Sequence analysis of minor protein genes of equine arteritis virus during persistent infection

The variability of the ORF2a, ORF2b, ORF3, and ORF4 genes of the equine arteritis virus (EAV) was analysed during a seven year observation of persistent infection in a stallion of the Malopolska breed. A total of 11 semen samples were collected between 2004 and 2011. RNA of EAV isolates obtained from the semen of the stallion was amplified, sequenced, and compared with the sequences of other strains available in GenBank. Multiple nucleotide substitutions were found in sequences of the analysed regions, however, neither deletion nor insertions were detected. The highest number of point mutations (11-6 synonymous and 5 non-synonymous) were found in the ORF2b gene, and the lowest number of substitutions (6-5 synonymous and one non-synonymous) were found in the ORF2a gene. None of the identified mutations affected any of the glycosylation or phosphorylation sites of the minor EAV protein. Phylogenetic analysis of the ORF3 gene of EAV isolates showed that they grouped together within the cluster of European strains of EAV. Additionally, the ORF3 gene sequences of the isolates showed high (86.4% - 98.3%) similarity to the previously isolated Polish EAV strains.