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Background: Quantitative molecular assays are increasingly used for detection of enteric viruses., Methods: We compared the clinical severity using modified Vesikari score (mVS) of enteric viruses detected by conventional assays (enzyme immunoassays [EIA] for rotavirus and adenovirus 40/41 and conventional polymerase chain reaction for astrovirus, sapovirus, and norovirus) and a quantitative molecular assay (TaqMan Array Card [TAC]) among children aged 0-59 months in the Global Enteric Multicenter Study. For rotavirus and adenovirus 40/41, we compared severity between EIA-positive and TAC-positive cases assigned etiologies using different cycle threshold (CT) cutoffs., Results: Using conventional assays, the median (interquartile range) mVS was 10 (8, 11) for rotavirus, 9 (7, 11) for adenovirus 40/41, 8 (6, 10) for astrovirus, sapovirus, and norovirus GII, and 7 (6, 9) for norovirus GI. Compared to rotavirus EIA-positive cases, the median mVS was 2 and 3 points lower for EIA-negative/TAC-positive cases with CT<32.6 and 32.6≤CT<35, respectively (p-value<.0001). Adenovirus 40/41 EIA-positive and EIA-negative/TAC-positive cases were similar, regardless of CT cutoff., Conclusions: Quantitative molecular assays compared to conventional assays, such as EIA, may influence severity of identified cases, especially for rotavirus. Cutoffs to assign etiology for quantitative assays should be considered in the design and interpretation of enteric virus studies., (Published by Oxford University Press on behalf of Infectious Diseases Society of America 2024.)

Cryptosporidium species are a major cause of diarrhea and associated with growth failure. There is currently only limited knowledge of the parasite's genomic variability. We report a genomic analysis of Cryptosporidium parvum isolated from Bangladeshi infants and reanalysis of sequences from the United Kingdom. Human isolates from both locations shared 154 variants not present in the cattle-derived reference genome, suggesting host-specific adaptation of the parasite. Remarkably 34.6% of single-nucleotide polymorphisms unique to human isolates were nonsynonymous and 8.2% of these were in secreted proteins. Linkage disequilibrium decay indicated frequent recombination. The genetic diversity of C. parvum has potential implications for vaccine and therapeutic design. Clinical Trials Registration. NCT02764918., (© The Author(s) 2023. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Background: The association between childhood diarrheal disease and linear growth faltering in developing countries is well described. However, the impact attributed to specific pathogens has not been elucidated, nor has the impact of recommended antibiotic treatment., Methods: The Global Enteric Multicenter Study enrolled children with moderate to severe diarrhea (MSD) seeking healthcare at 7 sites in sub-Saharan Africa and South Asia. At enrollment, we collected stool samples to identify enteropathogens. Length/height was measured at enrollment and follow-up, approximately 60 days later, to calculate change in height-for-age z scores (ΔHAZ). The association of pathogens with ΔHAZ was tested using linear mixed effects regression models., Results: Among 8077 MSD cases analyzed, the proportion with stunting (HAZ below -1) increased from 59% at enrollment to 65% at follow-up (P < .0001). Pathogens significantly associated with linear growth decline included Cryptosporidium (P < .001), typical enteropathogenic Escherichia coli (P = .01), and untreated Shigella (P = .009) among infants (aged 0-11 months) and enterotoxigenic E. coli encoding heat-stable toxin (P < .001) and Cryptosporidium (P = .03) among toddlers (aged 12-23 months). Shigella-infected toddlers given antibiotics had improved linear growth (P = .02)., Conclusions: Linear growth faltering among children aged 0-23 months with MSD is associated with specific pathogens and can be mitigated with targeted treatment strategies, as demonstrated for Shigella., (© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases Society of America.)

Background: The Global Enteric Multicenter Study (GEMS) determined the etiologic agents of moderate-to-severe diarrhea (MSD) in children under 5 years old in Africa and Asia. Here, we describe the prevalence and antimicrobial susceptibility of nontyphoidal Salmonella (NTS) serovars in GEMS and examine the phylogenetics of Salmonella Typhimurium ST313 isolates., Methods: Salmonella isolated from children with MSD or diarrhea-free controls were identified by classical clinical microbiology and serotyped using antisera and/or whole-genome sequence data. We evaluated antimicrobial susceptibility using the Kirby-Bauer disk-diffusion method. Salmonella Typhimurium sequence types were determined using multi-locus sequence typing, and whole-genome sequencing was performed to assess the phylogeny of ST313., Results: Of 370 Salmonella-positive individuals, 190 (51.4%) were MSD cases and 180 (48.6%) were diarrhea-free controls. The most frequent Salmonella serovars identified were Salmonella Typhimurium, serogroup O:8 (C2-C3), serogroup O:6,7 (C1), Salmonella Paratyphi B Java, and serogroup O:4 (B). The prevalence of NTS was low but similar across sites, regardless of age, and was similar among both cases and controls except in Kenya, where Salmonella Typhimurium was more commonly associated with cases than controls. Phylogenetic analysis showed that these Salmonella Typhimurium isolates, all ST313, were highly genetically related to isolates from controls. Generally, Salmonella isolates from Asia were resistant to ciprofloxacin and ceftriaxone, but African isolates were susceptible to these antibiotics., Conclusions: Our data confirm that NTS is prevalent, albeit at low levels, in Africa and South Asia. Our findings provide further evidence that multidrug-resistant Salmonella Typhimurium ST313 can be carried asymptomatically by humans in sub-Saharan Africa., (© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases Society of America.)

