Functional Single-Nucleotide Polymorphisms in the MBL2 and TLR3 Genes Influence Disease Severity in Influenza A (H1N1)pdm09 Virus-Infected Patients from Maharashtra, India.

The clinical outcome in influenza A (H1N1)pdm09 virus-infected subjects is determined by several factors, including host genetics. In the present study, single-nucleotide polymorphisms (SNPs) in the IFITM, MBL2, TLR3, TLR8, DDX58, IFIH1, CD55, and FCGR2, genes were investigated in influenza A (H1N1)pdm09 virus-infected subjects to find out their association with disease severity. Influenza A (H1N1)pdm09 virus-infected subjects with severe disease (n = 86) and mild disease (n = 293) from western India were included in the study. The SNPs were investigated by PCR-based methods. The results revealed a higher frequency of TLR3 rs5743313 T/T genotype [odds ratio (OR) with 95% confidence interval (CI) 2.55 (1.08-6.04) p = 0.039] and TLR3 two-locus haplotype rs3775291-rs3775290 T-A [OR with 95% CI 7.94 (2.05-30.68)] in severe cases. Lower frequency of the mutant allele of MBL2 rs1800450 [OR with 95% CI 0.51 (0.27-0.87), p = 0.01] and TLR3 two-locus haplotype rs3775291-rs3775290 T-G [OR with 95% CI 0.48 (0.27-0.85)] was observed in severe cases compared with cases with mild disease. Higher frequency of TLR3 two-locus haplotype rs3775291-rs3775290 T-A was observed in severe cases [OR with 95% CI 7.9 (2.0-30.7)]. The allele and genotype frequencies of other SNPs were not different between the study categories. The results suggest that the functional SNPs in MBL2 and TLR3 are associated with severe disease in influenza A (H1N1)pdm09 virus-infected subjects.

TNFA and IL10 Polymorphisms and IL-6 and IL-10 Levels Influence Disease Severity in Influenza A(H1N1)pdm09 Virus Infected Patients.

Cytokines are key modulators of immune response, and dysregulated production of proinflammatory and anti-inflammatory cytokines contributes to the pathogenesis of influenza A(H1N1)pdm09 virus infection. Cytokine production is impacted by single nucleotide polymorphisms (SNPs) in the genes coding for them. In the present study, SNPs in the IL6, TNFA, IFNG, IL17A, IL10, and TGFB were investigated for their association with disease severity and fatality in influenza A(H1N1)pdm09-affected patients with mild disease (n = 293) and severe disease (n = 86). Among those with severe disease, 41 patients had fatal outcomes. In a subset of the patients, levels of IL-2, IL-4, IL-6, IL-10, TNF, IFN-γ, and IL-17 were assayed in the plasma for their association with severe disease. The frequency of TNFA rs1800629 G/A allele was significantly higher in severe cases and survived severe cases group compared to that of those with mild infection (OR with 95% for mild vs. severe cases 2.95 (1.52-5.73); mild vs. survived severe cases 4.02 (1.84-8.82)). IL10 rs1800896-rs1800872 G-C haplotype was significantly lower (OR with 95% 0.34 (0.12-0.95)), while IL10 rs1800896-rs1800872 G-A haplotype was significantly higher (OR with 95% 12.11 (2.23-76.96)) in fatal cases group compared to that of the mild group. IL-6 and IL-10 levels were significantly higher in fatal cases compared to that of survived severe cases. IL-6 levels had greater discriminatory power than IL-10 to predict progression to fatal outcome in influenza A(H1N1)pdm09 virus-infected patients. To conclude, the present study reports the association of TNFA and IL10 SNPs with severe disease in Influenza A(H1N1)pdm09 virus-infected subjects. Furthermore, IL-6 levels can be a potential biomarker for predicting fatal outcomes in Influenza A(H1N1)pdm09 virus infected subjects.

