Identification of Enteric Pathogen Reservoirs and Transmission Pathways Associated with Short Childhood Stature in the Kolkata Indian Site of the Global Enteric Multicenter Study.

Age-stratified path analyses modeled associations between enteric pathogen reservoirs, transmission pathways and height-for-age z-scores (HAZ) to identify determinants of childhood growth in the Kolkata, India site of the Global Enteric Multicenter Study (GEMS). Models tested direct associations of potential pathogen reservoirs with HAZ at 60-day follow-up in separate moderate and severe diarrhea (MSD) case and control cohorts or indirectly when mediated by enteric infections. In the MSD cohort, rotavirus and typical EPEC (tEPEC) infections among children 0-11 months of age and ST-ETEC infections among children 12-23 months of age were associated with lower HAZ. Handwashing after defecating and before cooking reduced impaired growth through reductions in rotavirus and tEPEC infections. Water storage increased rotavirus and ST-ETEC infection risks, resulting in increased impaired growth, but was reduced with reported child feces disposal. The GII norovirus variant was inversely associated with HAZ among children 12-59 months of age in the control cohort. Reported handwashing before the handling of children reduced GII infections and impaired growth. Boiling water and the disposal of children's feces mediated by stored water were positively associated with HAZ. The targeting of pathogen-specific reservoirs and transmission pathways may more effectively improve childhood linear growth in South Asian urban communities.

Adverse outcomes in patients hospitalized with pneumonia at age 60 or more: A prospective multi-centric hospital-based study in India.

BACKGROUND: Limited data exists regarding risk factors for adverse outcomes in older adults hospitalized with Community-Acquired Pneumonia (CAP) in low- and middle-income countries such as India. This multisite study aimed to assess outcomes and associated risk factors among adults aged ≥60 years hospitalized with pneumonia. METHODS: Between December 2018 and March 2020, we enrolled ≥60-year-old adults admitted within 48 hours for CAP treatment across 16 public and private facilities in four sites. Clinical data and nasal/oropharyngeal specimens were collected by trained nurses and tested for influenza, respiratory syncytial virus (RSV), and other respiratory viruses (ORV) using the qPCR. Participants were evaluated regularly until discharge, as well as on the 7th and 30th days post-discharge. Outcomes included ICU admission and in-hospital or 30-day post-discharge mortality. A hierarchical framework for multivariable logistic regression and Cox proportional hazard models identified risk factors (e.g., demographics, clinical features, etiologic agents) associated with critical care or death. FINDINGS: Of 1,090 CAP patients, the median age was 69 years; 38.4% were female. Influenza viruses were detected in 12.3%, RSV in 2.2%, and ORV in 6.3% of participants. Critical care was required for 39.4%, with 9.9% in-hospital mortality and 5% 30-day post-discharge mortality. Only 41% of influenza CAP patients received antiviral treatment. Admission factors independently associated with ICU admission included respiratory rate >30/min, blood urea nitrogen>19mg/dl, altered sensorium, anemia, oxygen saturation <90%, prior cardiovascular diseases, chronic respiratory diseases, and private hospital admission. Diabetes, anemia, low oxygen saturation at admission, ICU admission, and mechanical ventilation were associated with 30-day mortality. CONCLUSION: High ICU admission and 30-day mortality rates were observed among older adults with pneumonia, with a significant proportion linked to influenza and RSV infections. Comprehensive guidelines for CAP prevention and management in older adults are needed, especially with the co-circulation of SARS-CoV-2.

Cost of acute respiratory illness episode and its determinants among community-dwelling older adults: a four-site cohort study from India.

Introduction: Advocacy for the provision of public health resources, including vaccine for the prevention of acute respiratory illnesses (ARIs) among older adults in India, needs evidence on costs and benefits. Using a cohort of community-dwelling adults aged 60 years and older in India, we estimated the cost of ARI episode and its determinants. Methods: We enrolled 6016 participants in Ballabgarh, Chennai, Kolkata and Pune from July 2018 to March 2020. They were followed up weekly to identify ARI and classified them as acute upper respiratory illness (AURI) or pneumonia based on clinical features based on British Thoracic Society guidelines. All pneumonia and 20% of AURI cases were asked about the cost incurred on medical consultation, investigation, medications, transportation, food and lodging. The cost of services at public facilities was supplemented by WHO-Choosing Interventions that are Cost-Effective(CHOICE) estimates for 2019. Indirect costs incurred by the affected participant and their caregivers were estimated using human capital approach. We used generalised linear model with log link and gamma family to identify the average marginal effect of key determinants of the total cost of ARI. Results: We included 2648 AURI and 1081 pneumonia episodes. Only 47% (range 36%-60%) of the participants with pneumonia sought care. The mean cost of AURI episode was US$13.9, while that of pneumonia episode was US$25.6, with indirect costs comprising three-fourths of the total. The cost was higher among older men by US$3.4 (95% CI: 1.4 to 5.3), those with comorbidities by US$4.3 (95% CI: 2.8 to 5.7) and those who sought care by US$17.2 (95% CI: 15.1 to 19.2) but not by influenza status. The mean per capita annual cost of respiratory illness was US$29.5. Conclusion: Given the high community disease and cost burden of ARI, intensifying public health interventions to prevent and mitigate ARI among this fast-growing older adult population in India is warranted.

