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Abstract: Picobirnaviruses (PBVs) with bisegmented small RNA genome profile (1.75 and 1.55kbp for segment 1 and 2, respectively) were detected from 1999 to 2003 in faecal specimens of acute watery diarrhoea cases, largely children (n =20) and an adult in Kolkata, India. Varying degrees of dehydration necessitated their visit to hospital for further treatment and management of acute watery diarrhoea. PBV was associated with rotavirus (n =3) or astrovirus (n =3) and with both in one case. No co-infection with norovirus, sapovirus or adenovirus was detected in the picobirnavirus positive cases. No co-infection with parasites (Cryptosporidium spp., Giardia spp., Entamoeba spp., helminths) or bacteria (Vibrio spp., Shigella spp., Escherichia coli) was detected among the picobirnavirus positive cases. There was a single instance of co-infection with Salmonella spp. (n =1). PBVs not associated with serious diarrhoea illness and showing large genome profile (2.3–2.6 and 1.5–1.9kbp for segment 1 and 2, respectively) have earlier been reported in adult individuals and recently among children from a slum community in Kolkata, India. The short genome profile PBVs associated with acute watery diarrhoea may be another emerging diarrhoeagenic virus in Kolkata, India. Molecular characterization using reported primers PicoB25–PicoB43 for Genogroup I and PicoB23–PicoB24 for Genogroup II in RT-PCR showed the presence of Genogroup I PBVs (n =6) and Genogroup II PBVs (n =5), while some could not be amplified (n =3) with these primers. Sequence analysis of Genogroup I amplicons indicated remarkable sequence heterogeneity. After more than a decade, four PBV positives of Genogroup II were detected during this study. Phylogenetic analysis showed varying degree of genetic diversity amongst PBV strains from Kolkata and other countries. [Copyright &y& Elsevier]

Abstract: The study is aimed to determine the seasonal distribution and clinical characteristics of astroviruses associated with acute watery diarrhoea among children in Kolkata and characterize them at the molecular level. Method of study: Faecal specimens of acute watery diarrhoea cases (n =857) and non-diarrhoeic samples (n =211) from the hospitals and a nearby field community were screened with IDEIA Astrovirus detection kit; astrovirus co-infections with rotavirus and/or picobirnavirus were detected by RNA-PAGE and silver staining. Further RT-PCR was carried out using specific primers, viz. Mon340 (+) and Mon348 (−) targeting a highly conserved domain of ORF1a (289bp) of human astroviruses. Results: Astrovirus infection was detected in 50 cases (50/857); astroviruses were detected mostly in children aged 6–12 months (50%); all non-diarrhoeic samples (n =211) were negative for astrovirus. In 52% of astrovirus positive cases, the virus was detected as the sole agent; mixed infections were also detected with other diarrhoeic pathogens such as rotavirus (32%), picobirnavirus (2%), rotavirus and picobirnavirus (2%), picobirnavirus and Enterotoxigenic E. coli (ETEC) (2%), rotavirus and ETEC (2%), rotavirus and Enteroaggregative E. coli (EAEC) (2%), Enteropathogenic E. coli (EPEC) (2%), Shigella flexneri type 3a (2%) and Ascaris (2%). RT-PCR and sequencing of amplicons of astroviruses from Kolkata, with specific primers targeted to the conserved domain of ORF1a (289bp) of the astrovirus genome, showed maximum homology to the astrovirus strain (“5–158”) from Seoul (98%). Results and Conclusions: Clinical characteristics of the diarrhoeic children in Kolkata indicated that astrovirus infections were detected throughout the year and were associated with varying degree of dehydration and acute watery diarrhoea. In-depth molecular epidemiological surveillance of astroviruses in Kolkata is essential for better understanding of their overall genetic nature. [Copyright &y& Elsevier]

Abstract: Picobirnaviruses are a group of unclassified, non-enveloped, small spherical viruses, 35–41nm in diameter without any apparent surface morphology. They have characteristic bisegmented double stranded RNA genome of two types namely large profile (2.3–2.6kbp for the larger and 1.5–1.9kbp for the smaller segment, respectively) or small profile (1.75 and 1.55kbp for segments 1 and 2, respectively). Human picobirnaviruses (n =12 positives; 2/56 diarrhoeic children and 10/607 non-diarrhoeic children) with large (n =11) or small (n =1) genome pattern were observed in faecal specimens of children from a slum community by silver stained PAGE gels. Faecal specimen from four asymptomatic cases (P597_02_IND, K135_02_IND, A373_03_IND, A356_03_IND) and one diarrhoeic case (K135_03_IND) had genogroup I picobirnaviruses (1-CHN-97 like) showing amplicons within the 201bp region, with primers PicoB25–PicoB43, targeting the conserved domain of RNA-dependent RNA polymerase (RdRp) gene. It was interesting to note that only the PBV strain P597_02_IND from Kolkata with large genome was closely related to a reported strain (similarity with 2-GA-91 from USA was 87% at the nucleotide level and 90% at the amino acid level). Sequence analysis showed three conserved amino acid domains as well as a highly conserved D-S—D motif, characteristic of RNA-dependent RNA polymerase gene of bisegmented, double stranded RNA viruses. Sequence data of the picobirnavirus A356_03_IND indicated strong heterogeneity with all other picobirnavirus strains sequenced till date. After nearly a decade a genogroup II picobirnavirus strain (R227_03_IND) was isolated from a diarrhoea case in the community, with small genome profile and amplified with specific primers PicoB23–PicoB24; but the sequence data showed that it was divergent from the hitherto reported prototype strain 4-GA-91 of genogroup II human picobirnaviruses. [Copyright &y& Elsevier]