Clinical severity of enteric viruses detected using a quantitative molecular assay compared to conventional assays in the Global Enteric Multicenter Study.

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.

A Description of the Statistical Methods for the Vaccine Impact on Diarrhea in Africa (VIDA) Study.

BACKGROUND: Diarrheal diseases remain a health threat to children in low- and middle-income countries. The Vaccine Impact on Diarrhea in Africa (VIDA) study was a 36-month, prospective, matched case-control study designed to estimate the etiology, incidence, and adverse clinical consequences of moderate-to-severe diarrhea (MSD) in children aged 0-59 months. VIDA was conducted following rotavirus vaccine introduction at 3 censused sites in sub-Saharan Africa that participated in the Global Enteric Multicenter Study (GEMS) ∼10 years earlier. We describe the study design and statistical methods of VIDA and where they differ from GEMS. METHODS: We aimed to enroll 8-9 MSD cases every 2 weeks from sentinel health centers in 3 age strata (0-11, 12-23, 24-59 months) and 1 to 3 controls matched by age, sex, date of case enrollment, and village. Clinical, epidemiological, and anthropometric data were collected at enrollment and ∼60 days later. A stool specimen collected at enrollment was analyzed by both conventional methods and quantitative PCR for enteric pathogens. For the matched case-control study, we estimated the population-based, pathogen-specific attributable fraction (AF) and attributable incidence adjusted for age, site, and other pathogens, and identified episodes attributable to a specific pathogen for additional analyses. A prospective cohort study nested within the original matched case-control study allowed assessment of (1) the association between potential risk factors and outcomes other than MSD status and (2) the impact of MSD on linear growth. CONCLUSIONS: GEMS and VIDA together comprise the largest and most comprehensive assessment of MSD conducted to date in sub-Saharan Africa populations at highest risk for morbidity and mortality from diarrhea. The statistical methods used in VIDA have endeavored to maximize the use of available data to produce more robust estimates of the pathogen-specific disease burden that might be prevented by effective interventions.

Drivers of Decline in Diarrhea Mortality Between GEMS and VIDA Studies.

BACKGROUND: Statistical modeling suggests that decreasing diarrhea-associated mortality rates in recent decades are largely attributed to improved case management, rotavirus vaccine, and economic development. METHODS: We examined data collected in 2 multisite population-based diarrhea case-control studies, both conducted in The Gambia, Kenya, and Mali: the Global Enteric Multicenter Study (GEMS; 2008-2011) and Vaccine Impact on Diarrhea in Africa (VIDA; 2015-2018). Population-level diarrhea mortality and risk factor prevalence, estimated using these study data, were used to calculate the attribution of risk factors and interventions for diarrhea mortality using a counterfactual framework. We performed a decomposition of the effects of the changes in exposure to each risk factor between GEMS and VIDA on diarrhea mortality for each site. RESULTS: Diarrhea mortality among children under 5 in our African sites decreased by 65.3% (95% confidence interval [CI]: -80.0%, -45.0%) from GEMS to VIDA. Kenya and Mali had large relative declines in diarrhea mortality between the 2 periods with 85.9% (95% CI: -95.1%, -71.5%) and 78.0% (95% CI: -96.0%, 36.3%) reductions, respectively. Among the risk factors considered, the largest declines in diarrhea mortality between the 2 study periods were attributed to reduction in childhood wasting (27.2%; 95% CI: -39.3%, -16.8%) and an increased rotavirus vaccine coverage (23.1%; 95% CI: -28.4%, -19.4%), zinc for diarrhea treatment (12.1%; 95% CI: -16.0%, -8.9%), and oral rehydration salts (ORS) for diarrhea treatment (10.2%). CONCLUSIONS: The VIDA study sites demonstrated exceptional reduction in diarrhea mortality over the last decade. Site-specific differences highlight an opportunity for implementation science in collaboration with policymakers to improve the equitable coverage of these interventions globally.