Cohort profile: Indian Network of Population-Based Surveillance Platforms for Influenza and Other Respiratory Viruses among the Elderly (INSPIRE).

PURPOSE: We describe here a multicentric community-dwelling cohort of older adults (>60 years of age) established to estimate incidence, study risk factors, healthcare utilisation and economic burden associated with influenza and respiratory syncytial virus (RSV) in India. PARTICIPANTS: The four sites of this cohort are in northern (Ballabgarh), southern (Chennai), eastern (Kolkata) and western (Pune) parts of India. We enrolled 5336 participants across 4220 households and began surveillance in July 2018 for viral respiratory infections with additional participants enrolled annually. Trained field workers collected data about individual-level and household-level risk factors at enrolment and quarterly assessed frailty and grip strength. Trained nurses surveilled weekly to identify acute respiratory infections (ARI) and clinically assessed individuals to diagnose acute lower respiratory infection (ALRI) as per protocol. Nasal and oropharyngeal swabs are collected from all ALRI cases and one-fifth of the other ARI cases for laboratory testing. Cost data of the episode are collected using the WHO approach for estimating the economic burden of seasonal influenza. Handheld tablets with Open Data Kit platform were used for data collection. FINDINGS TO DATE: The attrition of 352 participants due to migration and deaths was offset by enrolling 680 new entrants in the second year. All four sites reported negligible influenza vaccination uptake (0.1%-0.4%), low health insurance coverage (0.4%-22%) and high tobacco use (19%-52%). Ballabgarh had the highest proportion (54.4%) of households in the richest wealth quintile, but reported high solid fuel use (92%). Frailty levels were highest in Kolkata (11.3%) and lowest in Pune (6.8%). The Chennai cohort had highest self-reported morbidity (90.1%). FUTURE PLANS: The findings of this cohort will be used to inform prioritisation of strategies for influenza and RSV control for older adults in India. We also plan to conduct epidemiological studies of SARS-CoV-2 using this platform.

Pathogens Associated With Linear Growth Faltering in Children With Diarrhea and Impact of Antibiotic Treatment: The Global Enteric Multicenter Study.

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.

Diarrhoeal disease and subsequent risk of death in infants and children residing in low-income and middle-income countries: analysis of the GEMS case-control study and 12-month GEMS-1A follow-on study.

BACKGROUND: The Global Enteric Multicenter Study (GEMS) was a 3-year case-control study that measured the burden, aetiology, and consequences of moderate-to-severe diarrhoea (MSD) in children aged 0-59 months. GEMS-1A, a 12-month follow-on study, comprised two parallel case-control studies, one assessing MSD and the other less-severe diarrhoea (LSD). In this report, we analyse the risk of death with each diarrhoea type and the specific pathogens associated with fatal outcomes. METHODS: GEMS was a prospective, age-stratified, matched case-control study done at seven sites in Africa and Asia. Children aged 0-59 months with MSD seeking care at sentinel health centres were recruited along with one to three randomly selected matched community control children without diarrhoea. In the 12-month GEMS-1A follow-on study, children with LSD and matched controls, in addition to children with MSD and matched controls, were recruited at six of the seven sites; only cases of MSD and controls were enrolled at the seventh site. We compared risk of death during the period between enrolment and one follow-up household visit done about 60 days later (range 50-90 days) in children with MSD and LSD and in their respective controls. Approximately 50 pathogens were detected using, as appropriate, classic bacteriology, immunoassays, gel-based PCR and reverse transcriptase PCR, and quantitative real-time PCR (qPCR). Specimens from a subset of GEMS cases and controls were also tested by a TaqMan Array Card that compartmentalised probe-based qPCR for 32 enteropathogens. FINDINGS: 223 (2·0%) of 11 108 children with MSD and 43 (0·3%) of 16 369 matched controls died between study enrolment and the follow-up visit at about 60 days (hazard ratio [HR] 8·16, 95% CI 5·69-11·68, p<0·0001). 12 (0·4%) of 2962 children with LSD and seven (0·2%) of 4074 matched controls died during the follow-up period (HR 2·78, 95% CI 0·95-8·11, p=0·061). Risk of death was lower in children with dysenteric MSD than in children with non-dysenteric MSD (HR 0·20, 95% CI 0·05-0·87, p=0·032), and lower in children with LSD than in those with non-dysenteric MSD (HR 0·29, 0·14-0·59, p=0·0006). In children younger than 24 months with MSD, infection with typical enteropathogenic Escherichia coli, enterotoxigenic E coli encoding heat-stable toxin, enteroaggregative E coli, Shigella spp (non-dysentery cases), Aeromonas spp, Cryptosporidium spp, and Entamoeba histolytica increased risk of death. Of 61 deaths in children aged 12-59 months with non-dysenteric MSD, 31 occurred among 942 children qPCR-positive for Shigella spp and 30 deaths occurred in 1384 qPCR-negative children (HR 2·2, 95% CI 1·2-3·9, p=0·0090), showing that Shigella was strongly associated with increased risk of death. INTERPRETATION: Risk of death is increased following MSD and, to a lesser extent, LSD. Considering there are approximately three times more cases of LSD than MSD in the population, more deaths are expected among children with LSD than in those with MSD. Because the major attributable LSD-associated and MSD-associated pathogens are the same, implementing vaccines and rapid diagnosis and treatment interventions against these major pathogens are rational investments. FUNDING: Bill & Melinda Gates Foundation.

