Comparison of four regions in the replicase gene of heterologous infectious bronchitis virus strains

Infectious bronchitis virus (IBV) produces six subgenomic (sg) mRNAs, each containing a 64 nucleotide (nt) leader sequence, derived from the 5′ end of the genome by a discontinuous process. Several putative functional domains such as a papain-like proteinase (PL(pro)), main protease (M(pro)), RNA-dependent RNA polymerase (RdRp), and RNA helicase encoded by the replicase gene are important for virus replication. We have sequenced four regions of the replicase genes corresponding to the 5′-terminal sequence, PL(pro), M(pro), and RdRp domains from 20 heterologous IBV strains, and compared them with previously published coronavirus sequences. All the coronavirus 5′-termini and PL(pro) domains were divergent, unlike the M(pro) and the RdRp domains that were highly conserved with 28% and 48% conserved residues, respectively. Among IBV strains, the 5′ untranslated region including the leader sequence was highly conserved (>94% identical); whereas, the N-terminal coding region and the PL(pro) domains were highly variable ranging from 84.6% to 100%, and 77.6% to 100% identity, respectively. The IBV M(pro) and RdRp domains were highly conserved with 82.7% and 92.7% conserved residues, respectively. The BJ strain was the most different from other IBVs in all four regions of the replicase. Phylogeny-based clustering based on replicase genes was identical to the antigen-based classification of coronaviruses into three groups. However, the IBV strain classification based on replicase gene domains did not correlate with that of the type-specific antigenic groups. The replicase gene sequences of many IBVs recovered from infected chickens were identical to those of vaccine viruses irrespective of serotype, suggesting that either there has been an exchange of genetic material among vaccine and field isolates or that there is a convergent evolution to a specific replicase genotype. There was no correlation between the genotype of any region of the replicase gene and pathotype, suggesting that the replicase is not the sole determinant of IBV pathogenicity.