub-Saharan Africa and South Asia. METHODS: We prospectively studied the natural history of E. histolytica colonization and diarrhea among infants in an urban slum of Dhaka, Bangladesh. RESULTS: Approximately 80% of children were infected with E. histolytica by the age of 2 years. Fecal anti-galactose/N-acetylgalactosamine lectin immunoglobulin A was associated with protection from reinfection, while a high parasite burden and expansion of the Prevotella copri level was associated with diarrhea. CONCLUSIONS: E. histolytica infection was prevalent in this population, with most infections asymptomatic and diarrhea associated with both the amount of parasite and the composition of the microbiota.

Geophagy Is Associated with Growth Faltering in Children in Rural Bangladesh.

OBJECTIVE: To determine the relationship between geophagy (mouthing of dirt, sand, clay, or mud) and growth faltering in young children. STUDY DESIGN: We examined linear growth as height and weight standardized by age and sex, and weight standardized by height, in a cohort of children aged 6-36 months in rural Mirzapur, Bangladesh. We determined geophagy behavior at baseline through caregiver report. Anthropometric measurements were assessed at baseline and at a 1-year follow-up. RESULTS: We found that among children not stunted at baseline, those with caregiver-reported geophagy at baseline grew less over 1 year compared with their peers, with a difference in the change of standardized height for age and sex of -0.31 (95% CI, -0.61 to -0.01). CONCLUSION: These findings show that caregiver-reported geophagy was associated with growth faltering in a pediatric population in rural Bangladesh. Future studies are needed to learn more about this exposure pathway and its relevance to child growth.

Crohn's Disease: Evolution, Epigenetics, and the Emerging Role of Microbiome-Targeted Therapies.

Crohn's disease (CD) is a chronic, systemic, immune-mediated inflammation of the gastrointestinal tract. Originally described in 1932 as non-caseating granulomatous inflammation limited to the terminal ileum, it is now recognized as an expanding group of heterogeneous diseases defined by intestinal location, extent, behavior, and systemic extraintestinal manifestations. Joint diseases, including inflammatory spondyloarthritis and ankylosing spondylitis, are the most common extraintestinal manifestations of CD and share more genetic susceptibility loci than any other inflammatory bowel disease (IBD) trait. The high frequency and overlap with genes associated with infectious diseases, specifically Mendelian susceptibility to mycobacterial diseases (MSMD), suggest that CD may represent an evolutionary adaptation to environmental microbes. Elucidating the diversity of the enteric microbiota and the protean mucosal immune responses in individuals may personalize microbiome-targeted therapies and molecular classifications of CD. This review will focus on CD's natural history and therapies in the context of epigenetics, immunogenetics, and the microbiome.

Opportunities to assess factors contributing to the development of the intestinal microbiota in infants living in developing countries.

Recent evidence suggests that establishment of a healthy gut microbiota shortly after birth is important to achieve optimal growth and development of children. Being born into a resource-poor environment presents challenges to the establishment of a healthy gut microbial flora in the newborn. Among these challenges are births that occur at home, traditional pre-lacteal feeding of newborns leading to failure to initiate lactation, poor sanitation and water quality, early environmental exposure to, and infection with, enteric or other pathogens, suboptimal breast feeding duration and intensity, deficiencies in weaning and childhood diets contributing to micro- and macro-nutrient deficiencies, and the frequent use of antibiotics. These factors should be considered in the design and implementation of preventive and therapeutic interventions aimed at improving the health and development of these children.

Seroepidemiology of hepatitis E virus infection in an urban population in Zambia: strong association with HIV and environmental enteropathy.

BACKGROUND: Hepatitis E virus (HEV) infection causes major epidemics of infectious hepatitis, with high mortality rates in pregnant women. Recent reports indicate that HEV coinfections with human immunodeficiency virus (HIV) may have a more protracted course. However, the impact of HEV infections in communities heavily affected by HIV remains poorly studied. We set out to examine age-related seroprevalence in a community where we have previously carried out studies on environmental enteropathy. METHODS: Blood samples from 194 children and 106 adults were examined for immunoglobulin G and immunoglobulin M antibodies for HEV. HEV data were correlated with HIV status and morphometric analysis of small intestinal biopsies. RESULTS: Seroprevalence rose throughout childhood, from 8% in children aged 1-4 years, to 36% in children aged 10-14 years. In adults, the overall prevalence was 42%, with 28% in HIV-seronegative adults and 71% in HIV-seropositive adults (odds ratio, 6.2; 95% confidence interval, 2.2-18; P = .0001). In adults, villous height and crypt depth measurements showed that HEV seropositivity was associated with worse enteropathy (P = .05 and P = .005, respectively). CONCLUSIONS: HEV infection is common in Zambia. In adults it is strongly associated with HIV status, and also with environmental enteropathy.

Assessment of environmental enteropathy in the MAL-ED cohort study: theoretical and analytic framework.

Individuals in the developing world live in conditions of intense exposure to enteric pathogens due to suboptimal water and sanitation. These environmental conditions lead to alterations in intestinal structure, function, and local and systemic immune activation that are collectively referred to as environmental enteropathy (EE). This condition, although poorly defined, is likely to be exacerbated by undernutrition as well as being responsible for permanent growth deficits acquired in early childhood, vaccine failure, and loss of human potential. This article addresses the underlying theoretical and analytical frameworks informing the methodology proposed by the Etiology, Risk Factors and Interactions of Enteric Infections and Malnutrition and the Consequences for Child Health and Development (MAL-ED) cohort study to define and quantify the burden of disease caused by EE within a multisite cohort. Additionally, we will discuss efforts to improve, standardize, and harmonize laboratory practices within the MAL-ED Network. These efforts will address current limitations in the understanding of EE and its burden on children in the developing world.