貉源犬瘟热病毒的分离鉴定

Isolation and Identification of Canine Distemper Virus from Raccoon Dog

采集大庆某貉养殖场病貉的病料,处理后接种于非洲绿猴肾细胞(Vero)进行病毒分离。对分离毒株进行了中和试验、血凝试验、理化性质的鉴定,并用反转录-聚合酶链式反应(RT-PCR)检测感染细胞中的病毒核酸。病料接种Vero细胞72h后产生明显的细胞病变(CPE);病毒分离株可以被犬瘟热病毒阳性血清中和,中和效价为1∶25;分离株对氯仿、乙醚敏感,对酸和热抵抗力弱,可以凝集鸡红细胞,其凝集作用可被犬瘟热病毒阳性血清所抑制;RT-PCR检测病毒细胞培养液,扩增出的片段长287bp,与预期设计的长度相同,经测序发现与MS01株的同源性为99%。结果表明,分离的病毒株为犬瘟热病毒,命名为CDV-DQ株。

Raccoon dog specimen from daqing were collected for virus isolation in vero cell culture. The isolated virus was confirmed by a series of systemic identification, such as serology test , physics and chemistry characteristics and molecular virology test. The results showed that the specimens could cause cytopathogenic efect （CPE） in vero cell after subculturing 72 h. The isolated virus could be neutralized with positive canine sera of CDV with the neutralization titer of 1 ： 2^5. The agglutination titer of isolated virus was 1 ： 16 for the chicken erythrocytes. The isolated virus was sensitive for chloroform, aether and acid. It was completely inactivated at 60 ℃ within 1 h. A DNA fragment of 287 bp was amplified from infected vero cells by RT- PCR. The agreement of N gene fragment sequence between the isolated virus and CDV MS01 from GenBank was 99%. From the results described above,the isolated virus was CDV and designated as CDV-DQ.