Analysis of the complete genomic sequences of two virus subpopulations of the Australian infectious bronchitis virus vaccine VicS

Abstract

Although sequencing of the 3′ end of the genome of Australian infectious bronchitis viruses (IBVs) has shown that their structural genes are distinct from those of IBVs found in other countries, their replicase genes have not been analysed. To examine this, the complete genomic sequences of the two subpopulations of the VicS vaccine, VicS-v and VicS-del, were determined. Compared with VicS-v, the more attenuated VicS-del strain had two nonsynonymous changes in the non-structural protein 6 (nsp6), a transmembrane (TM) domain that may participate in autocatalytic release of the 3-chymotrypsin-like protease, a polymorphic difference at the end of the S2 gene, which coincided with the body transcription-regulating sequence (B-TRS) of mRNA 3 and a truncated open reading frame for a peptide encoded by gene 4 (4b). These genetic differences could be responsible for the differences between these variants in pathogenicity in vivo, and replication in vitro. Phylogenetic analysis of the whole genome showed that VicS-v and VicS-del did not cluster with strains from other countries, supporting the hypothesis that Australian IBV strains have been evolving independently for some time, and analyses of individual polymerase peptide and S glycoprotein genes suggested a distant common ancestor with no recent recombination. This study suggests the potential role of the TM domain in nsp6, the integrity of the S2 protein and the B-TRS 3, and the putative accessory protein 4b, as well as the 3′ untranslated region, in the virulence and replication of IBV and has provided a better understanding of relationships between the Australian vaccine strain of IBV and those used elsewhere.

Full genome analysis of Australian infectious bronchitis viruses suggests frequent recombination events between vaccine strains and multiple phylogenetically distant avian coronaviruses of unknown origin

“…Phylogenetic analysis of different strains of IBVs isolated from different countries (Fig. 1) supported previous suggestions that Australian strains of IBV have been diverging as an independent cluster (Quinteros et al, 2015). However, the representation of genetic divergence by a phylogenetic tree alone may not be accurate, because once a branch is created no further potential interactions between the branches are represented (Thor et al, 2011).…”

“…The IBV Network Splitstree (Fig. 1) suggested considerable intra-clade interactions, with a significantly lower number of inter-clade interactions, supporting suggestions that Australian IBVs have been genetically isolated for a considerable time (Ignjatovic et al, 2006;Quinteros et al, 2015). This could be explained by the geographic location of Australia as an island and the enclosed nature of the Australian ecosystem, and also by the biosecurity barriers imposed by Australian authorities on biological imports.…”

“…RNA was fragmented using RNase III (Life Technologies) and purified using magnetic beads. To construct the libraries, the RNA was reverse transcribed and the cDNA purified and amplified following the manufacturer's protocols, and then sequenced (Quinteros et al, 2015). Sequencing was performed using the PGM Ion Torrent TM platform (Life Technologies), with a 314 chip and the 200-base sequencing Ion OneTouch Kit v2.…”

“…A multiple sequence alignment of the complete genome sequences of Australian and international isolates of IBV was constructed with Clustal-Omega (Sievers et al, 2011), using the default settings. This included previously determined genomic sequences of two variants of the VicS vaccine strain (Quinteros et al, 2015). This was used to prepare a network tree using SplitsTree4 (Huson and Bryant, 2006), version 4.14.3, using the maximum likelihood (F84) distance model, the NeighborNet network algorithm, 1000 bootstrap replicates and an estimated proportion of invariant sites of 0.37.…”

Analysis of antigenicity and pathogenicity reveals major differences among QX-like infectious bronchitis viruses and other serotypes

“…As we know, the pathogenicity of IBV is determined by multiple factors, which may involve its tissue tropism, replication efficacy and its ability to deal with the host's immune system. The spike protein was shown to be an important virulence determination of IBV and some other protein genes have also been shown to exhibit obvious influence to IBV virulence (Armesto et al, 2009;Casais et al, 2003;Quinteros et al, 2015;Wickramasinghe et al, 2011). So the specific amino acids differences between them in the spike protein possibly contributed to the difference in the two viruses (SD and SZ).…”