Post-monsoon waterlogging-associated upsurge of cholera cases in and around Kolkata metropolis, 2015.

The Infectious Diseases and Beliaghata General Hospital, Kolkata, India witnessed a sudden increase in admissions of diarrhoea cases during the first 2 weeks of August 2015 following heavy rainfall. This prompted us to investigate the event. Cases were recruited through hospital-based surveillance along with the collection of socio-demographic characteristics and clinical profile using a structured questionnaire. Stool specimens were tested at bacteriological laboratory of the National Institute of Cholera and Enteric Diseases (NICED), Kolkata. Admission of 3003 diarrhoea cases, clearly indicated occurrence of outbreak in Kolkata municipal area as it was more than two standard deviation of the mean number (911; s.d. = 111) of diarrhoea admissions during the same period in previous 7 years. Out of 164 recruited cases, 25% were under-5 children. Organisms were isolated from 80 (49%) stool specimens. Vibrio cholerae O1 was isolated from 50 patients. Twenty-eight patients had this organism as the sole pathogen. Among 14 infants, five had cholera. All V. cholerae O1 isolates were resistant to nalidixic acid, followed by co-trimoxazole (96%), streptomycin (92%), but sensitive to fluroquinolones. We confirmed the occurrence of a cholera outbreak in Kolkata during August 2015 due to V. cholerae O1 infection, where infants were affected.

The incidence, aetiology, and adverse clinical consequences of less severe diarrhoeal episodes among infants and children residing in low-income and middle-income countries: a 12-month case-control study as a follow-on to the Global Enteric Multicenter Study (GEMS).

BACKGROUND: Diarrheal diseases remain a leading cause of illness and death among children younger than 5 years in low-income and middle-income countries. The Global Enteric Multicenter Study (GEMS) has described the incidence, aetiology, and sequelae of medically attended moderate-to-severe diarrhoea (MSD) among children aged 0-59 months residing in censused populations in sub-Saharan Africa and south Asia, where most child deaths occur. To further characterise this disease burden and guide interventions, we extended this study to include children with episodes of less-severe diarrhoea (LSD) seeking care at health centres serving six GEMS sites. METHODS: We report a 1-year, multisite, age-stratified, matched case-control study following on to the GEMS study. Six sites (Bamako, Mali; Manhiça, Mozambique; Basse, The Gambia; Mirzapur, Bangladesh; Kolkata, India; and Bin Qasim Town, Karachi, Pakistan) participated in this study. Children aged 0-59 months at each site who sought care at a sentinel hospital or health centre during a 12-month period were screened for diarrhoea. New (onset after ≥7 diarrhoea-free days) and acute (onset within the previous 7 days) episodes of diarrhoea in children who had sunken eyes, whose skin lost turgor, who received intravenous hydration, who had dysentery, or who were hospitalised were eligible for inclusion as MSD. The remaining new and acute diarrhoea episodes among children who sought care at the same health centres were considered LSD. We aimed to enrol the first eight or nine eligible children with MSD and LSD at each site during each fortnight in three age strata: infants (aged 0-11 months), toddlers (aged 12-23 months), and young children (aged 24-59 months). For each included case of MSD or LSD, we enrolled one to three community control children without diarrhoea during the previous 7 days. From patients and controls we collected clinical and epidemiological data, anthropometric measurements, and faecal samples to identify enteropathogens at enrolment, and we performed a follow-up home visit about 60 days later to ascertain vital status, clinical outcome, and interval growth. Primary outcomes were to characterise, for MSD and LSD, the pathogen-specific attributable risk and population-based incidence values, and to assess the frequency of adverse clinical consequences associated with these two diarrhoeal syndromes. FINDINGS: From Oct 31, 2011, to Nov 14, 2012, we recruited 2368 children with MSD, 3174 with LSD, and one to three randomly selected community control children without diarrhoea matched to cases with MSD (n=3597) or LSD (n=4236). Weighted adjusted population attributable fractions showed that most attributable cases of MSD and LSD were due to rotavirus, Cryptosporidium spp, enterotoxigenic Escherichia coli encoding heat-stable toxin (with or without genes encoding heat-labile enterotoxin), and Shigella spp. The attributable incidence per 100 child-years for LSD versus MSD, by age stratum, for rotavirus was 22·3 versus 5·5 (0-11 months), 9·8 versus 2·9 (12-23 months), and 0·5 versus 0·2 (24-59 months); for Cryptosporidium spp was 3·6 versus 2·3 (0-11 months), 4·3 versus 0·6 (12-23 months), and 0·3 versus 0·1 (24-59 months); for enterotoxigenic E coli encoding heat-stable toxin was 4·2 versus 0·1 (0-11 months), 5·2 versus 0·0 (12-23 months), and 1·1 versus 0·2 (24-59 months); and for Shigella spp was 1·0 versus 1·3 (0-11 months), 3·1 versus 2·4 (12-23 months), and 0·8 versus 0·7 (24-59 months). Participants with both MSD and LSD had significantly more linear growth faltering than controls at follow-up. INTERPRETATION: Inclusion of participants with LSD markedly expands the population of children who experience adverse clinical and nutritional outcomes from acute diarrhoeal diseases. Since MSD and LSD have similar aetiologies, interventions targeting rotavirus, Shigella spp, enterotoxigenic E coli producing heat-stable toxin, and Cryptosporidium spp might substantially reduce the diarrhoeal disease burden and its associated nutritional faltering. FUNDING: Bill & Melinda Gates Foundation.

