Complete genome sequence analysis of chicken astrovirus isolate from India.

OBJECTIVE: Chicken astroviruses have been known to cause severe disease in chickens leading to increased mortality and "white chicks" condition. Here we aim to characterize the causative agent of visceral gout suspected for astrovirus infection in broiler breeder chickens. METHODS: Total RNA isolated from allantoic fluid of SPF embryo passaged with infected chicken sample was sequenced by whole genome shotgun sequencing using ion-torrent PGM platform. The sequence was analysed for the presence of coding and non-coding features, its similarity with reported isolates and epitope analysis of capsid structural protein. RESULTS: The consensus length of 7513 bp genome sequence of Indian isolate of chicken astrovirus was obtained after assembly of 14,121 high quality reads. The genome was comprised of 13 bp 5'-UTR, three open reading frames (ORFs) including ORF1a encoding serine protease, ORF1b encoding RNA dependent RNA polymerase (RdRp) and ORF2 encoding capsid protein, and 298 bp of 3'-UTR which harboured two corona virus stem loop II like "s2m" motifs and a poly A stretch of 19 nucleotides. The genetic analysis of CAstV/INDIA/ANAND/2016 suggested highest sequence similarity of 86.94% with the chicken astrovirus isolate CAstV/GA2011 followed by 84.76% with CAstV/4175 and 74.48%% with CAstV/Poland/G059/2014 isolates. The capsid structural protein of CAstV/INDIA/ANAND/2016 showed 84.67% similarity with chicken astrovirus isolate CAstV/GA2011, 81.06% with CAstV/4175 and 41.18% with CAstV/Poland/G059/2014 isolates. However, the capsid protein sequence showed high degree of sequence identity at nucleotide level (98.64-99.32%) and at amino acids level (97.74-98.69%) with reported sequences of Indian isolates suggesting their common origin and limited sequence divergence. The epitope analysis by SVMTriP identified two unique epitopes in our isolate, seven shared epitopes among Indian isolates and two shared epitopes among all isolates except Poland isolate which carried all distinct epitopes.

Evidence of genetic drift and reassortment in infectious bursal disease virus and emergence of outbreaks in poultry farms in India.

Recurrent outbreaks of infectious bursal disease (IBD) have become a burning problem to the poultry industry worldwide. Here, we performed genetic analysis of IBD virus (IBDV) field isolates from recent outbreaks in various poultry farms in India. The sequence analysis of IBDV VP2 hypervariable region revealed amino acid pattern similar to that of very virulent (222A, 242I, 253Q, 256I, 272I, 279D, 284A, 294I, 299S and 330S) and intermediate plus virulent (222A, 242I, 253Q, 256I, 272T, 279N, 284A, 294I, 299S and 330S) type whereas analysis of VP1 revealed presence of sequence similar to that of very virulent (61I, 145T) and unique (61I, 141I, 143D, 145S) type in field isolates. Among the eight field isolates, two isolates contained very virulent type VP2 and unique type VP1, three contained intermediate plus virulent type VP2 and unique type VP1 whereas five contained both VP2 and VP1 of very virulent type. The phylogenetic analysis based on VP2 nucleotide sequence showed clustering of all eight isolates close to known very virulent strains whereas based on VP1, five isolates formed unique cluster and three isolates were placed close to very virulent strains. The isolates forming unique VP1 cluster showed highest similarity with classical virulent IBDVs suggesting their possible evolution from segment B of non-very virulent IBDVs. Interestingly, these five isolates were responsible for outbreaks in four different farms located at three different geographic locations in India. These observations indicates genetic reassortment between segment A and segment B from co-infecting IBDV strains leading to emergence of very virulent strains and their widespread prevalence in Indian poultry farms. The presence of 272I and 279D in VP2 protein of five field isolates may explain possible cause of Gumboro intermediate plus vaccine failure in prevention of the outbreaks. However, mortality caused by other three strains which are antigenically similar to VP1 of intermediate plus vaccine strains could not be explained and the possible role of their unique VP1 in enhancing the pathogenesis needs to be investigated further.