Prevalence and genetic diversity of gastroenteritis viruses in hospitalized children < 5 years of age in Maharashtra state, Western India, 2017-2019.

Four gastroenteritis viruses were responsible for 54% of the acute gastroenteritis (AGE) cases in children hospitalized between May 2017 and December 2019 in Pune city of Maharashtra state, Western India. The majority (79%) of the children were <2 years of age. The prevalence of Rotavirus A (RVA) was 30.5% followed by 14.3% for norovirus, 8.4% for adenovirus, and 5.5% for astrovirus. The severity of the disease was highest in patients with coinfections compared with the patients with a single infection or negative for all (p = 0.024). Genotyping analysis showed that the majority of the RVA-positive samples (66%) could be typed as G3P[8], 63.6% of the norovirus as GII.4 Sydney [P16], 44% of the adenovirus as type 41%, and 56.2% of the astrovirus as astrovirus type 1. The almost equivalent prevalence of rotavirus and nonrotaviruses and acute gastroenteritis (AGE) cases without known etiology in around 46% of the cases was noted in the present study. Our data highlight that after the recent inclusion of rotavirus vaccines as a part of the National Immunization schedule in India, conducting extensive AGE surveillance in children should include nonrotaviruses such as norovirus.

Gut microbial diversity during pregnancy and early infancy: an exploratory study in the Indian population.

The gut microbial community is known to influence the human health and disease state and is shaped by various factors since birth. It is now evident that understanding the alterations in these commensal microbes during crucial stages of life is of utmost importance to determine and predict the health status of an individual. To study the gut microbiota in two such vital stages, pregnancy and infancy, we analyzed gut microbial communities from 20 mother-infant dyads at different stages of pregnancy and early infancy. In total, we analyzed 80 fecal samples for profiling the gut microbial community using 16S rRNA gene-based sequencing. We observed no significant alterations in the gut bacterial diversity during pregnancy; however, significant alterations were observed during the period from birth to six months in infants, with a reduction in Staphylococcus and Enterococcus and an increase in Bifidobacterium and Streptococcus with a more stable microbial community at the age of six months.

Clinical and genetic characteristics of unusual G12P[11] rotavirus strains recovered from neonates: A study from Pune, Western India.

Rotavirus infections in neonates are generally nosocomial, and differ from pediatric infections both clinically and epidemiologically. These infections are predominantly asymptomatic and often associated with unusual strains. Globally, so far limited data is available on rotavirus infections in neonates admitted at Neonatal Intensive Care Unit. The aim of the present study is to determine the prevalence of rotavirus among neonates and to study their genetic characteristics. Stool specimens (n = 701) collected from neonates (n = 621) admitted during April 2016 to March 2018 mainly for prematurity, low birth weight and associated respiratory distress syndrome from two hospitals from Pune were tested for rotavirus, genotyped and representative strains were sequenced for the genes encoding outer capsid proteins, VP7 and VP4. Rotavirus was detected in 24.31% neonates. Majority of rotavirus infected neonates (98.68%) were asymptomatic. Peak rotavirus antigen detection (91.38%) occurred during the first 2 weeks of admission. Low, very low and normal birth weight neonates with gestational age ≥28 weeks had significantly higher rotavirus infection than those with extreme low birth weight with gestational age <28 weeks. Rotaviral infections occurred almost evenly throughout the year without an apparent peak in colder months. Predominance of unusual G12P[11] strains (97.1%) was observed. Phylogenetic analysis of the partial VP7 coding gene revealed all G12 strains clustered in lineage III and shared 96.94%-100% (nucleotide) and 96.26%-100% (amino acid) identities among themselves, and 95.69%-98.98% (nucleotide) and 94.77%-98.98% (amino acid) with other lineage III G12 strains respectively. Similarly VP4 partial gene sequences of P[11] study strains shared 97.5%-100% (nucleotide and amino acid) identities among themselves and highest 93.34%-94.53% (nucleotide) and 93.57%-94.64% (amino acid) identity with vaccine strain 116E, G9P[11]. The study highlights high frequency of unusual G12P[11] strains among neonates for the first time in western India and reaffirms limited strain diversity in this population. The knowledge of neonatal strains is important for estimating the efficacies of rotavirus vaccines.