Colonization factors among enterotoxigenic Escherichia coli isolates from children with moderate-to-severe diarrhea and from matched controls in the Global Enteric Multicenter Study (GEMS).

BACKGROUND: Enterotoxigenic Escherichia coli (ETEC) encoding heat-stable enterotoxin (ST) alone or with heat-labile enterotoxin (LT) cause moderate-to-severe diarrhea (MSD) in developing country children. The Global Enteric Multicenter Study (GEMS) identified ETEC encoding ST among the top four enteropathogens. Since the GEMS objective was to provide evidence to guide development and implementation of enteric vaccines and other interventions to diminish diarrheal disease morbidity and mortality, we examined colonization factor (CF) prevalence among ETEC isolates from children age <5 years with MSD and from matched controls in four African and three Asian sites. We also assessed strength of association of specific CFs with MSD. METHODOLOGY/PRINCIPAL FINDINGS: MSD cases enrolled at healthcare facilities over three years and matched controls were tested in a standardized manner for many enteropathogens. To identify ETEC, three E. coli colonies per child were tested by polymerase chain reaction (PCR) to detect genes encoding LT, ST; confirmed ETEC were examined by PCR for major CFs (Colonization Factor Antigen I [CFA/I] or Coli Surface [CS] antigens CS1-CS6) and minor CFs (CS7, CS12, CS13, CS14, CS17, CS18, CS19, CS20, CS21, CS30). ETEC from 806 cases had a single toxin/CF profile in three tested strains per child. Major CFs, components of multiple ETEC vaccine candidates, were detected in 66.0% of LT/ST and ST-only cases and were associated with MSD versus matched controls by conditional logistic regression (p≤0.006); major CFs detected in only 25.0% of LT-only cases weren't associated with MSD. ETEC encoding exclusively CS14, identified among 19.9% of 291 ST-only and 1.5% of 259 LT/ST strains, were associated with MSD (p = 0.0011). No other minor CF exhibited prevalence ≥5% and significant association with MSD. CONCLUSIONS/SIGNIFICANCE: Major CF-based efficacious ETEC vaccines could potentially prevent up to 66% of pediatric MSD cases due to ST-encoding ETEC in developing countries; adding CS14 extends coverage to ~77%.

Upsurge and spread of G3 rotaviruses in Eastern India (2014-2016): Full genome analyses reveals heterogeneity within Wa-like genomic constellation.

Advent of new strains and shift in predominantly circulating genotypes are characteristics of group- A rotavirus (RVA), one of the major causes of childhood gastroenteritis. During diarrheal disease surveillance at Kolkata, India (2014-2016), a shift in circulating RVA strains from G1P[8] to G3P[8] was seen. Stool samples from children (n = 3048) with acute gastroenteritis were tested of which 38.7% were RVA positive. G1 was the predominant strain (65.3%) in 2014-2015 whereas in late 2015 and 2016, G3 became the preponderant strain (44.6%). In the past decade G3 strains were not observed in this region, we conducted whole genome sequencing of representative strains to gain insight into the phenomenon of emergence and genetic constellation of these circulating human G3 strains. The analyses revealed intergenogroup reassortment in G3P[4] strains (among Wa and DS-1-like genogroup) whereas G3P[8] strains were authentic Wa-like. Phylogenetic analysis revealed Kolkata G3 strains as polymorphic and thus they formed two sub-clusters due to antigenic differences in their VP7 protein. One of the sub-clusters had the wild-type threonine at 87 amino acid position while another sub-cluster had an isoleucine mutation. Presence of additional N-linked glycosylation site at amino acid 283 of VP7 glycoprotein suggests that the major neutralizing epitope on the VP7 (G3) of RotaTeq vaccine differs from the currently circulating G3 strains. The study is important as efficiency of rotavirus vaccine depends on the circulating heterogeneous genotype constellations. Continuous monitoring of circulating RVA strains in endemic settings like India is therefore important in pre- and post-vaccination period to monitor the emergence of new reassortant genotypes in addition to assessing vaccine efficacy.

