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BackgroundThere is no standardized approach to comparing socioeconomic status (SES) across multiple sites in epidemiological studies. This is particularly problematic when cross-country comparisons are of interest. We sought to develop a simple measure of SES that would perform well across diverse, resource-limited settings.MethodsA cross-sectional study was conducted with 800 children aged 24 to 60 months across eight resource-limited settings. Parents were asked to respond to a household SES questionnaire, and the height of each child was measured. A statistical analysis was done in two phases. First, the best approach for selecting and weighting household assets as a proxy for wealth was identified. We compared four approaches to measuring wealth: maternal education, principal components analysis, Multidimensional Poverty Index, and a novel variable selection approach based on the use of random forests. Second, the selected wealth measure was combined with other relevant variables to form a more complete measure of household SES. We used child height-for-age Z-score (HAZ) as the outcome of interest.ResultsMean age of study children was 41 months, 52% were boys, and 42% were stunted. Using cross-validation, we found that random forests yielded the lowest prediction error when selecting assets as a measure of household wealth. The final SES index included access to improved water and sanitation, eight selected assets, maternal education, and household income (the WAMI index). A 25% difference in the WAMI index was positively associated with a difference of 0.38 standard deviations in HAZ (95% CI 0.22 to 0.55).ConclusionsStatistical learning methods such as random forests provide an alternative to principal components analysis in the development of SES scores. Results from this multicountry study demonstrate the validity of a simplified SES index. With further validation, this simplified index may provide a standard approach for SES adjustment across resource-limited settings.

Background Rotavirus is the most common cause of severe dehydrating gastroenteritis in developing countries. Safe, effective, and affordable rotavirus vaccines are needed for developing countries. Methods In a double-blind placebo controlled multicentre trial, 6799 infants aged 6 to 7 weeks were randomised to receive three doses of an oral human-bovine natural reassortant vaccine (116E) or placebo at ages 6, 10, and 14 weeks. Primary outcome was severe (≥11 on the Vesikari scale) rotavirus gastroenteritis. Efficacy outcomes and adverse events were ascertained through active surveillance. Findings At analyses, the median age was 17·2 months; over 96% subjects received all three doses of the vaccine/placebo and ~1% were lost to follow up. 4532 and 2267 subjects were randomly assigned to receive vaccine and placebo, respectively. The per protocol analyses included 4354 subjects in the vaccine and 2187 subjects in the placebo group. 71 events of severe rotavirus gastroenteritis were reported in 4752 person years among the vaccinees compared to 76 events in 2360 person years in the placebo recipients; vaccine efficacy against severe rotavirus gastroenteritis was 53·6% (95% CI 35·0–66·9; P<0·001) and 56·4% (95% CI 36·6–70·1; P <0·001) in the first year of life. The number of infants needed to be immunized to prevent one severe rotavirus gastroenteritis episode was 55 (95% CI 37–97). The incidence of severe rotavirus gastroenteritis/100 person years was 1·5 in vaccine and 3·2 in placebo group and an incidence rate ratio of 0·46 (95% CI 0·33–0·65). The absolute rate reduction for severe rotavirus gastroenteritis was 1·7 (95% CI 2·5–0·9). Efficacy against severe gastroenteritis of any aetiology was 18·6% (95% CI 1·9–32·3); it was 24·1% (95% CI 5·8–38·7) in the first year of life. The prevalence of immediate, solicited, and serious adverse events were similar in both groups. There were six cases of intussusception amongst 4532 vaccinees and two amongst 2267 placebo recipients (P=0·73). All intussusception cases occurred after the third dose. Among vaccine and placebo recipients, the minimum interval between dosing and intussusception was 112 and 36 days, respectively. Interpretation The monovalent human-bovine (116E) rotavirus vaccine is effective and well-tolerated in Indian infants.

