Adaptive Evolution of the Fox Coronavirus Based on Genome-Wide Sequence Analysis.

Purpose: To report the first complete fox coronavirus (CoV) genome sequence obtained through genome-wide amplifications and to understand the adaptive evolution of fox CoV. Methods: Anal swab samples were collected from 35 foxes to detect the presence of CoV and obtain the virus sequence. Phylogenetic analysis was conducted using MrBayes. The possibility of recombination within these sequences was assessed using GARD. Analysis of the levels of selection pressure experienced by these sequences was assessed using methods on both the PAML and Data Monkey platforms. Results: Of the 35 samples, two were positive, and complete genome sequences for the viruses were obtained. Phylogenetic analysis, using Bayesian methods, of these sequences, together with other CoV sequences, revealed that the fox CoV sequences clustered with canine coronavirus (CCoV) sequences, with sequences from other carnivores more distantly related. In contrast to the feline, ferret and mink CoV sequences that clustered into species-specific clades, the fox CoV fell within the CCoV clade. Minimal evidence for recombination was found among the sequences. A total of 7, 3, 14, and 2 positively selected sites were identified in the M, N, S, and 7B genes, respectively, with 99, 111, and 581 negatively selected sites identified in M, N, and S genes, respectively. Conclusion: The complete genome sequence of fox CoV has been obtained for the first time. The results suggest that the genome sequence of fox CoV may have experienced adaptive evolution in the genes replication, entry, and virulence. The number of sites in each gene that experienced negative selection is far greater than the number that underwent positive selection, suggesting that most of the sequence is highly conserved and important for viral survive. However, positive selection at a few sites likely aided these viruses to adapt to new environments.

Adaptive Evolution of Feline Coronavirus Genes Based on Selection Analysis.

PURPOSE: We investigated sequences of the feline coronaviruses (FCoV), which include feline enteric coronavirus (FECV) and feline infectious peritonitis virus (FIPV), from China and other countries to gain insight into the adaptive evolution of this virus. METHODS: Ascites samples from 31 cats with suspected FIP and feces samples from 8 healthy cats were screened for the presence of FCoV. Partial viral genome sequences, including parts of the nsp12-nsp14, S, N, and 7b genes, were obtained and aligned with additional sequences obtained from the GenBank database. Bayesian phylogenetic analysis was conducted, and the possibility of recombination within these sequences was assessed. Analysis of the levels of selection pressure experienced by these sequences was assessed using methods on both the PAML and Datamonkey platforms. RESULTS: Of the 31 cats investigated, two suspected FIP cats and one healthy cat tested positive for FCoV. Phylogenetic analysis showed that all of the sequences from mainland China cluster together with a few sequences from the Netherlands as a distinct clade when analyzed with FCoV sequences from other countries. Fewer than 3 recombination breakpoints were detected in the nsp12-nsp14, S, N, and 7b genes, suggesting that analyses for positive selection could be conducted. A total of 4, 12, 4, and 4 positively selected sites were detected in the nsp12-nsp14, S, N, and 7b genes, respectively, with the previously described site 245 of the S gene, which distinguishes FIPV from FECV, being a positive selection site. Conversely, 106, 168, 25, and 17 negative selection sites in the nsp12-14, S, N, and 7b genes, respectively, were identified. CONCLUSION: Our study provides evidence that the FCoV genes encoding replicative, entry, and virulence proteins potentially experienced adaptive evolution. A greater number of sites in each gene experienced negative rather than positive selection, which suggests that most of the protein sequence must be conservatively maintained for virus survival. A few of the sites showing evidence of positive selection might be associated with the more severe pathology of FIPV or help these viruses survive other harmful conditions.