犬细小病毒CPV-JZ2021株的分离鉴定及遗传进化分析

Isolation and identification of canine parvovirus CPV-JZ2021 strain and it’s analysis of genetic evolution

为研究犬细小病毒的生物学特性和遗传进化情况,从湖北馨爱宠物医院有限公司收集疑似病犬的粪便,无菌处理病料后接种猫肾细胞(CRFK)进行病毒的培养,至出现明显的细胞病变。通过PCR扩增、电镜形态学观察、间接免疫荧光检测、血凝试验等进行病毒鉴定,结果表明,成功分离出一株犬细小病毒,并命名为CPV-JZ2021。该病毒的血凝效价为2^(9),TCID_(50)为10^(3.4)/0.1 mL。全基因组测序结果表明,该病毒全基因组序列长4567 bp。构建的系统发育树表明,该病毒与四川、河南、广东和北京等地毒株亲缘关系较近,处于同一进化分支。其中与河南株CPV-HN1587的同源性达99.2%。通过对VP2基因推断的氨基酸序列进行分析,确定该犬细小病毒属于New CPV-2a型。相比较国外已经报道的New CPV-2a型毒株,CPV-JZ2021存在多个关键氨基酸变异。该研究为犬细小病毒的流行情况及新疫苗的开发提供了参考依据。

In order to study the biological characteristics,and genetic evolution of canine parvovirus,the feces of suspected dogs were collected from Hubei Xin-ai Pet Hospital Co.Ltd..After sterility treatment,CRFK cells were inoculated with virus culture until obvious cytopathic changes appeared.The virus was verified by PCR,electron microscopy morphological observation,indirect immunofluorescence test and hemagglutination test.The results showed that a canine parvovirus was successfully isolated and named CPVJZ2021.The hemagglutination titer of the virus is 2^(9),and the TCID_(50) is 10^(3.4)/0.1 mL.Whole-genome sequencing showed that the length of the whole-genome sequence was 4567 bp.The phylogenetic tree showed that the isolated strain was closely related to strains from Sichuan,Henan,Guangdong and Beijing,and belonged to the same evolutionary branch.The homology with CPV-HN1587 is 99.2%.The canine parvovirus belongs to the New CPV-2 a type according to amino acid sequence of the VP2 gene.Compared with the New CPV-2 a strains that have been reported abroad,CPV-JZ2021 has many key amino acid variations.This study provides a reference for the prevalence of canine parvovirus and the development of new vaccines.