Stunting Following Moderate-to-Severe Diarrhea Among Children Aged <5 Years in Africa Before and After Rotavirus Vaccine Introduction: A Comparison of the Global Enteric Multicenter Study and the Vaccine Impact on Diarrhea in Africa (VIDA) Study.

BACKGROUND: Studies conducted before rotavirus vaccine introduction found that moderate-to-severe diarrhea (MSD) in children aged <5 years was associated with stunting at follow-up. It is unknown whether the reduction in rotavirus-associated MSD following vaccine introduction decreased the risk of stunting. METHODS: The Global Enteric Multicenter Study (GEMS) and the Vaccine Impact on Diarrhea in Africa (VIDA) study, two comparable matched case-control studies, were conducted during 2007-2011 and 2015-2018, respectively. We analyzed data from 3 African sites where rotavirus vaccine was introduced after GEMS and before starting VIDA. Children with acute MSD (<7 days onset) were enrolled from a health center and children without MSD (diarrhea-free for ≥7 days) were enrolled at home within 14 days of the index MSD case. The odds of being stunted at a follow-up visit 2-3 months after enrollment for an episode of MSD was compared between GEMS and VIDA using mixed-effects logistic regression models controlling for age, sex, study site, and socioeconomic status. RESULTS: We analyzed data from 8808 children from GEMS and 10 579 from VIDA. Among those who were not stunted at enrollment in GEMS, 8.6% with MSD and 6.4% without MSD became stunted during the follow-up period. In VIDA, 8.0% with MSD and 5.5% children without MSD developed stunting. An episode of MSD was associated with higher odds of being stunted at follow-up compared with children without MSD in both studies (adjusted odds ratio [aOR], 1.31; 95% confidence interval [CI]: 1.04-1.64 in GEMS and aOR, 1.30; 95% CI: 1.04-1.61 in VIDA). However, the magnitude of association was not significantly different between GEMS and VIDA (P = .965). CONCLUSIONS: The association of MSD with subsequent stunting among children aged <5 years in sub-Saharan Africa did not change after rotavirus vaccine introduction. Focused strategies are needed for prevention of specific diarrheal pathogens that cause childhood stunting.

Management of Diarrhea in Young Children in Sub-Saharan Africa: Adherence to World Health Organization Recommendations During the Global Enteric Multisite Study (2007-2011) and the Vaccine Impact of Diarrhea in Africa (VIDA) Study (2015-2018).

BACKGROUND: Reducing diarrhea-related morbidity and mortality is a global priority, particularly in low-resource settings. We assessed adherence to diarrhea case management indicators in the Global Enteric Multisite Study (GEMS) and Vaccine Impact of Diarrhea in Africa (VIDA) study. METHODS: GEMS (2007-2010) and VIDA (2015-2018) were age-stratified case-control studies of moderate-to-severe diarrhea (MSD) in children aged <5 years. In this case-only analysis, we included children enrolled in The Gambia, Kenya, and Mali. A case with no dehydration received adherent care at home if they were offered more than usual fluids and at least the same as usual to eat. Children with diarrhea and some dehydration are to receive oral rehydration salts (ORS) in the facility. The recommendation for severe dehydration is to receive ORS and intravenous fluids in the facility. Adherent care in the facility included a zinc prescription independent of dehydration severity. RESULTS: For home-based management of children with MSD and no signs of dehydration, 16.6% in GEMS and 15.6% in VIDA were adherent to guidelines. Adherence to guidelines in the facility was likewise low during GEMS (some dehydration, 18.5%; severe dehydration, 5.5%). The adherence to facility-based rehydration and zinc guidelines improved during VIDA to 37.9% of those with some dehydration and 8.0% of children with severe dehydration. CONCLUSIONS: At research sites in The Gambia, Kenya, and Mali, suboptimal adherence to diarrhea case management guidelines for children aged <5 years was observed. Opportunities exist for improvement in case management for children with diarrhea in low-resource settings.