Antibody secreting B cells and plasma antibody response to rotavirus vaccination in infants from Kolkata India.

BACKGROUND: Assessing immune response after rotavirus vaccination consists in measuring serum or plasma IgA and IgG antibodies, but these assays provide very little information about the mucosal immune response. Thus the development of assays for detection of mucosal immune response following rotavirus vaccination is essential. We evaluate to assess circulating antibody-secreting cells (ASCs) as a potential means to evaluate mucosal immune responses to rotavirus vaccine. METHODS: 372 subjects, aged 6 weeks, were enrolled in the study. All the subjects were assigned to receive two doses of Rotarix® vaccine. Using a micro-modified whole blood-based ELISPOT assay, circulating rotavirus type-specific IgA- and IgG-ASCs, including gut homing ß7+ ASCs, were enumerated on week 6 before the first dose of Rotarix vaccination at 7 weeks of age and week 18 after the second vaccination at 17 weeks of age. Plasma samples collected before vaccination, and after two doses of Rotarix® vaccination were tested for plasma rotavirus IgA titers. RESULTS: Two doses of Rotarix® provided to induce sero-protective titer of ≥ 20 Units in 35% of subjects. Total blood IgA- ASC responses were detected in 26.4% of subjects who were non-responder before vaccination. Among responders, 47% of the subjects also have sero-protective plasma IgA titers. DISCUSSION: Our results suggest that virus-specific blood gut homing ASCs were detected and provide insight into mucosal immune response after rotavirus vaccination. Further studies are needed to evaluate the duration of such immune responses and to assess the programmatic utility of this whole blood-based mucosal ASC testing for the rotavirus immunization program.

Use of oral cholera vaccine as a vaccine probe to define the geographical dimensions of person-to-person transmission of cholera.

BACKGROUND: Cholera is known to be transmitted from person to person, and inactivated oral cholera vaccines (OCVs) have been shown to confer herd protection via interruption of this transmission. However, the geographic dimensions of chains of person-to-person transmission of cholera are uncertain. The ability of OCVs to confer herd protection was used to define these dimensions in two cholera-endemic settings, one in rural Bangladesh and the other in urban India. METHODS: Two large randomized, placebo-controlled trials of inactivated OCVs, one in rural Matlab, Bangladesh and the other in urban Kolkata, India, were reanalyzed. Vaccine herd protection was evaluated by relating the risk of cholera in placebo recipients to vaccine coverage of surrounding residents residing within concentric rings. In Matlab, concentric rings in 100-m increments up to 700m were evaluated; in Kolkata, 50-m increments up to 350m were evaluated. RESULTS: One hundred and eight cholera cases among 24667 placebo recipients were detected during 1year of post-vaccination follow-up at Matlab; 128 cholera cases among 34968 placebo recipients were detected during 3 years of follow-up in Kolkata. Consistent inverse relationships were observed between vaccine coverage of the ring and the risk of cholera in the central placebo recipient for rings with radii up to 500m in Matlab and up to 150m in Kolkata. CONCLUSIONS: These results suggest that the dimensions of chains of person-to-person transmission in endemic settings can be quite large and may differ substantially from setting to setting. Using OCVs as 'probes' to define these dimensions can inform geographical targeting strategies for the deployment of these vaccines in endemic settings.

Species A rotaviruses isolated from hospitalized patients over 5 years of age in Kolkata, India, in 2012/13.

In hospital-based diarrhoeal disease surveillance at Infectious Diseases & Beliaghata-General Hospital (May-2012 to April-2013), Kolkata, India, stool samples were collected from patients < 5 years (n = 830) and > 5 years of age (n = 728) hospitalized with diarrhea. Group-A rotavirus (GARV) was identified by ELISA followed by multiplex RT-PCR. In children < 5 years of age, 53.4% of the samples were positive for GARV. In patients > 5 years to 90 years old, only 6.04% (n = 44) tested positive for GARV. G2P[4] strains (n = 16 [36.36%]) were the most prevalent, followed by G9P[4] strains (n = 13 [29.54%]), while P[4]-(n = 30 [68.18%]) was most prevalent among the P genotypes. The GARV strains G2, G9 and P[4] detected in adults clustered together in the phylogenetic tree with the GARV strains identified in children (< 5 years) during the same period. Rotavirus positivity was high among female patients (75%), suggesting that caregivers (mother/grandmother/older-siblings) may get infected through young children or may act as carriers for transmission.