Association of Single Nucleotide Polymorphisms in TNFA and IL10 Genes with Disease Severity in Influenza A/H1N1pdm09 Virus Infections: A Study from Western India.

Susceptibility to severe influenza A/H1N1pdm09 virus is multifactorial. The present study was carried out in 246 patients infected with A/H1N1pdm09 virus to find out whether single nucleotide polymorphisms (SNPs) in the genes coding for proinflammatory and anti-inflammatory cytokines are associated with disease severity. Among the cases, 129 had mild disease, whereas 117 had severe disease. There were 27 fatal cases. TNFA rs1800629, IFNG rs2430561, IL10 rs1800872, IL10 rs1800896, and CCL2 rs1024611 SNPs were genotyped by polymerase chain reaction-based methods. A significantly higher frequency of TNFA rs1800629 "G/A" genotype was observed in severe and fatal cases compared with mild and survived cases, respectively. In a dominant mode, IL10 rs1800896 "G" allele was significantly negatively associated with disease severity. IL10 rs1800896 "C/A" genotype was significantly associated with fatality in influenza A/H1N1pdm09 infections. The results suggest that SNPs in the IL10 and TNFA genes might be associated with disease severity in influenza A/H1N1pdm09-infected patients.

A cross-sectional comparative study of gut bacterial community of Indian and Finnish children.

The human gut microbiome plays a crucial role in the compositional development of gut microbiota. Though well documented in western pediatrics population, little is known about how various host conditions affect populations in different geographic locations such as the Indian subcontinent. Given the impact of distinct environmental conditions, our study assess the gut bacterial diversity of a small cohort of Indian and Finnish children and investigated the influence of FUT2 secretor status and birth mode on the gut microbiome of these populations. Using multiple profiling techniques, we show that the gut bacterial community structure in 13-14-year-old Indian (n = 47) and Finnish (n = 52) children differs significantly. Specifically, Finnish children possessed higher Blautia and Bifidobacterium, while genera Prevotella and Megasphaera were predominant in Indian children. Our study also demonstrates a strong influence of FUT2 and birth mode variants on specific gut bacterial taxa, influence of which was noticed to differ between the two populations under study.

Changing trends in circulating rotavirus strains in Pune, western India in 2009-2012: emergence of a rare G9P[4] rotavirus strain.

BACKGROUND: A vast diversity in rotaviruses at inter- and intra-genotypic level underscores the need for monitoring of circulating rotavirus strains. The aim of this study was to update the data on rotavirus disease and strains for the period from January 2009 to December 2012 in Pune, western India which has been one of the sites of the Indian Rotavirus Strain Surveillance Network since November 2005. METHODS: Children aged <5 years admitted for acute gastroenteritis in three different hospitals from Pune city were included in the study. The stool specimens were collected and tested for rotavirus antigen by a commercial enzyme immunoassay. The rotavirus strains were genotyped by multiplex reverse transcription polymerase chain reaction. RESULTS: During the study period, we found 35.1% of 685 stool specimens contained rotavirus antigen. Frequency of rotavirus detection was greatest (58.5%) among children aged 7-12 months. The G1P[8] (31.4%), G2P[4] (20.2%) and G9P[8] (11.8%) strains were the most common types. We noted predominance of G1P[8] strains (39.6%-46.1%) in all the years of study except 2009 wherein G9P[8] strains scored highest level (15.3%). Subsequent to this, we identified G9P[8] strains at the second highest position in 2010, their sudden decline and rise in G9P[4] strains in 2011-2012. We detected G12 strains in combination with P[6] and P[8] at variable rates (0-10.2%) and highest level (27.1%) of mixed rotavirus infections in 2009 as compared to 2010-2012 (0-3.8%). CONCLUSION: The study highlights the huge burden of rotavirus disease and changing profile of circulating rotavirus strains displaying emergence of G9P[4] reassortant strains in Pune, western India and emphasizes the need to analyze the entire genomic constellation of rotavirus strains for better evaluation of the impact of rotavirus.

