Detection of low-concentration host mRNA transcripts in Malawian children at risk for environmental enteropathy.

BACKGROUND AND OBJECTIVE: Transcriptomic analysis of fecal samples is an emerging method for the diagnosis of gastrointestinal pathology because it is noninvasive and requires minute volumes of analyte; however, detection of mRNA in low copy numbers in human stool is challenging. Our objective was to develop a method for detecting human mRNA suggestive of environmental enteropathy (EE) in feces. METHODS: Stool samples from 70 Malawian children, 34 without EE and 36 with EE, as defined by dual sugar absorption, were used to develop the methodology for mRNA detection. Multiple RNA isolation techniques and polymerase chain reaction formats were tested to detect 38 potential mRNA biomarkers suggestive of EE, and the results compared. RESULTS: RNA isolation using magnetic bead extraction best recovered host mRNA in stool, and digital droplet polymerase chain reaction was the most sensitive format to detect low copy numbers of mRNA. In all of the 70 samples, >20 copies of glyceraldehyde-3-phosphate dehydrogenase/200 mg of stool were detected. Copy numbers of potential biomarkers were normalized to glyceraldehyde-3-phosphate dehydrogenase, to account for interspecimen differences in concentration of human mRNA. Of the 38 transcripts chosen for initial evaluation, 24 had copy numbers >10 in all of the samples tested. Of the 6 potential markers measured in all of the 70 samples, REG4 best differentiated children with and without EE. CONCLUSIONS: A reproducible and reliable method to quantify human mRNA in stool present in low copy numbers has been developed, and may prove useful in investigations of EE and possibly other inflammatory gut conditions.

Abnormal gut integrity is associated with reduced linear growth in rural Malawian children.

The aim of the present study was to investigate the relation of environmental enteropathy, as measured by the dual sugar absorption test, to linear growth faltering in 2- to 5-year-old Malawian children. Dietary quality, food insecurity, anthropometry, and site-specific sugar testing were measured in 418 children, and anthropometry was reassessed 3 months later. A linear regression model predicting linear growth was created. Better growth was associated with less urinary lactulose excretion, more clean water usage, not sleeping with animals, and no previous history of malnutrition. Eighty-seven percent of children studied demonstrated evidence of environmental enteropathy. In conclusion, abnormal gut integrity is associated with reduced linear growth in a population of rural African preschool-age children.

The duration of diarrhea and fever is associated with growth faltering in rural Malawian children aged 6-18 months.

Nutrition support programs that only focus upon better complementary feeding remain an insufficient means of limiting growth faltering in vulnerable populations of children. To determine if symptoms of acute infections correlate with the incidence of growth faltering in rural Malawian children, the associations between fever, diarrhea, and cough with anthropometric measures of stunting, wasting, and underweight were investigated. Data were analyzed from a trial where 209 children were provided with adequate complementary food and followed fortnightly from 6-18 months of age. Linear mixed model analysis was used to test for associations. Diarrheal disease was inversely associated with changes in height-for-age Z-score (HAZ), mid-upper arm circumference Z-score (MUACZ), and weight-for-age Z-score (WAZ). Fever was also inversely associated with changes in MUACZ and WAZ. These results suggest that initiatives to reduce febrile and diarrheal diseases are needed in conjunction with improved complementary feeding to limit growth faltering in rural Malawi.

Investigation of food acceptability and feeding practices for lipid nutrient supplements and blended flours used to treat moderate malnutrition.

OBJECTIVE: To examine acceptability and feeding practices associated with different supplementary food items and identify practices associated with weight gain. METHODS: Caregivers (n = 409) whose children had been enrolled in a trial comparing a fortified corn-soy blended flour (CSB++), soy ready-to-use supplementary food (RUSF), and soy/whey RUSF answered a questionnaire administered by health workers in their homes. RESULTS: No significant differences in acceptability of food types were found. CSB++ was more likely than soy RUSF or soy/whey RUSF to be shared (21% vs 3% vs 8%, respectively, P < .001). Children who received soy/whey RUSF were more likely to feed themselves than children who received soy RUSF or CSB++ (11% vs 4% vs 3%, respectively, P < .05). Refusing food was associated with slower weight gain. CONCLUSIONS AND IMPLICATIONS: Despite similar acceptability, feeding practices differed among food types. Increased nonstaple food consumption is associated with weight gain.