Moderate-to-Severe Diarrhea and Stunting Among Children Younger Than 5 Years: Findings From the Vaccine Impact on Diarrhea in Africa (VIDA) Study.

BACKGROUND: Stunting affects >20% of children <5 years old worldwide and disproportionately impacts underserved communities. The Vaccine Impact on Diarrhea in Africa (VIDA) Study examined the association between an episode of moderate-to-severe diarrhea (MSD) and the risk of subsequent stunting in children <5 years living in 3 sub-Saharan African countries. METHODS: In this prospective, matched, case-control study among children <5 years, data were collected over 36 months from 2 groups. "Children with MSD" visited a health center within 7 days of illness onset experiencing ≥3 loose stools/day plus sunken eyes, poor skin turgor, dysentery, intravenous rehydration, or hospitalization. "Children without MSD" were enrolled from the community within 14 days of the index MSD child; they were diarrhea-free during the previous 7 days and were matched to the index case by age, sex, and residence. Using generalized linear mixed-effects models, we estimated the effect of an MSD episode on odds of being stunted, defined as height-for-age z-scores <-2, at a follow-up visit 2-3 months post-enrollment. RESULTS: The proportion of stunting at enrollment was similar when 4603 children with MSD and 5976 children without MSD were compared (21.8% vs 21.3%; P = .504). Among children not stunted at enrollment, those with MSD had 30% higher odds of being stunted at follow-up than children without MSD after controlling for age, sex, study site, and socioeconomic status (adjusted OR: 1.30; 95% CI: 1.05-1.62: P = .018). CONCLUSIONS: Children <5 years in sub-Saharan Africa without stunting experienced an increased likelihood of stunting during 2-3 months following an episode of MSD. Strategies for control of early childhood diarrhea should be integrated into programs intended to reduce childhood stunting.

Clinical and Epidemiologic Features of Cryptosporidium-Associated Diarrheal Disease Among Young Children Living in Sub-Saharan Africa: The Vaccine Impact on Diarrhea in Africa (VIDA) Study.

BACKGROUND: As part of the Vaccine Impact on Diarrhea in Africa (VIDA) Study, we examined the prevalence, clinical presentation, and seasonality of Cryptosporidium in children to understand its relative burden after the introduction of rotavirus vaccine. METHODS: VIDA was a 3-year, age-stratified, matched case-control study of medically attended acute moderate-to-severe diarrhea (MSD) in children aged 0-59 months residing in censused populations at sites in Kenya, Mali, and The Gambia. Clinical and epidemiologic data were collected at enrollment, and a stool sample was tested for enteropathogens by quantitative PCR. An algorithm was created based on the organism's cycle threshold (Ct) and association with MSD to identify the subset of Cryptosporidium PCR-positive (Ct <35) cases most likely to be attributed to MSD. Clinical outcomes were assessed at 2-3 months after enrollment. RESULTS: One thousand one hundred six (22.9%) cases of MSD and 873 controls (18.1%) were PCR positive for Cryptosporidium; 465 cases (42.0%) were considered attributable to Cryptosporidium, mostly among children 6-23 months. Cryptosporidium infections peaked in The Gambia and Mali during the rainy season, while in Kenya they did not have clear seasonality. Compared with cases with watery MSD who had a negative PCR for Cryptosporidium, cases with watery MSD attributed to Cryptosporidium were less frequently dehydrated but appeared more severely ill using a modified Vesikari scale (38.1% vs 27.0%; P < 0.001), likely due to higher rates of hospitalization and intravenous fluid administration, higher prevalence of being wasted or very thin very thin (23.4% vs 14.7%; P < 0.001), and having severe acute malnutrition (midupper arm circumference <115 mm, 7.7% vs 2.5%; P < 0.001). On follow-up, Cryptosporidium-attributed cases had more prolonged and persistent episodes (43.2% vs 32.7%; P <0 .001) and linear growth faltering (change in height-for-age z score between enrollment and follow-up: -0.29 vs -0.17; P < 0.001). CONCLUSIONS: The burden of Cryptosporidium remains high among young children in sub-Saharan Africa. Its propensity to cause illness and further impact children longer term by compromising nutritional status early in life calls for special attention to enable appropriate management of clinical and nutritional consequences.