A randomized Phase III clinical trial to assess the efficacy of a bovine-human reassortant pentavalent rotavirus vaccine in Indian infants.

Rotavirus is the most common cause of moderate-to-severe infant diarrhoea in developing countries, resulting in enormous morbidity, mortality, and economic burden. A bovine-human reassortant pentavalent rotavirus vaccine (BRV-PV) targeting the globally most common strains was developed in India and tested in a randomized, double-blind, placebo-controlled end-point driven Phase III efficacy clinical trial implemented at six sites across India. Infants 6 to 8weeks of age were randomized (1:1) to receive three oral doses of BRV-PV or placebo at 6, 10, and 14weeks of age along with routine vaccines. Home visit surveillance was conducted to detect severe rotavirus gastroenteritis (SRVGE) and safety outcomes until the children reached two years of age. A total of 3749 infants received BRV-PV while 3751 received placebo. At the time of the primary end-point (when the minimum number of cases needed for analysis were accrued) the vaccine efficacy against SRVGE was 36% (95% CI 11.7, 53.6, p=0.0067) in the per protocol (PP) analysis, and 41.9% (95% CI 21.1, 57.3, p=0.0005) in the intent to treat (ITT) analysis. Vaccine efficacy over the entire follow-up period (until children reached two years of age) was 39.5% (95% CI 26.7, 50, p<0.0001) in the PP analysis and 38.8% (95% CI, 26.4, 49, p<0.0001) in the ITT analysis. Vaccine efficacy against the very severe rotavirus cases (VSRVGE, Vesikari score≥16) was 60.5% (95% CI 17.7, 81, p=0.0131) at the time of the primary analysis and 54.7% (95% CI 29.7, 70.8, p=0.0004) for the complete follow-period in the PP population. The incidence of solicited, unsolicited, and serious adverse events were similar in both the vaccine and placebo groups. Likewise, the number of intussusceptions and deaths were similar between both groups. Thus, BRV-PV is an effective, well tolerated and safe vaccine in Indian infants. (Trial registration: Clinical Trials.Gov [NCT 02133690] and Clinical Trial Registry of India [CTRI/2013/05/003667]).

Comparison of IPV to tOPV week 39 boost of primary OPV vaccination in Indian infants: an open labelled randomized controlled trial.

BACKGROUND: The final endgame strategy of global polio eradication initiative includes switching from trivalent oral poliovirus vaccines (tOPV) to bivalent oral polio vaccine (bOPV), and introduction of inactivated poliovirus vaccine (IPV). This study compares IPV with tOPV week 39 boost in Indian infants. METHODS: Starting 28 March 2012, we enrolled 372 Indian infant-mother pairs from Kolkata, India in an open-label, block-randomized, controlled trial comparing a 39 week tOPV to an IPV boost among infants immunized with three doses of tOPV. The primary outcome was mucosal immunity to poliovirus as measured by fecal polio virus shedding after OPV challenge. The secondary outcome was humoral response as defined by >1:8 titers for neutralizing antibodies at week 40. Seroconversion was measured by change in level of antibody titers from week 18 to week 40. The analyses were performed by both intention-to-treat (ITT) and per-protocol (PP) comparing the occurrences of outcomes between the arms of the study. FINDINGS: Both the study arms provided equivalent mucosal immunity at 52 weeks with a total shedding prevalence of 28%. Vaccination with IPV resulted in significantly higher seroconversion rates for Polio type 2 (p = 0.03) and Polio type 3 (p < 0.01). CONCLUSIONS: This study indicates that an IPV boost at week 39 is equivalent to tOPV in intestinal immunity, and provides higher seroconversion compared to tOPV. The major limitation of the study was the additional OPV doses receive by infants during pulse polio immunization resulted in additional mucosal boosting, diminishing the impact of IPV or tOPV boost at week 39. However, IPV for OPV boost should prove to be a step forward in the global polio eradication initiative to reduce the problem of circulating vaccine-derived poliovirus (cVDPV).

Influence of maternal and socioeconomic factors on breast milk fatty acid composition in urban, low-income families.

