犬细小病毒吉林流行株的分离鉴定及VP2基因的遗传进化分析

Isolation and Identification of Canine Parvovirus Strain in Jilin Province and Genetic Evolution Analysis of VP2 Gene

从吉林省长春市某动物医院采集了3份CPV阳性宠物犬粪便样品,经PCR鉴定,使用F81细胞进行盲传,同时对病毒进行血凝、电镜观察及间接免疫荧光等检测后,对其VP2基因进行序列分析。结果显示,3株CPV分离株为CPV-2c型犬细小病毒(命名为CPV-JL21-L1、CPV-JL21-L2、CPV-JL21-L3),第20代病毒滴度均在10^(-7)~10^(-7.5)TCID50/0.1 mL之间,第20代血凝效价结果均保持在210~211。VP2基因序列遗传进化分析显示,3株CPV分离株与3株疫苗株VP2基因的同源性为98.6%~99.0%,与其他参考毒株同源性为98.6%~99.9%。此外,分离株CPV-JL21-L1和CPV-JL21-L3在80 L、101 T、267 Y、297 A、300 G、324 I、375 D、426 E和555 V位点与国内流行株相同,CPV-JL21-L2在267位点氨基酸发生了变异。本研究为吉林省CPV分子流行病学的调查提供理论依据。

In this study,three pieces of CPV positive fecal samples were collected from pet dogs in an animal hospital in Changchun City of Jilin Province.The positive results were confirmed by the PCR assay,and experienced blind passaged in the F81 cell line for CPV isolations.The VP2 gene sequences of the isolated CPV strains were analyzed with the hemagglutination(HA)tests and indirect immunofluorescence.The results showed that the three CPV isolates were CPV-2c type(named CPV-JL21-L1,CPV-JL21-L2,CPV-JL21-L3,respectively).The HA titers of the 20th CPV isolates were between 210 to 211,and the corresponding virus titers were 10^(-7)to 10^(-7.5)TCID_(50)/0.1 mL.The genetic evolution analyses based on VP2 genes demonstrated that the three CPV isolates shared 98.6%~99.0%and 98.6%~99.9%homology with three vaccine strains and other referred strains,respectively.In addition,the CPV-JL21-L1 and CPV-JL21-L3 share same site at80 L,101 T,267 Y,297 A,300 G,324 I,375 D,426 E and 555 V with epidemic strains in domestic and CPV-JL21-L2 showed anmio acid variation at 267 site.This study would provide an important theoretical basis for the recently CPV epidemiology study in Jilin Province.