Antibiotic-Prescribing Practices for Management of Childhood Diarrhea in 3 Sub-Saharan African Countries: Findings From the Vaccine Impact on Diarrhea in Africa (VIDA) Study, 2015-2018.

BACKGROUND: Despite antibiotic prescription being recommended for dysentery and suspected cholera only, diarrhea still triggers unwarranted antibiotic prescription. We evaluated antibiotic-prescribing practices and their predictors among children aged 2-59 months in the Vaccine Impact on Diarrhea in Africa (VIDA) Study performed in The Gambia, Mali, and Kenya. METHODS: VIDA was a prospective case-control study (May 2015-July 2018) among children presenting for care with moderate-to-severe diarrhea (MSD). We defined inappropriate antibiotic use as prescription or use of antibiotics when not indicated by World Health Organization (WHO) guidelines. We used logistic regression to assess factors associated with antibiotic prescription for MSD cases who had no indication for an antibiotic, at each site. RESULTS: VIDA enrolled 4840 cases. Among 1757 (36.3%) who had no apparent indication for antibiotic treatment, 1358 (77.3%) were prescribed antibiotics. In The Gambia, children who presented with a cough (adjusted odds ratio [aOR]: 2.05; 95% confidence interval [95% CI]: 1.21-3.48) were more likely to be prescribed an antibiotic. In Mali, those who presented with dry mouth (aOR: 3.16; 95% CI: 1.02-9.73) were more likely to be prescribed antibiotics. In Kenya, those who presented with a cough (aOR: 2.18; 95% CI: 1.01-4.70), decreased skin turgor (aOR: 2.06; 95% CI: 1.02-4.16), and were very thirsty (aOR: 4.15; 95% CI: 1.78-9.68) were more likely to be prescribed antibiotics. CONCLUSIONS: Antibiotic prescription was associated with signs and symptoms inconsistent with WHO guidelines, suggesting the need for antibiotic stewardship and clinician awareness of diarrhea case-management recommendations in these settings.

Prevalence of Salmonella in Stool During the Vaccine Impact on Diarrhea in Africa (VIDA) Study, 2015-2018.

BACKGROUND: Non-typhoidal Salmonella (NTS) is a common cause of gastroenteritis in young children, with limited data on NTS serovars and antimicrobial resistance in Africa. METHODS: We determined the prevalence of Salmonella spp. and frequency of antimicrobial resistance among serovars identified in stools of 0-59 month-old children with moderate-to-severe diarrhea (MSD) and controls enrolled in the Vaccine Impact on Diarrhea in Africa (VIDA) Study in The Gambia, Mali, and Kenya in 2015-2018, and compared with data from the Global Enteric Multicenter Study (GEMS; 2007-2010) and the GEMS-1A study (2011). Salmonella spp. was detected by quantitative real-time PCR (qPCR) and culture-based methods. Identification of serovars was determined by microbiological methods. RESULTS: By qPCR, the prevalence of Salmonella spp. among MSD cases was 4.0%, 1.6%, and 1.9% and among controls was 4.6%, 2.4%, and 1.6% in The Gambia, Mali, and Kenya, respectively, during VIDA. We observed year-to-year variation in serovar distribution and variation between sites. In Kenya, Salmonella enterica serovar Typhimurium decreased (78.1% to 23.1%; P < .001) among cases and controls from 2007 to 2018, whereas serogroup O:8 increased (8.7% to 38.5%; P = .04). In The Gambia, serogroup O:7 decreased from 2007 to 2018 (36.3% to 0%; P = .001) but S. enterica serovar Enteritidis increased during VIDA (2015 to 2018; 5.9% to 50%; P = .002). Only 4 Salmonella spp. were isolated in Mali during all 3 studies. Multidrug resistance was 33.9% in Kenya and 0.8% in The Gambia across all 3 studies. Ceftriaxone resistance was only observed in Kenya (2.3%); NTS isolates were susceptible to ciprofloxacin at all sites. CONCLUSIONS: Understanding variability in serovar distribution will be important for the future deployment of vaccines against salmonellosis in Africa.