The lipid composition of breast milk may have a significant impact on early infant growth and cognitive development. Comprehensive breast milk data is lacking from low-income populations in the Indian subcontinent impeding assessment of deficiencies and limiting development of maternal nutritional interventions. A single breast milk specimen was collected within 6 weeks postpartum from two low-income maternal cohorts of exclusively breastfed infants, from Dhaka, Bangladesh (n = 683) and Kolkata, India (n = 372) and assayed for percentage composition of 26 fatty acids. Mature milk (>15 days) in Dhaka (n = 99) compared to Kolkata (n = 372) was higher in total saturated fatty acid (SFA; mean 48% vs. 44%) and disproportionately lower in ω3-polyunsaturated fatty acid (PUFA), hence the ω6- and ω3-PUFA ratio in Dhaka were almost double the value in Kolkata. In both sites, after adjusting for days of lactation, increased maternal education was associated with decreased SFA and PUFA, and increasing birth order or total pregnancies was associated with decreasing ω6-PUFA or ω3-PUFA by a factor of 0.95 for each birth and pregnancy. In Dhaka, household prosperity was associated with decreased SFA and PUFA and increased ω6- and ω3-PUFA. Maternal height was associated with increased SFA and PUFA in Kolkata (1% increase per 1 cm), but body mass index showed no independent association with either ratio in either cohort. In summary, the socioeconomic factors of maternal education and household prosperity were associated with breast milk composition, although prosperity may only be important in higher cost of living communities. Associated maternal biological factors were height and infant birth order, but not adiposity. Further study is needed to elucidate the underlying mechanisms of these effects.

Molecular characterization of enteric adenovirus genotypes 40 and 41 identified in children with acute gastroenteritis in Kolkata, India during 2013-2014.

Human enteric adenovirus (HAdV) belonging to species F is one of the most common pathogens responsible for infantile gastroenteritis worldwide. This study was initiated to estimate prevalence and types of HAdV among children below 5 years of age seeking health care facility for treatment of diarrhea in Kolkata, Eastern India. A total of 1,562 stool specimens were tested during 2013-2014 and among them, 185 (11.8%) were positive for enteric HAdV. Maximum number of positive cases were observed among children between 6 and 12 months of age (13.9%). HAdV infection occurred at a low frequency throughout the year, with an increased incidence in the month of March-July in both the years. Among HAdV positive samples (n = 185), 44.8% showed coinfection with rotavirus. Genotyping based on hypervariable region of hexon and partial shaft region of fiber genes, revealed prevalence of HAdV-40 over HAdV-41 genotype during this study period. Nucleotide sequence analysis of HAdV-40 strains exhibited more than 99% similarity among themselves and 98.5% with the prototype strain Dugan. Sequence analysis of six hypervariable regions (HVRs) of hexon genes from all the HAdV-41 strains revealed co-circulation of both genome type cluster 1(GTC1) and GTC2. The presence of both types of GTCs reflects accumulation of amino acid (aa) mutations in HVR of hexon gene. A recombination event was evident in a subset of HAdV-41 strains where hexon gene belonged to GTC1 whereas, fiber gene clustered with GTC2. Sequence analysis of fiber gene shaft region of HAdV-41 strains revealed 15 aa deletion from the 15th repeat motif. J. Med. Virol. 89:606-614, 2017. © 2016 Wiley Periodicals, Inc.

Potential for Controlling Cholera Using a Ring Vaccination Strategy: Re-analysis of Data from a Cluster-Randomized Clinical Trial.

INTRODUCTION: Vaccinating a buffer of individuals around a case (ring vaccination) has the potential to target those who are at highest risk of infection, reducing the number of doses needed to control a disease. We explored the potential vaccine effectiveness (VE) of oral cholera vaccines (OCVs) for such a strategy. METHODS AND FINDINGS: This analysis uses existing data from a cluster-randomized clinical trial in which OCV or placebo was given to 71,900 participants in Kolkata, India, from 27 July to 10 September 2006. Cholera surveillance was then conducted on 144,106 individuals living in the study area, including trial participants, for 5 y following vaccination. First, we explored the risk of cholera among contacts of cholera patients, and, second, we measured VE among individuals living within 25 m of cholera cases between 8 and 28 d after onset of the index case. For the first analysis, individuals living around each index case identified during the 5-y period were assembled using a ring to define cohorts of individuals exposed to cholera index cases. An index control without cholera was randomly selected for each index case from the same population, matched by age group, and individuals living around each index control were assembled using a ring to define cohorts not exposed to cholera cases. Cholera attack rates among the exposed and non-exposed cohorts were compared using different distances from the index case/control to define the rings and different time frames to define the period at risk. For the VE analysis, the exposed cohorts were further stratified according to the level of vaccine coverage into high and low coverage strata. Overall VE was assessed by comparing the attack rates between high and low vaccine coverage strata irrespective of individuals' vaccination status, and indirect VE was assessed by comparing the attack rates among unvaccinated members between high and low vaccine coverage strata. Cholera risk among the cohort exposed to cholera cases was 5-11 times higher than that among the cohort not exposed to cholera cases. The risk gradually diminished with an increase in distance and time. The overall and indirect VE measured between 8 and 28 d after exposure to a cholera index case during the first 2 y was 91% (95% CI 62%-98%) and 93% (95% CI 44%-99%), respectively. VE persisted for 5 y after vaccination and was similar whether the index case was a young child (<5 y) or was older. Of note, this study was a reanalysis of a cholera vaccine trial that used two doses; thus, a limitation of the study relates to the assumption that a single dose, if administered quickly, will induce a similar level of total and indirect protection over the short term as did two doses. CONCLUSIONS: These findings suggest that high-level protection can be achieved if individuals living close to cholera cases are living in a high coverage ring. Since this was an observational study including participants who had received two doses of vaccine (or placebo) in the clinical trial, further studies are needed to determine whether a ring vaccination strategy, in which vaccine is given quickly to those living close to a case, is feasible and effective. TRIAL REGISTRATION: ClinicalTrials.gov NCT00289224.

