title: Detection and molecular characterization of multiple strains of Picobirnavirus causing mixed infection in a diarrhoeic child: Emergence of prototype Genogroup II-like strain in Kolkata, India.
abstract: BACKGROUND
  Picobirnaviruses (PBVs) associated with viral gastroenteritis were reported from humans and several animal species to date. PBVs belonging to family Picobirnaviridae under proposed order Diplornavirales are small, non-enveloped, with bisegmented dsRNA genome.
  
  
  METHODS
  PBV was detected by polyacrylamide gel electrophoresis (PAGE) and silver staining. Confirmatory RT-PCR using primer pair PicoB25 (+) and PicoB43 (-) for genogroup I PBV and PicoB23(+) and PicoB24(-) for genogroup II PBV, resulted in amplicons of 201bp and 369bp respectively. The amplicons of genogroup I PBV were cloned and sequenced; amplicon of genogroup II PBV was directly sequenced. Further, the phylogenetic relationship and genetic diversity of strains from Kolkata was compared with hitherto reported PBV strains.
  
  
  RESULTS
  In PAGE, a faecal specimen showed three sets of PBV with large profile bisegmented genomic RNA with slight variation in migration pattern. Molecular cloning experiments confirmed that PBV/ Human/INDIA/GPBV6/2007 had mixed infection comprising four different strains of PBV genogroup I [GPBV6C1P-GPBV6C4P] and one PBV genogroup II strain [GPBV6G2P].
  
  
  CONCLUSIONS
  Sequence comparison and phylogenetic analysis of gene segment 2 of GPBV6 clones (C1, C2, C3 and C4) revealed low nucleotide identities (59-63%) and distant genetic relatedness to other human and porcine genogroup I picobirnaviruses. The strain GPBV6G2P represents another PBV genogroup II strain after prototype strain 4-GA-91/USA as genogroup II PBVs have seldom been reported to date, except from Kolkata, India and Netherlands. We are reporting the first incidence of detection of multiple strain (mixed) infection of picobirnavirus [genogroups I and II] from a diarrhoeic child in a slum community of Kolkata, India.
authors.forename: Balasubramanian
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authors.lastname: Ganesh
authors.forename: Shigeo
authors.initials: S
authors.lastname: Nagashima
authors.forename: Souvik
authors.initials: S
authors.lastname: Ghosh
authors.forename: Seegekote M
authors.initials: SM
authors.lastname: Nataraju
authors.forename: Krishnan
authors.initials: K
authors.lastname: Rajendran
authors.forename: Byomkesh
authors.initials: B
authors.lastname: Manna
authors.forename: Thandavarayan
authors.initials: T
authors.lastname: Ramamurthy
authors.forename: Swapan K
authors.initials: SK
authors.lastname: Niyogi
authors.forename: Suman
authors.initials: S
authors.lastname: Kanungo
authors.forename: Dipika
authors.initials: D
authors.lastname: Sur
authors.forename: Nobumichi
authors.initials: N
authors.lastname: Kobayashi
authors.forename: Triveni
authors.initials: T
authors.lastname: Krishnan