Isolation,Identification,Phylogenetic And Pathogenicity Analysis Of Canine Parvovirus

Canine parvovirus(CPV-2)belongs to the Parvoviridae family and Parvovirus genus,which is an important canine pathogen causing canine parvovirus disease(CPVD)characterized by hemorrhagic enteritis and myocarditis in dogs.CPV-2 is believed to be a host range variant derived from carnivore parvoviruses such as feline panleukopenia virus(FPV)or FPV-like virus,which evolved via wild animals and acquired the ability to infect dogs.The fundamental measure to prevent CPVD lies in vaccination.However,CPV-2 is prone to gene mutation.At present,four antigenic variants have been found,namely CPV-2,CPV-2a,CPV-2b and CPV-2c.Although there were studies proving that existing vaccines can effectively protect against all antigenic variants,cases of immunization failure have been occasionally reported.The continuous evolution of CPV-2 may pose new challenges to the immune efficacy of existing vaccines.Therefore,it is necessary to continuously monitor the dynamic evolution of CPV-2 in Chins and pay attention to the possibility of immunization failure caused by emerging variants.This article studies and discusses CPV-2 from three aspects: The isolation and identification,the phylogenetic analysis and the pathogenicity analysis of CPV-2 strains.The isolation of virus is the basis of virological research and the development of new vaccines.Based on 36 CPV-2 positive or suspected samples collected from pet hospitals in Guangdong,Tianjin and other areas in China,24 strains were identified as CPV-2 by cytopathic observation,PCR detection,electron microscope observation,direct immunofluorescence staining and hemagglutination test.The results showed that these isolated strains have the typical characteristics of CPV-2,that is,it could proliferate on F81 cells and cause characteristic cytopathy such as cell elongation and shedding;the virus particles were round or polygonal with a diameter of about 22 nm;it could specifically bind the CPV-2 monoclonal antibody;it could agglutinate porcine erythrocytes.The VP2 gene extracted from 30 CPV-2 strains was sequenced.21 strains were identified as CPV-2c,7 strains were new CPV-2a and 2 strains were FPV according to the amino acid substitutions in the VP2 protein,indicating that CPV-2 has replaced new CPV-2a as the new dominant strain.Several novel substitutions were found in the VP2 protein,such as Ala5 Gly,Pro13Ser,Phe267 Tyr,Tyr324Ile and Gln370 Arg.The potential biological significance of these substitutions remains to be further elucidated.CPV-2 has been an important cause of morbidity and mortality in puppies since its emergence in 1978.Despite extensive vaccination,CPVD is still a common disease.CPVD is mainly divided into enteritis type and myocarditis type,and both types can be fatal while the curative effect is very limited.The pathology and the viral loads in tissues of two beagle dogs infected with CPV-2 were studied.The results showed that the pathological changes of these two infected dogs were mainly concentrated in the intestinal tract characterized by extensive necrosis and bleeding.Histopathological results showed the necrosis and abscission of intestinal villi,collapse or expansion of intestinal crypts,lymphopenia,and necrosis of cardiac myocytes;The immunohistochemical staining confirmed the distribution of CPV-2 in multiple important organs.By means of real-time fluorescent quantitative PCR(q PCR),CPV-2DNA was detected in all tissues examined,with the highest titres observed in the lymphoid tissue and the intestinal tissue,and the lowest loads in the muscle tissue.These results revealed that CPVD was a systemic disease,which could lead to multiple organ damages.Through isolation and identification,phylogenetic analysis and pathogenicity analysis of CPV-2,this study provides theoretical and experimental basis for better understanding the pathogenesis of CPV-2 and taking better preventive and therapeutic measures.