Differentiation of Sheep Pox and Goat Poxviruses by Sequence Analysis and PCR-RFLP of P32 Gene

Abstract

Sheep pox and Goat pox are highly contagious viral diseases of small ruminants. These diseases were earlier thought to be caused by a single species of virus, as they are serologically indistinguishable. P32, one of the major immunogenic genes of Capripoxvirus, was isolated and Sequenced from two Indian isolates of goat poxvirus (GPV) and a vaccine strain of sheep poxvirus (SPV). The sequences were compared with other P32 sequences of capripoxviruses available in the database. Sequence analysis revealed that sheep pox and goat poxviruses share 97.5 and 94.7% homology at nucleotide and amino acid level, respectively. A major difference between them is the presence of an additional aspartic acid at 55th position of P32 of sheep poxvirus that is absent in both goat poxvirus and lumpy skin disease virus. Further, six unique neutral nucleotide substitutions were observed at positions 77, 275, 403, 552, 867 and 964 in the sequence of goat poxvirus, which can be taken as GPV signature residues. Similar unique nucleotide signatures could be identified in SPV and LSDV sequences also. Phylogenetic analysis showed that members of the Capripoxvirus could be delineated into three distinct clusters of GPV, SPV and LSDV based on the P32 genomic sequence. Using this information, a PCR-RFLP method has been developed for unequivocal genomic differentiation of SPV and GPV.

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“…The GTPVs, and to a lesser extent the LSDV, lineages show more diversity than in the SPPV group. This observation was also observed previously for the P32 gene, but using a very limited number of isolates (Hosamani et al, 2004). Our GTPVs fall into three subgroups.…”

“…This study shows that GTPV and LSDV are more closely related to each other than to SPPV. This would support the hypothesis that they both emerged from a common ancestor close to the SPPVs, as proposed by others who carried out phylogenetic studies on different genome segments (Hosamani et al, 2004;Stram et al, 2008). This assumption is, however, in contradiction with another study, which concluded that small ruminant poxviruses may have emerged from a common LSDV-like ancestor, based on the observation in GTPV and SPPV of disrupted LSDV genes with putative virulence and host range functions (Tulman et al, 2002).…”

“…We even observed more divergence within each lineage than shown previously (Hosamani et al, 2004;Stram et al, 2008), thus identifying clearly separated sublineages with high bootstrap support. The multiple sequence alignments of the GPCR gene and the corresponding phylogenetic tree did not reveal specific differences between the vaccine and virulent strains of GTPV, LSDV and SPPV.…”

“…Our GTPVs fall into three subgroups. For LSDV, the subdivision into subgroups 1 and 2 was also supported by the work on P32 for our strains LSDV2, 3 and 4 (Hosamani et al, 2004).…”

“…Indeed, the rational design of CaPV subspecies primers in this region may permit the development of conventional and real-time PCR allowing the direct differential detection of the three CaPVs. Such a tool would be more robust and less sensitive to genetic drift than the two conventional PCRs described before, which both rely on subsequent analysis of the restriction profiles of the PCR products (Heine et al, 1999;Hosamani et al, 2004). This tool would also supersede a recently developed conventional PCR for LSDV, which does not allow the detection of SPPV (Stram et al, 2008).…”