Background: Shigella is a leading cause of childhood diarrhea and target for vaccine development. Microbiologic and clinical case definitions are needed for pediatric field vaccine efficacy trials., Methods: We compared characteristics of moderate to severe diarrhea (MSD) cases in the Global Enteric Multicenter Study (GEMS) between children with culture positive Shigella to those with culture-negative, quantitative polymerase chain reaction (qPCR)-attributable Shigella (defined by an ipaH gene cycle threshold <27.9). Among Shigella MSD cases, we determined risk factors for death and derived a clinical severity score., Results: Compared to culture-positive Shigella MSD cases (n = 745), culture-negative/qPCR-attributable Shigella cases (n = 852) were more likely to be under 12 months, stunted, have a longer duration of diarrhea, and less likely to have high stool frequency or a fever. There was no difference in dehydration, hospitalization, or severe classification from a modified Vesikari score. Twenty-two (1.8%) Shigella MSD cases died within the 14-days after presentation to health facilities, and 59.1% of these deaths were in culture-negative cases. Age <12 months, diarrhea duration prior to presentation, vomiting, stunting, wasting, and hospitalization were associated with mortality. A model-derived score assigned points for dehydration, hospital admission, and longer diarrhea duration but was not significantly better at predicting 14-day mortality than a modified Vesikari score., Conclusions: A composite severity score consistent with severe disease or dysentery may be a pragmatic clinical endpoint for severe shigellosis in vaccine trials. Reliance on culture for microbiologic confirmation may miss a substantial number of Shigella cases but is currently required to measure serotype specific immunity., (© The Author(s) 2020. Published by Oxford University Press for the Infectious Diseases Society of America.)

Background: Cryptosporidiosis is a major cause of childhood diarrhea in low- and middle-income countries and has been linked to impairment of child growth. This study investigated the burden of cryptosporidiosis and its impact on child growth in both a rural and an urban site in Bangladesh., Methods: Pregnant women in the second trimester were identified at 2 sites in Bangladesh, 1 urban and 1 rural. Their offspring were enrolled at birth into the study (urban, n = 250; rural, n = 258). For 2 years, the children were actively monitored for diarrhea and anthropometric measurements were obtained every 3 months. Stool samples were collected monthly and during diarrheal episodes with Cryptosporidium infection and causative species determined by quantitative polymerase chain reaction assays., Results: Cryptosporidium infections were common at both sites and mostly subclinical. In the urban site, 161 (64%) children were infected and 65 (26%) had ≥2 infections. In the rural site, 114 (44%) were infected and 24 (9%) had multiple infections. Adjusted for potential confounders, cryptosporidiosis was associated with a significantly greater drop in the length-for-age z score (LAZ) at 24 months from LAZ at enrollment (Δ-LAZ), an effect greatest in the children with multiple episodes of cryptosporidiosis. The most common species in Mirpur was Cryptosporidium hominis, whereas Cryptosporidium meleagridis predominated in Mirzapur., Conclusions: Cryptosporidiosis is common in early childhood and associated with early growth faltering in Bangladeshi children. Predominant Cryptosporidium species differed between the 2 sites, suggesting different exposures or modes of transmission but similar consequences for child growth., Clinical Trials Registration: NCT02764918.

We studied the genetic diversity of Cryptosporidium hominis infections in slum-dwelling infants from Dhaka over a 2-year period. Cryptosporidium hominis infections were common during the monsoon, and were genetically diverse as measured by gp60 genotyping and whole-genome resequencing. Recombination in the parasite was evidenced by the decay of linkage disequilibrium in the genome over <300 bp. Regions of the genome with high levels of polymorphism were also identified. Yet to be determined is if genomic diversity is responsible in part for the high rate of reinfection, seasonality, and varied clinical presentations of cryptosporidiosis in this population.

