猪繁殖与呼吸综合征商品活疫苗中盖他病毒的分离与鉴定  

Isolation and Identification of the Getah Virus from the Contaminated PRRS Commercial Live Vaccine

作　　者：周峰[1] 王傲杰 常洪涛[1] 王新港[1] 崔丹丹[1] 陈陆[1] 杜季梅[1] 刘红英[1] 杨霞[1] 王新卫[1] 王川庆[1] ZHOU Feng;WANG Aojie;CHANG Hongtao;WANG Xingang;CUI DANDan;CHEN Lu;DU Jimei;LIU Hongying;YANG Xia;WANG Xinwei;WANG Chuanqing(College of Animal Science and Veterinary Medicine,Henan Agricultural University, Zhengzhou 450002,China)

机构地区：[1]河南农业大学牧医工程学院,郑州450002

出　　处：《病毒学报》2019年第3期486-496,共11页Chinese Journal of Virology

基　　金：河南省高等学校重点科研项目计划(项目号:18A230008);题目:猪盖塔病毒分离培养平台的建立与在疫苗研究中的应用;国家自然科学基金(项目号:3177131377);题目:伪狂犬病病毒编码prv--miR--LLT11a靶向调节SLA-I机制及免疫学效应~~

摘　　要：在疫苗生产过程中,外源病毒造成疫苗污染的现象屡有发生。本文在对某猪场的猪繁殖与呼吸综合征(Porcine reproductive and respiratory syndrome, PRRS)减毒活疫苗进行外源病毒检测时,发现盖他病毒(Getah virus,GETV)为阳性。经病毒分离、纯化及鉴定,获得一株GETV分离物,将其命名为GETV-V1。GETV-V1与GenBank中GETV全基因序列相似性为97.3%～99.2%。遗传进化分析显示,GETV-V1与日本毒株14-I-605-C1、12IH26及中国毒株HB0234等处于同一进化分支,与本室分离的猪源GETV毒株HNJZ-S1的全基因序列(KY363862)相似性为99.1%,两者E2基因推导氨基酸序列相似性为99.8%。对同一厂家4批次市售同种疫苗和相应批次留样疫苗RT-PCR检测均为GETV阳性,第三方实验室和疫苗生产厂家分别独立进行的RT-PCR检测得出同样结果。测序结果表明,所有被测样品中E2基因与GETV已知相应序列的相似性均在98%以上。本研究首次报道了我国PRRS减毒活疫苗中GETV的污染,不仅表明猪用活疫苗外源病毒监测标准有待改进,而且提示要重视该病毒污染在疫苗生产、保存和使用过程中潜在的公共卫生风险。Contamination occurs frequently during the development and production of vaccines.We detected exogenous viruses of a live attenuated porcine reproductive and respiratory syndrome virus(PRRSV)vaccine from a pig farm using reverse transcription-polymerase chain reaction(RT-PCR)and found that the vaccine was contaminated by the Getah virus(GETV).The virus was isolated,passaged,purified by plaques and named"GETV-V1".According to complete-genome analysis,the identity of the sequence between GETV-V1 and reference sequence(GenBank)was 97.3%~99.2%,and the identity was the highest with the Korean pig strain AY702913(Accession number:AY702913).Genetic-evolution analysis showed that GETV-V1 was closely related to the Japanese strains 14-I-605-C1,14-I-605-C2 and 12 IH26,and Chinese strain HB0234,and was in the same evolutionary branch.The identity of the whole gene sequence between the porcine GETV strain HNJZ-S1(Accession number:KY363862,previously isolated by our lab)and GETV-V1 was 99.1%,while E2-gene deduced amino-acid sequence analysis showed that the identity was 99.8%.The E2 gene of the GETV was detected by RT-PCR in four batches of the vaccine derived from the same manufacturer.All specimens of different batches of the same vaccine derived from the manufacturer above were GETV-positive as well when tested by using RT-PCR conducted by a third-party laboratory and vaccine manufacturer’s lab.Sequencing of E2 gene amplified by RT-PCR showed that the identity of sequences between the strains detected in present study and published strains of GETV was>98%.In this study,the contamination of a live attenuated PRRS vaccine with GETV was reported for the first time in China and the results indicated that the monitoring method for exogenous viruses of live vaccines for pigs needs to be improved and people should pay more attention to the effect of live vaccine contaminated with GETV on the potential public health risk.

关 键 词：猪繁殖与呼吸综合征(PRRS)疫苗 盖他病毒(GETV)污染 分离鉴定 RT-PCR 透射电镜(TEM) 间接免疫荧光试验(IFA) 序列分析 公共卫生安全 

分 类 号：S852.65[农业科学—基础兽医学] S855[农业科学—兽医学]