Effectiveness of an Indian-made attenuated influenza A(H1N1)pdm 2009 vaccine: a case control study.

A live attenuated influenza A(H1N1)pdm 2009 vaccine was developed and distributed in India in 2010. We estimated the vaccine effectiveness (VE) against laboratory-confirmed pandemic H1N1 (pH1N1) infections in patients with influenza-like illness who visited five tertiary care hospitals in Pune, India during June-December 2010. Swab specimens were analyzed for influenza pH1N1 by reverse transcriptase polymerase chain reaction (RT-PCR). VE was estimated using the test-negative case-control study design and logistic regression. A total of 784 patients (253 cases, 531 controls) were analyzed. The unadjusted overall VE was 75.5% (95% confidence interval [CI] 42.1-89.7), while the adjusted VE was 76% (95% CI 42.1-89.7). We conclude that the live attenuated influenza A(H1N1)pdm 2009 vaccine was effective in our study population, which has opened prospects for using this platform for trivalent formulations.

Safety and immunogenicity of a new purified vero cell rabies vaccine (PVRV) administered by intramuscular and intradermal routes in healthy volunteers.

BACKGROUND: Rabies is 100% fatal but preventable with modern vaccines and immunoglobulins. There is a huge demand for rabies vaccines in developing countries of Asia and Africa. We have developed a new purified vero cell rabies vaccine (PVRV) and evaluated its safety and immunogenicity in healthy volunteers by intramuscular (IM) and intradermal (ID) routes of vaccination. METHODOLOGY: Sixty adults aged between 18 and 50 years were recruited in this actively controlled Phase I clinical study and were randomized to receive three 1 ml or 0.1 ml doses of new PVRV intramuscularly or intradermally on days 0, 7 and 21. The control group received commercially available PVRV (Verorab) by intramuscular route. Adverse events (AEs) were recorded with diary cards till day 28 post-vaccination. Immunogenicity was assessed on day 0, 7, 21 and 42 by rapid fluorescence focus inhibition test (RFFIT). RESULTS: In all, 116 solicited local and systemic events were reported across the three groups. Most were mild and resolved without sequelae. Also the few unsolicited events, deemed unrelated to the study vaccines, caused no problems. No significant changes in the routine laboratory parameters were found. Two doses of a vaccine elicited protective titres (≥ 0.5 IU/ml) in all subjects, the GMTs varying between 0.57 and 0.69 IU/ml on day 7, 3.07 and 3.97 IU/ml on day 21, and 6.12 and 8.52 IU/ml on day 42 post-vaccination. CONCLUSIONS: PVRV was well tolerated and showed good immunogenicity regardless of whether administered intramuscularly or, using a tenth of that volume, intradermally. Further studies with this new vaccine are warranted.

Epidemiological, clinical, and molecular features of norovirus infections in western India.

