Despite numerous nutrition support programs and infectious disease interventions, a reduction in linear growth (stunting) in children remains a prevalent public health problem (1). Stunting, a height-for-age z score (HAZ) <−2, affects an estimated 32% of children younger than 5 years in the developing world (1). Growth faltering among these children is not entirely explained by inadequate nutrition or acute infection (1–3). Environmental enteropathy (EE), an asymptomatic, diffuse villous atrophy of the small bowel associated with T-cell inflammatory infiltration and reduced intestinal integrity, is present in 20% to 80% of seemingly healthy, poor individuals living in the developing world (4–7). This damage promotes a chronic inflammatory state and reduces intestinal nutrient absorption, potentially contributing to malnutrition and poor growth.
EE can be assessed with a dual-sugar permeability test, in which mannitol and lactulose are consumed under controlled conditions and quantified in subsequent urine collection (7–9). Elevated lactulose excretion and/or reduced mannitol excretion reflect increased permeability of the gut mucosa and reduced surface area (10). Although studies have shown that increased permeability and/or surface area correlates with stunting in infants and toddlers, whether this relationship exists in older preschool children has not been described. We hypothesized that despite their greater resistance to environmental stressors compared with infants and toddlers, older children also would be at risk for stunting related to EE as reflected by increased gut permeability.
A total of 418 2- to 5-year-old children were recruited from 5 rural subsistence farming areas in southern Malawi in June and July of 2011 from a population enrolled in a gut microbiome study (11). These children were originally recruited because they were twins born in proximity to one of the surveillance sites, consuming a diet primarily of maize and legumes and had growth monitored every 2 to 3 months since birth. Exclusion criteria included any chronic debilitating illness, including known HIV infection or obvious congenital abnormalities, evidence of acute malnutrition, and a recent history or present case of diarrhea or hematochezia. The study was approved by the College of Medicine research ethics committee of the University of Malawi, the Washington University Human Research Protection Office, and the Baylor College of Medicine human investigations review board. Written and oral consent, as well as information regarding each child's household, hygienic practices, and recent health, were obtained at enrollment. Weight was measured using an electronic scale to the nearest 5 g. Length was measured to the nearest 0.5 cm using a canvas mat or the nearest 0.2 cm using a rigid length board. Mid-upper arm circumference was measured with a standard insertion tape to the nearest 0.2 cm. Edema was assessed, and household food insecurity and individual dietary diversity scores were measured using adapted versions of the indicator guides provided by Food and Nutrition Technical Assistance Project (12,13). Approximately 3 months after their initial visit, additional anthropometric measurements and health history were collected.
Caregivers were directed not to feed the child for 12 hours before participation. Children were given the nonmetabolized sugars, lactulose (5 g) and mannitol (1 g), dissolved in 100 mL of water. Urine was then collected for 4 hours. Ten milligrams of merthiolate was added to limit bacterial degradation of the sugars. Children were encouraged to drink water to facilitate urination and mothers were instructed not to breast-feed their children during this time. The total urine volume was measured and a 4-mL aliquot was transferred into a cryovial, flash frozen in liquid nitrogen, and transported at −80°C to Baylor College of Medicine.
The concentrations of lactulose and mannitol in the urine specimens were then analyzed by high-performance liquid chromatography as previously described (14). The assays are sensitive to 1-μg/mL lactulose and mannitol, and the coefficient of variation is ≤5% (14).
Anthropometric indices based on the World Health Organization's 2006 Child Growth Standards were calculated using Anthro version 3.1 (WHO, Geneva). The lactulose:mannitol ratio (L:M) and urinary excretion of lactulose and mannitol (as percentage of dose administered) were calculated. An L:M ratio ≥0.10 was defined as abnormal (14). Change in HAZ was compared between the children divided approximately into quartiles defined by the L:M and lactulose excretion. Comparisons of the means of each quartile were made using the Tukey-Kramer test.
Nonparametric bivariate correlations between change in HAZ and the demographic, sanitation, anthropometric, and gut function and integrity characteristics were calculated. Spearman correlation coefficients with a level of statistical significance <0.20 were used in linear regression modeling to determine the best predictors of linear growth. Backward linear regression modeling using change in HAZ as the dependent variable was then conducted (SPSS for Windows, version 20, SPSS Inc, Chicago, IL), eliminating terms with P > 0.10.
A total of 418 children completed the study (Table 1), of which 364 (87%) had a L:M suggestive of EE. Change in HAZ was associated with greater lactulose excretion, but not increased L:M (Fig. 1).
Urinary lactulose excretion, initial weight-for-height z score, initial HAZ, the number of bicycles in the home, the presence of animals sleeping in the home, the times per day clean water was brought to the home, and whether the child had previously been treated for malnutrition were placed into the linear regression model. The linear regression model to predict change in HAZ (r 2 = 0.52, F = 54.2, P < 0.001) found that urinary lactulose excretion, among other factors, was a predictor of linear growth (Table 2).
Most of the rural Malawian children selected for study had evidence of EE and abnormal gut integrity that was associated with reduced linear growth. The study is limited in that these study children were not surveyed for acute infectious episodes during the period in which growth was measured. Acute infections are likely to affect linear growth substantially in that they create a short-term inflammatory state associated with catabolic metabolism. The study population is typical of rural subsistence farmers in southern and eastern Africa, and thus these data should be used with caution when considering urban populations or populations outside of sub-Saharan Africa.
Other studies that have examined the relation between gut integrity and linear growth have found correlations between EE, as determined by dual-sugar absorption testing, and HAZ (8,9). A study in Nepal found that the lactulose:creatinine ratio explained 9% of the height deficit during a 7-month period in both “squatter” and middle-class populations of 3- to 18-month-old children (9). Degree of increase in the mannitol excretion, but not lactulose, has also been positively correlated with change in body weight and weight-for-length z score during a 3-month period in malnourished Bangladeshi children ages 6 to 24 months (15). In Gambian children ages 2 to 15 months, 43% of observed growth faltering could be explained by the L:M, but lactulose excretion alone was not correlated with growth outcomes (5,7). Differences in the pattern of the perturbation of the mannitol and lactulose excretion and L:M between different populations may represent different insults to the children's gastrointestinal mucosa mediated by nutritional status, age, or environment. Our study is the first in older preschool children to describe this association.
EE, although asymptomatic, has been found to be associated with growth faltering (10), diarrhea (14), and kwashiorkor (16). Stunting has been associated with lower school performance and decreased adult physical productivity (17). It has been hypothesized that poor hygiene and living in a contaminated environment can lead to or exacerbate EE (1,6). Further research is needed to determine the etiology of EE to develop appropriate interventions.
The high prevalence of EE found in this population is similar to that found in young children in other rural Malawian villages (14). The analyses highlight the importance of EE to growth in rural African children. It is interesting to note that neither maternal education nor socioeconomic status were significant predictors of linear growth, although this population had a limited variability in both of these parameters. The pervasiveness of EE in this population and its association with reduced linear growth demonstrate the importance of further investigation into appropriate and successful interventions for ameliorating EE.
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Keywords:Copyright 2012 by ESPGHAN and NASPGHAN
environmental enteropathy; intestinal integrity; lactulose mannitol test; linear growth