Molecular characterization of the features of antigenic and pathogenic variant strains of infectious bronchitis virus.

摘要

Antigenic and pathogenic variants of avian infectious bronchitis virus (IBV), a coronavirus, were molecularly characterized to delineate the virus evolution. For rapid detection of IBV strains from field outbreaks, we describe a slot blot hybridization assay that can distinguish unknown IBV strains by serotype if they bear at least 95% sequence identity to a reference strain within the hypervariable regions of S1 gene. To elucidate the role of replicase gene for the emergence of antigenic variants, four functional regions in the replicase gene of heterologous IBV strains were sequenced, and compared with previously published IBV and other coronavirus sequences. Sequence analysis revealed that: (i) the main protease and RNA-dependent RNA polymerase regions are highly conserved among all coronaviruses unlike the 5' -terminal and papain-like proteinase regions, (ii) the replicase-based clustering of heterologous IBV strains does not correlate with antigen-based S1 phylogeny, (iii) several widely used IBV vaccines and field strains isolated over past decades have closely related replicase genes suggestive of a possible common ancestry or a converging evolution, and (iv) the replicase gene is unlikely to be the sole determinant of IBV pathogenicity. To delineate the origin of California 99 (Cal99) variant, the complete genome of virus was sequenced. Extensive sequence analysis of the structural protein genes revealed that the Cal99 virus is very similar to the ArkDPI strain, except in the S1 gene and stretches of sequence in the S2 and M protein genes. The Cal99 acquired new antigenic properties through extensive mutations in S1 gene. The virus most likely acquired two gene fragments, one in the 3 ' end of Cal99 S2 gene which is similar to Conn strain and another in the 5' end of M protein gene which is similar to Mass strain, by recombinations. Since Ark, Conn and Mass strains are used as live vaccines in California, we theorize that both point mutations and recombination among vaccine strains contributed to the emergence of Cal99 variant. Finally, we performed a challenge study with prototype Cal99 strain and demonstrated that the virus causes late-onset respiratory disease, with a severity comparable to that of M41 challenge strain. Based on these results we suggest that the variants may arise from IBV vaccines through genetic modifications, which may confer changes in pathogenicity and antigenicity.