中华按蚊源日本脑炎病毒的分离鉴定及其分子特征的分析  

Isolation,identification and molecular characteristic of a Japanese encephalitis virus isolate from Anopheles sinensis

作　　者：刘茜倩[1,2] 魏建超[1] 钟登科[3] 吴亚玲[1] 石坤[1] 张克龙[1] 陆美林[1] 肖长广 李蓓蓓[1] 刘珂[1] 邵东华[1] 邱亚峰[1] 石元元[1] 马志永[1] 范红结[2] 

机构地区：[1]中国农业科学院上海兽医研究所,上海200241 [2]南京农业大学动物医学院,江苏南京210095 [3]上海农林职业技术学院,上海201600

出　　处：《中国兽医科学》2017年第1期16-22,共7页Chinese Veterinary Science

基　　金：国家自然科学基金项目(31302116);国家国际科技合作专项(2014DFE30140);国家公益性行业(农业)科研专项(201203082)

摘　　要：为了解上海市蚊源日本脑炎病毒(JEV)的分子特征,2014年从上海市嘉定区猪场采集蚊媒样品1万余份,采用BHK21和C6/36细胞培养法进行病毒分离,通过RT-PCR和IFA技术对分离株进行鉴定。结果表明,分离到1株JEV,命名为JD-15,源自中华按蚊。用JD-15株分别感染BHK21和C6/36细胞,72 h后所有细胞均发生明显病变(CPE),其细胞半数感染量(TCID50)为1.96×106PFU。用分离株JD-15脑内接种3周龄乳鼠,结果测得半数致死量(LD50)为4.2×101PFU,表明该毒株具有较强的神经毒力。对JD-15株的E基因进行克隆测序及进化分析,结果显示,该株JEV属于基因Ⅲ型,与减毒活疫苗株SA-14-14-2的E基因核苷酸序列的同源性为98.1%,氨基酸序列的同源性为97.5%。本研究为乙型脑炎的流行病学调查提供了科学依据,对上海市蚊虫及蚊媒病的防治具有重要意义。To investigate the molecular characteristics of JEV in Shanghai from mosquitoes,more than ten thousand samples from mosquitoes were collected from Jiading District＇s pigpens in Shanghai 2014. BHK21 and C6/36 cell culture techniques were used for viral isolation. RT-PCR and IFA techniques were used for molecular identification. The results indicated that a new strain named JD-15 was isolated from Anopheles sinensis. All BHK21 cells and C6/36 cells showed cytopathic effects at 72 h after inoculation of the isolated strain. The titer was 1.96 ×101 PFU. The median lethal dose of the isolate measured by intracerebral injection in mice was 4.2 ×101 PFU,which indicated the isolate possessed strong neurovirulence. The cloning and sequence analysis based on the amplified E gene indicated that the isolate belonged to Genotype Ⅲ of JEV. Meanwhile,homology of the nucleotide and amino acid sequences between the isolated JD-15 and vaccine stain SA-14-14-2 was 98.1%and 97.5%,respectively. This study was to provide a scientific basis evidence for epidemiological survey of JEW disease. It will be of great significance for prevention and controlof Shanghai local mosquito and mosquito-borne diseases.

关 键 词：日本脑炎病毒 分离鉴定 进化分析 

分 类 号：S852.659.6[农业科学—基础兽医学]