Isolation, Identification Of Canine Parvovirus And Phylogeny Analysis Of The VP2Gene Sequences

Canine parvovirus enteritis which caused by canine parvovirus (canine parvovirus, CPV) isan acute, highly contagious disease of dogs and carnivores, and is one of most important diseasein pets, army dogs, patrol dogs. CPV was first isolated from dogs with diarrhea disease in1978by the scholars of Australian and Canadian, and then was widely spread in the world. In recentyears, with the variation of key amino acids of CPV capsid protein, new antigenic variants(CPV-2a, CPV-2b, CPV-2c three kinds of subtypes) was appeared constantly, and graduallyreplace the original CPV-2subtype become epidemic strain.The isolation and phylogenetic analysis of prevalent strain of canine parvovirus is of greatsignificance for understanding the epidemic, variation of CPV and development of new vaccines.In this study, tissue samples of canine from different regions of Hebei, Shanghai, Jilin provinceon2013were detected by PCR with specific primers of CPV, of which12samples were positive.The virus was isolated by inoculation of the homogeneous of positive samples into feline kidneycells (F81). The biology, morphology, physical and chemical characterization of the isolatedviruses were identified by HA/HI test, transmission electron microscopy and sequencing. It wasshowed that9strain canine parvoviruses were successfully isolated, and the virus could induceCPE in F81. The virus particles are cubic symmetry, with the diameter of20-24nm, no envelope,can agglutinate red blood cells of pigs, and could be inhibited by positive sera of canineparvovirus.The sequencing results of VP2gene showed that the length of VP2gene of9strains CPVwere both1755bp, encoding584amino acids, of which6strains belong to CPV-2a subtype,3strains belong to CPV-2b. Among the9strains of CPV, nucleotide homology of VP2gene was99.3%-99.9%, the homology of amino acid sequence was99.3%-100%. Homology in nucleotidewas98.8%-99.9%, and the homology of amino acid was98.6%-100%between domestic andforeign CPV isolates. Phylogenetic analysis of the isolated strains compared with the referencestrains showed that the genetic evolutionary relationship was closed to Chinese isolated strain,and far from the isolates from other countries. This study provides the experimental basis for thefurther study of epidemiology and genetic evolution of canine parvovirus in China.