Epidemiology of Enteroaggregative, Enteropathogenic, and Shiga Toxin-Producing Escherichia coli Among Children Aged <5 Years in 3 Countries in Africa, 2015-2018: Vaccine Impact on Diarrhea in Africa (VIDA) Study.

BACKGROUND: To address knowledge gaps regarding diarrheagenic Escherichia coli (DEC) in Africa, we assessed the clinical and epidemiological features of enteroaggregative E. coli (EAEC), enteropathogenic E. coli (EPEC), and Shiga toxin-producing E. coli (STEC) positive children with moderate-to-severe diarrhea (MSD) in Mali, The Gambia, and Kenya. METHODS: Between May 2015 and July 2018, children aged 0-59 months with medically attended MSD and matched controls without diarrhea were enrolled. Stools were tested conventionally using culture and multiplex polymerase chain reaction (PCR), and by quantitative PCR (qPCR). We assessed DEC detection by site, age, clinical characteristics, and enteric coinfection. RESULTS: Among 4840 children with MSD and 6213 matched controls enrolled, 4836 cases and 1 control per case were tested using qPCR. Of the DEC detected with TAC, 61.1% were EAEC, 25.3% atypical EPEC (aEPEC), 22.4% typical EPEC (tEPEC), and 7.2% STEC. Detection was higher in controls than in MSD cases for EAEC (63.9% vs 58.3%, P < .01), aEPEC (27.3% vs 23.3%, P < .01), and STEC (9.3% vs 5.1%, P < .01). EAEC and tEPEC were more frequent in children aged <23 months, aEPEC was similar across age strata, and STEC increased with age. No association between nutritional status at follow-up and DEC pathotypes was found. DEC coinfection with Shigella/enteroinvasive E. coli was more common among cases (P < .01). CONCLUSIONS: No significant association was detected between EAEC, tEPEC, aEPEC, or STEC and MSD using either conventional assay or TAC. Genomic analysis may provide a better definition of the virulence factors associated with diarrheal disease.

Shigella in Africa: New Insights From the Vaccine Impact on Diarrhea in Africa (VIDA) Study.

BACKGROUND: We evaluated the burden of Shigella spp from children aged 0-59 months with medically attended moderate-to-severe diarrhea and matched controls at sites in Mali, The Gambia, and Kenya participating in the Vaccine Impact on Diarrhea in Africa (VIDA) study from 2015 to 2018. METHODS: Shigella spp were identified using coprocultures and serotyping in addition to quantitative polymerase chain reaction (qPCR). Episode-specific attributable fractions (AFe) for Shigella were calculated using Shigella DNA quantity; cases with AFe ≥0.5 were considered to have shigellosis. RESULTS: The prevalence of Shigella was determined to be 359 of 4840 (7.4%) cases and 83 of 6213 (1.3%) controls by culture, and 1641 of 4836 (33.9%) cases and 1084 of 4846 (22.4%) controls by qPCR (cycle threshold <35); shigellosis was higher in The Gambia (30.8%) than in Mali (9.3%) and Kenya (18.7%). Bloody diarrhea attributed to Shigella was more common in 24- to 59-month-old children (50.1%) than 0- to 11-month-old infants (39.5%). The Shigella flexneri serogroup predominated among cases (67.6% of isolates), followed by Shigella sonnei (18.2%), Shigella boydii (11.8%), and Shigella dysenteriae (2.3%). The most frequent S. flexneri serotypes were 2a (40.6%), 1b (18.8%), 6 (17.5%), 3a (9.0%), and 4a (5.1%). Drug-specific resistance among 353 (98.3%) Shigella cases with AMR data was as follows: trimethoprim-sulfamethoxazole (94.9%), ampicillin (48.4%), nalidixic acid (1.7%), ceftriaxone (0.3%), azithromycin (0.3%), and ciprofloxacin (0.0%). CONCLUSIONS: A high prevalence of shigellosis continues in sub-Saharan Africa. Strains are highly resistant to commonly used antibiotics while remaining susceptible to ciprofloxacin, ceftriaxone, and azithromycin.