Human astroviruses are a leading cause of gastrointestinal disease. Since their discovery in 1975, 8 closely related serotypes have been described in humans, and more recently, two new astrovirus species, astrovirus MLB1 and astrovirus VA1, were identified in diarrhea patients. In this study, we used consensus astrovirus primers targeting the RNA polymerase to define the diversity of astroviruses present in pediatric patients with diarrhea on two continents. From 416 stool specimens comprising two different cohorts from Vellore, India, 35 samples were positive. These positive samples were analyzed further by either sequencing of the ~400 bp amplicon generated by the consensus PCR or by performing additional RT-PCR specific for individual astroviruses. 19 samples contained the classic human astrovirus serotypes 1-8 while 7 samples were positive for the recently described astrovirus MLB1. Strikingly, from samples that were positive in the consensus PCR screen but negative in the specific PCR assays, five samples contained sequences that were highly divergent from all previously described astroviruses. Sequence analysis suggested that three novel astroviruses, tentatively named astroviruses VA2, MLB2 and VA3, were present in these five patient specimens (AstV-VA2 in 2 patients, AstV-MLB2 in 2 patients and AstV-VA3 in one patient). Using the same RT-PCR screening strategy, 13 samples out of 466 tested stool specimens collected in St. Louis, USA were positive. Nine samples were positive for the classic human astroviruses. One sample was positive for AstV-VA2, and 3 samples were positive for AstV-MLB2 demonstrating that these two viruses are globally widespread. Collectively, these findings underscore the tremendous diversity of astroviruses present in fecal specimens from diarrhea patients. Given that a significant fraction of diarrhea etiologies is currently unknown, it is plausible that these or other yet unrecognized astroviruses may be responsible for at least part of the undiagnosed cases.

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.

BackgroundHuman immunology is a growing field of research in which experimental, clinical, and analytical methods of many life science disciplines are utilized. Classic epidemiological study designs, including observational longitudinal birth cohort studies, offer strong potential for gaining new knowledge and insights into immune response to pathogens in humans. However, rigorous discussion of methodological issues related to designs and statistical analysis that are appropriate for longitudinal studies is lacking.MethodsIn this communication we address key questions of quality and validity of traditional and recently developed statistical tools applied to measures of immune responses. For this purpose we use data on humoral immune response (IR) associated with the first cryptosporidial diarrhea in a birth cohort of children residing in an urban slum in south India. The main objective is to detect the difference and derive inferences for a change in IR measured at two time points, before (pre) and after (post) an event of interest. We illustrate the use and interpretation of analytical and data visualization techniques including generalized linear and additive models, data-driven smoothing, and combinations of box-, scatter-, and needle-plots.ResultsWe provide step-by-step instructions for conducting a thorough and relatively simple analytical investigation, describe the challenges and pitfalls, and offer practical solutions for comprehensive examination of data. We illustrate how the assumption of time irrelevance can be handled in a study with a pre-post design. We demonstrate how one can study the dynamics of IR in humans by considering the timing of response following an event of interest and seasonal fluctuation of exposure by proper alignment of time of measurements. This alignment of calendar time of measurements and a child's age at the event of interest allows us to explore interactions between IR, seasonal exposures and age at first infection.ConclusionsThe use of traditional statistical techniques to analyze immunological data derived from observational human studies can result in loss of important information. Detailed analysis using well-tailored techniques allows the depiction of new features of immune response to a pathogen in longitudinal studies in humans. The proposed staged approach has prominent implications for future study designs and analyses.

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Detection and quantification of enteropathogens in stool specimens is useful for diagnosing the cause of diarrhea but is technically challenging. Here we evaluate several important determinants of quantification: specimen collection, nucleic acid extraction, and extraction and amplification efficiency. First, we evaluate the molecular detection and quantification of pathogens in rectal swabs versus stool, using paired flocked rectal swabs and whole stool collected from 129 children hospitalized with diarrhea in Tanzania. Swabs generally yielded a higher quantification cycle (Cq) (average 29.7, standard deviation 3.5 vs. 25.3 ± 2.9 from stool, P<0.001) but were still able to detect 80% of pathogens with a Cq < 30 in stool. Second, a simplified total nucleic acid (TNA) extraction procedure was compared to separate DNA and RNA extractions and showed 92% (318/344) sensitivity and 98% (951/968) specificity, with no difference in Cq value for the positive results (ΔCq(DNA+RNA-TNA) = -0.01 ± 1.17, P = 0.972, N = 318). Third, we devised a quantification scheme that adjusts pathogen quantity to the specimen’s extraction and amplification efficiency, and show that this better estimates the quantity of spiked specimens than the raw target Cq. In sum, these methods for enteropathogen quantification, stool sample collection, and nucleic acid extraction will be useful for laboratories studying enteric disease.