The study was conducted to investigate the molecular epidemiology of noroviruses (NoVs) from western India. A total of 830 fecal specimens were collected during July 2005-June 2007 from children, < or =7 years of age suffering from acute gastroenteritis in Pune, Nagpur, and Aurangabad cities. All the specimens were subjected to RT-PCR, sequencing and phylogenetic analysis for detection and characterization of Genogroup I (GI) and GII NoVs. NoV positivity varied between 6.3% and 12.6% in different cities with the predominance of GII (96.6%). NoV infections were very common in the patients < or =2 years of age. A majority (55%) of the patients suffered from severe disease, however, vomiting was not experienced in 35%. Coinfections with rotaviruses were found in 10% cases. Summer month seasonality supported NoV infections in western India. The phylogenetic analysis of partial RNA polymerase and VP1 (capsid) genes identified 2 GI (GI. 2 and GI.6) and 5 GII (GII.4, GII.6, GII.7, GII.8, and GII.14) genetic clusters with possible occurrence of "2007 new-variant" of GII.4. Six different combinations of RdRp and capsid genes (GII.b/GII.3, GII.b/GII.4, GII.d/GII.3, GII.b/GII.18, GII.1/GII.12 and GII.3/GII.13) were also identified. GII.4 (52%) prevailed in 2005-2006 while the predominance of probable recombinant NoV strains (58%) was noted in 2006-2007 with the contribution of GII.b/GII.3 at 79% level. GII.b/GII.18 type identified in 37% infections in 2005-2006 was completely replaced by GII.b/GII.3 type in 2006-2007. This is the first report that highlights the norovirus epidemiology and strain diversity demonstrating possible circulation of new variants in patients with acute gastroenteritis from western India.

Immunohistochemical analysis of Mallory bodies in Wilsonian and non-Wilsonian hepatic copper toxicosis.

Patients with Wilson's disease (WD), Indian childhood cirrhosis (ICC), and idiopathic copper toxicosis (ICT) develop severe liver disease morphologically characterized by ballooning of hepatocytes, inflammation, cytoskeletal alterations, and Mallory body (MB) formation, finally leading to mostly micronodular cirrhosis. The pathogenesis of MBs in copper toxicosis is still unresolved. Immunohistochemical analysis of MBs in different types of copper intoxication revealed that keratin, p62, and ubiquitin are integral components. Thus MBs associated with copper intoxication resemble those present in alcoholic steatohepatitis (ASH) and nonalcoholic steatohepatitis (NASH). p62 is a multifunctional immediate early gene product that, on the one hand, is involved in stress-induced cell signaling (particularly that of oxidative stress) by acting as an adapter protein linking receptor-interacting protein (RIP) with the atypical protein kinase C. On the other hand, p62 binds with high affinity to polyubiquitin and ubiquitinated proteins. In conclusion, p62 accumulation in WD, ICC, and ICT and deposition in MBs indicates a central role of protein misfolding induced by oxidative stress in copper-induced liver toxicity. By sequestering potentially harmful misfolded ubiquitinated proteins as inert cytoplasmic inclusion bodies (e.g., as MBs), p62 may be a major player in an important cellular rescue mechanism in oxidative hepatocyte injury.

Higher offspring birth weight predicts the metabolic syndrome in mothers but not fathers 8 years after delivery: the Pune Children's Study.

In Europid populations, low birth weight of offspring predicts insulin resistance in the mother and cardiovascular disease in both parents. We investigated the association between birth weight of offspring and obesity and cardiovascular risk in the parents of 477 8-year-old children born at the King Edward Memorial Hospital, Pune, India. Eight years after the birth of the child, mothers (33 years of age, n = 459) of heavier babies were taller and more obese (BMI, fat mass, and waist circumference, all P < 0.001) than mothers of lighter babies. Increasing offspring birth weight predicted higher homeostasis model assessment for insulin resistance (P < 0.01) and metabolic syndrome in mothers (P < 0.001) (adjusted for offspring sex and birth order, maternal age, and socioeconomic status) but not hyperglycemia. Fathers (39 years of age, n = 398) of heavier babies were taller and heavier, independent of maternal size (P < 0.01, both), but were not more insulin resistant. Unlike other reports, lower offspring birth weight did not predict insulin resistance in fathers. Thus, urban Indian parents have a higher risk of being obese 8 years after delivery of a heavier child. Mothers but not fathers of heavier babies also have a higher risk of being insulin resistant and developing the metabolic syndrome. Our findings highlight the need for a better understanding of the relation between fetal growth and future health before contemplating public health interventions to improve fetal growth.