Transmission of Nipah Virus - 14 Years of Investigations in Bangladesh.

BACKGROUND: Nipah virus is a highly virulent zoonotic pathogen that can be transmitted between humans. Understanding the dynamics of person-to-person transmission is key to designing effective interventions. METHODS: We used data from all Nipah virus cases identified during outbreak investigations in Bangladesh from April 2001 through April 2014 to investigate case-patient characteristics associated with onward transmission and factors associated with the risk of infection among patient contacts. RESULTS: Of 248 Nipah virus cases identified, 82 were caused by person-to-person transmission, corresponding to a reproduction number (i.e., the average number of secondary cases per case patient) of 0.33 (95% confidence interval [CI], 0.19 to 0.59). The predicted reproduction number increased with the case patient's age and was highest among patients 45 years of age or older who had difficulty breathing (1.1; 95% CI, 0.4 to 3.2). Case patients who did not have difficulty breathing infected 0.05 times as many contacts (95% CI, 0.01 to 0.3) as other case patients did. Serologic testing of 1863 asymptomatic contacts revealed no infections. Spouses of case patients were more often infected (8 of 56 [14%]) than other close family members (7 of 547 [1.3%]) or other contacts (18 of 1996 [0.9%]). The risk of infection increased with increased duration of exposure of the contacts (adjusted odds ratio for exposure of >48 hours vs. ≤1 hour, 13; 95% CI, 2.6 to 62) and with exposure to body fluids (adjusted odds ratio, 4.3; 95% CI, 1.6 to 11). CONCLUSIONS: Increasing age and respiratory symptoms were indicators of infectivity of Nipah virus. Interventions to control person-to-person transmission should aim to reduce exposure to body fluids. (Funded by the National Institutes of Health and others.).

Characteristics of Salmonella Recovered From Stools of Children Enrolled in the Global Enteric Multicenter Study.

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 Clinical Presentation of Culture-positive and Culture-negative, Quantitative Polymerase Chain Reaction (qPCR)-Attributable Shigellosis in the Global Enteric Multicenter Study and Derivation of a Shigella Severity Score: Implications for Pediatric Shigella Vaccine Trials.

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.

Changing Contact Patterns Over Disease Progression: Nipah Virus as a Case Study.

Contact patterns play a key role in disease transmission, and variation in contacts during the course of illness can influence transmission, particularly when accompanied by changes in host infectiousness. We used surveys among 1642 contacts of 94 Nipah virus case patients in Bangladesh to determine how contact patterns (physical and with bodily fluids) changed as disease progressed in severity. The number of contacts increased with severity and, for case patients who died, peaked on the day of death. Given transmission has only been observed among fatal cases of Nipah virus infection, our findings suggest that changes in contact patterns during illness contribute to risk of infection.

Determinants of linear growth faltering among children with moderate-to-severe diarrhea in the Global Enteric Multicenter Study.