Background: The parasitic causes of diarrhea have historically been identified by use of microscopy; however, the use of this technique does not allow one to distinguish between subspecies or genotypes of parasites. Our objective was to determine, by use of modern diagnostic methods, the proportion of diarrhea cases in Bangladesh attributable to Cryptosporidium hominis, Cryptosporidium parvum, Entamoeba histolytica, and Giardia lamblia assemblages A and B., Methods: A prospective case-control study was performed involving 3646 case patients (both children and adults) who presented with diarrhea to the Dhaka hospital of the International Centre for Diarrhoeal Disease Research, Bangladesh, and 2575 control subjects with asymptomatic infection. Parasitic infection was detected by use of a stool parasite antigen test, and the parasite load and the species and/or genotypes were determined by use of polymerase chain reaction (PCR)., Results: Cryptosporidium species and E. histolytica were more prevalent in patients with acute diarrhea than in healthy control subjects, for all ages (2.1% vs. 1.4%; P = .039) and, specifically, for those 0-12 months of age (2.2% vs. 0.4%; P = .009). G. lamblia assemblage A was also more prevalent in case patients with diarrhea than in healthy control subjects (20% vs. 5%; P = .001). For case patients with diarrhea, the parasite load in feces, as measured by quantitative real-time PCR cycle threshold, was not higher that that for control subjects with asymptomatic infection. Case patients with diarrhea and cryptosporidiosis were less likely to have abdominal pain, compared with control subjects (15% vs. 37%; P = .001); case patients with amebiasis more likely to have visible blood in stool, compared with control subjects (8% vs. 1.6%; P = .001); and case patients with giardiasis more likely to be dehydrated, compared with control subjects (81% vs. 71%; P = .001)., Conclusion: E. histolytica, C. hominis, C. parvum, and G. lamblia assemblage A infections are important causes of diarrheal illness in Bangladesh.

Background: Infection with Vibrio cholerae induces durable immunity against subsequent disease, a process hypothesized to reflect anamnestic immune responses at the intestinal mucosa. The presence of antigen-specific memory B cells may therefore be a more direct measure of protection than serum antibody responses., Methods: We measured immunoglobulin (Ig) G memory B cells specific to cholera toxin B subunit (CTB) in 14 patients up to 90 days after V. cholerae O1 infection, by polyclonal stimulation of peripheral blood mononuclear cells followed by standard enzyme-linked immunospot assay., Results: All patients generated CTB-specific IgG memory B cell responses by day 30 (mean, 0.10% of total circulating IgG memory B cells; range, 0.037%-0.28%), which persisted to day 90 (mean, 0.07%; range, 0.003%-0.27%). In contrast, circulating CTB-specific IgG antibody-secreting cells and serum vibriocidal and anti-CTB antibody responses peaked on day 7 and declined to undetectable or significantly lower levels by day 90., Conclusions: CTB-specific IgG memory B cell responses are detectable in the circulation at least 3 months after V. cholerae O1 infection and remain measurable even after serum antibody titers have declined to undetectable or considerably lower levels. This suggests that antigen-specific memory B cells may be an important long-term marker of the immune response to cholera.

Background: We estimated the effects of rainfall and temperature on the number of non-cholera diarrhoea cases and identified population factors potentially affecting vulnerability to the effect of the climate factors in Dhaka, Bangladesh., Methods: Weekly rainfall, temperature and number of hospital visits for non-cholera diarrhoea were analysed by time-series regression. A Poisson regression model was used to model the relationships controlling for seasonally varying factors other than the weather variables. Modifications of weather effects were investigated by fitting the models separately to incidence series according to their characteristics (sex, age, socio-economic, hygiene and sanitation status)., Results: The number of non-cholera diarrhoea cases per week increased by 5.1% (95% CI: 3.3-6.8) for every 10 mm increase above the threshold of 52 mm of average rainfall over lags 0-8 weeks. The number of cases also increased by 3.9% (95% CI: 0.6-7.2) for every 10 mm decrease below the same threshold of rainfall. Ambient temperature was also positively associated with the number of non-cholera diarrhoea cases. There was no evidence for the modification of both 'high and low rainfall' effects by individual characteristics, while the effect of temperature was higher amongst those individuals at a lower educational attainment and unsanitary toilet users., Conclusions: The number of non-cholera diarrhoea cases increased both above and below a threshold level with high and low rainfall in the preceding weeks. The number of cases also increased with higher temperature, particularly in those individuals at a lower socio-economic and sanitation status.

The serum vibriocidal antibody is the only recognized predictor of protection from cholera, but no seroepidemiological data have been gathered since the emergence of Vibrio cholerae O139. We assessed the association between the vibriocidal antibody titer and protection from cholera in an endemic setting. Although a higher baseline vibriocidal titer correlated with protection from V. cholerae O1, infection still developed in some contacts with very high titers. No association between baseline vibriocidal titer and protection from V. cholerae O139 infection was found. Our findings suggest that the vibriocidal antibody is an incomplete predictor of protection from V. cholerae infection.