The Burden of Cryptosporidium Diarrheal Disease among Children < 24 Months of Age in Moderate/High Mortality Regions of Sub-Saharan Africa and South Asia, Utilizing Data from the Global Enteric Multicenter Study (GEMS).

BACKGROUND: The importance of Cryptosporidium as a pediatric enteropathogen in developing countries is recognized. METHODS: Data from the Global Enteric Multicenter Study (GEMS), a 3-year, 7-site, case-control study of moderate-to-severe diarrhea (MSD) and GEMS-1A (1-year study of MSD and less-severe diarrhea [LSD]) were analyzed. Stools from 12,110 MSD and 3,174 LSD cases among children aged <60 months and from 21,527 randomly-selected controls matched by age, sex and community were immunoassay-tested for Cryptosporidium. Species of a subset of Cryptosporidium-positive specimens were identified by PCR; GP60 sequencing identified anthroponotic C. parvum. Combined annual Cryptosporidium-attributable diarrhea incidences among children aged <24 months for African and Asian GEMS sites were extrapolated to sub-Saharan Africa and South Asian regions to estimate region-wide MSD and LSD burdens. Attributable and excess mortality due to Cryptosporidium diarrhea were estimated. FINDINGS: Cryptosporidium was significantly associated with MSD and LSD below age 24 months. Among Cryptosporidium-positive MSD cases, C. hominis was detected in 77.8% (95% CI, 73.0%-81.9%) and C. parvum in 9.9% (95% CI, 7.1%-13.6%); 92% of C. parvum tested were anthroponotic genotypes. Annual Cryptosporidium-attributable MSD incidence was 3.48 (95% CI, 2.27-4.67) and 3.18 (95% CI, 1.85-4.52) per 100 child-years in African and Asian infants, respectively, and 1.41 (95% CI, 0.73-2.08) and 1.36 (95% CI, 0.66-2.05) per 100 child-years in toddlers. Corresponding Cryptosporidium-attributable LSD incidences per 100 child-years were 2.52 (95% CI, 0.33-5.01) and 4.88 (95% CI, 0.82-8.92) in infants and 4.04 (95% CI, 0.56-7.51) and 4.71 (95% CI, 0.24-9.18) in toddlers. We estimate 2.9 and 4.7 million Cryptosporidium-attributable cases annually in children aged <24 months in the sub-Saharan Africa and India/Pakistan/Bangladesh/Nepal/Afghanistan regions, respectively, and ~202,000 Cryptosporidium-attributable deaths (regions combined). ~59,000 excess deaths occurred among Cryptosporidium-attributable diarrhea cases over expected if cases had been Cryptosporidium-negative. CONCLUSIONS: The enormous African/Asian Cryptosporidium disease burden warrants investments to develop vaccines, diagnostics and therapies.

Use of the data system for field management of a clinical study conducted in Kolkata, India.

BACKGROUND: Designing an appropriate data system is important to the success of a clinical study. However, little information is available on this topic. We share our experiences on designing, developing, and implementation of a data system for management of data and field activities of a complex clinical study. METHODS: The data system was implemented aiming at determining the biological basis for the underperformance of oral vaccines, such as polio and rotavirus vaccines in children at a site in Kolkata, India. The system included several functionalities to control data and field activities. It was restricted to authorized users based on their access privileges. A relational database platform was chosen, and Microsoft Visual FoxPro 7.0 (Microsoft Corporation, Seattle, WA, USA) was used to develop the system. The system was installed at the clinic and data office to facilitate both the field and data management activities. RESULTS: Data were doubly entered by two different data operators to identify keypunching errors in the data. Outliers, duplication, inconsistencies, missing entries, and linkage were also checked. Every modification and users log-in/log-out information was auto-recorded in an audit trail. The system offered tools for preparation of visit schedule of the participants. A visit considered as protocol deviation was documented by the system. The system alerted field staff to every upcoming visit date to organize the field activities and to inform participants which day to come. The system also produced a growth chart for evaluating nutritional status and referring the child to a specialized clinic if found to be severely malnourished. CONCLUSION: The data system offered unique features for controlling for both data and field activities, which led to minimize drop-out rates as well as protocol deviations. Such system is warranted for a successful clinical study.