BACKGROUND: Moderate-to-severe diarrhea (MSD) in the first 2 years of life can impair linear growth. We sought to determine risk factors for linear growth faltering and to build a clinical prediction tool to identify children most likely to experience growth faltering following an episode of MSD. METHODS: Using data from the Global Enteric Multicenter Study of children 0-23 months old presenting with MSD in Africa and Asia, we performed log-binomial regression to determine clinical and sociodemographic factors associated with severe linear growth faltering (loss of ≥ 0.5 length-for-age z-score [LAZ]). Linear regression was used to estimate associations with ΔLAZ. A clinical prediction tool was developed using backward elimination of potential variables, and Akaike Information Criterion to select the best fit model. RESULTS: Of the 5902 included children, mean age was 10 months and 43.2% were female. Over the 50-90-day follow-up period, 24.2% of children had severe linear growth faltering and the mean ΔLAZ over follow-up was - 0.17 (standard deviation [SD] 0.54). After adjustment for age, baseline LAZ, and site, several factors were associated with decline in LAZ: young age, acute malnutrition, hospitalization at presentation, non-dysenteric diarrhea, unimproved sanitation, lower wealth, fever, co-morbidity, or an IMCI danger sign. Compared to children 12-23 months old, those 0-6 months were more likely to experience severe linear growth faltering (adjusted prevalence ratio [aPR] 1.97 [95% CI 1.70, 2.28]), as were children 6-12 months of age (aPR 1.72 [95% CI 1.51, 1.95]). A prediction model that included age, wasting, stunting, presentation with fever, and presentation with an IMCI danger sign had an area under the ROC (AUC) of 0.67 (95% CI 0.64, 0.69). Risk scores ranged from 0 to 37, and a cut-off of 21 maximized sensitivity (60.7%) and specificity (63.5%). CONCLUSION: Younger age, acute malnutrition, MSD severity, and sociodemographic factors were associated with short-term linear growth deterioration following MSD. Data routinely obtained at MSD may be useful to predict children at risk for growth deterioration who would benefit from interventions.

Mild Respiratory Illness Among Young Children Caused by Highly Pathogenic Avian Influenza A (H5N1) Virus Infection in Dhaka, Bangladesh, 2011.

Background: In March 2011, a multidisciplinary team investigated 2 human cases of highly pathogenic avian influenza A(H5N1) virus infection, detected through population-based active surveillance for influenza in Bangladesh, to assess transmission and contain further spread. Methods: We collected clinical and exposure history of the case patients and monitored persons coming within 1 m of a case patient during their infectious period. Nasopharyngeal wash specimens from case patients and contacts were tested with real-time reverse-transcription polymerase chain reaction, and virus culture and isolates were characterized. Serum samples were tested with microneutralization and hemagglutination inhibition assays. We tested poultry, wild bird, and environmental samples from case patient households and surrounding areas for influenza viruses. Results: Two previously healthy case patients, aged 13 and 31 months, had influenzalike illness and fully recovered. They had contact with poultry 7 and 10 days before illness onset, respectively. None of their 57 contacts were subsequently ill. Clade 2.2.2.1 highly pathogenic avian influenza H5N1 viruses were isolated from the case patients and from chicken fecal samples collected at the live bird markets near the patients' dwellings. Conclusion: Identification of H5N1 cases through population-based surveillance suggests possible additional undetected cases throughout Bangladesh and highlights the importance of surveillance for mild respiratory illness among populations frequently exposed to infected poultry.

Convergence of Humans, Bats, Trees, and Culture in Nipah Virus Transmission, Bangladesh.

Preventing emergence of new zoonotic viruses depends on understanding determinants for human risk. Nipah virus (NiV) is a lethal zoonotic pathogen that has spilled over from bats into human populations, with limited person-to-person transmission. We examined ecologic and human behavioral drivers of geographic variation for risk of NiV infection in Bangladesh. We visited 60 villages during 2011-2013 where cases of infection with NiV were identified and 147 control villages. We compared case villages with control villages for most likely drivers for risk of infection, including number of bats, persons, and date palm sap trees, and human date palm sap consumption behavior. Case villages were similar to control villages in many ways, including number of bats, persons, and date palm sap trees, but had a higher proportion of households in which someone drank sap. Reducing human consumption of sap could reduce virus transmission and risk for emergence of a more highly transmissible NiV strain.

Evaluating Hospital-Based Surveillance for Outbreak Detection in Bangladesh: Analysis of Healthcare Utilization Data.

BACKGROUND: The International Health Regulations outline core requirements to ensure the detection of public health threats of international concern. Assessing the capacity of surveillance systems to detect these threats is crucial for evaluating a country's ability to meet these requirements. METHODS AND FINDINGS: We propose a framework to evaluate the sensitivity and representativeness of hospital-based surveillance and apply it to severe neurological infectious diseases and fatal respiratory infectious diseases in Bangladesh. We identified cases in selected communities within surveillance hospital catchment areas using key informant and house-to-house surveys and ascertained where cases had sought care. We estimated the probability of surveillance detecting different sized outbreaks by distance from the surveillance hospital and compared characteristics of cases identified in the community and cases attending surveillance hospitals. We estimated that surveillance detected 26% (95% CI 18%-33%) of severe neurological disease cases and 18% (95% CI 16%-21%) of fatal respiratory disease cases residing at 10 km distance from a surveillance hospital. Detection probabilities decreased markedly with distance. The probability of detecting small outbreaks (three cases) dropped below 50% at distances greater than 26 km for severe neurological disease and at distances greater than 7 km for fatal respiratory disease. Characteristics of cases attending surveillance hospitals were largely representative of all cases; however, neurological disease cases aged <5 y or from the lowest socioeconomic group and fatal respiratory disease cases aged ≥60 y were underrepresented. Our estimates of outbreak detection rely on suspected cases that attend a surveillance hospital receiving laboratory confirmation of disease and being reported to the surveillance system. The extent to which this occurs will depend on disease characteristics (e.g., severity and symptom specificity) and surveillance resources. CONCLUSION: We present a new approach to evaluating the sensitivity and representativeness of hospital-based surveillance, making it possible to predict its ability to detect emerging threats.

Use of quantitative molecular diagnostic methods to identify causes of diarrhoea in children: a reanalysis of the GEMS case-control study.

BACKGROUND: Diarrhoea is the second leading cause of mortality in children worldwide, but establishing the cause can be complicated by diverse diagnostic approaches and varying test characteristics. We used quantitative molecular diagnostic methods to reassess causes of diarrhoea in the Global Enteric Multicenter Study (GEMS). METHODS: GEMS was a study of moderate to severe diarrhoea in children younger than 5 years in Africa and Asia. We used quantitative real-time PCR (qPCR) to test for 32 enteropathogens in stool samples from cases and matched asymptomatic controls from GEMS, and compared pathogen-specific attributable incidences with those found with the original GEMS microbiological methods, including culture, EIA, and reverse-transcriptase PCR. We calculated revised pathogen-specific burdens of disease and assessed causes in individual children. FINDINGS: We analysed 5304 sample pairs. For most pathogens, incidence was greater with qPCR than with the original methods, particularly for adenovirus 40/41 (around five times), Shigella spp or enteroinvasive Escherichia coli (EIEC) and Campylobactor jejuni o C coli (around two times), and heat-stable enterotoxin-producing E coli ([ST-ETEC] around 1·5 times). The six most attributable pathogens became, in descending order, Shigella spp, rotavirus, adenovirus 40/41, ST-ETEC, Cryptosporidium spp, and Campylobacter spp. Pathogen-attributable diarrhoeal burden was 89·3% (95% CI 83·2-96·0) at the population level, compared with 51·5% (48·0-55·0) in the original GEMS analysis. The top six pathogens accounted for 77·8% (74·6-80·9) of all attributable diarrhoea. With use of model-derived quantitative cutoffs to assess individual diarrhoeal cases, 2254 (42·5%) of 5304 cases had one diarrhoea-associated pathogen detected and 2063 (38·9%) had two or more, with Shigella spp and rotavirus being the pathogens most strongly associated with diarrhoea in children with mixed infections. INTERPRETATION: A quantitative molecular diagnostic approach improved population-level and case-level characterisation of the causes of diarrhoea and indicated a high burden of disease associated with six pathogens, for which targeted treatment should be prioritised. FUNDING: Bill & Melinda Gates Foundation.