2009年重庆地区手足口病病毒分离鉴定  

Isolation and identification of viruses from patient with hand,foot and mouth disease in Chongqing area in 2009

作　　者：蔡丽君[1] 王均[1] 常国辉[2] 许红梅[1] 

机构地区：[1]重庆医科大学附属儿童医院感染消化科,重庆400014 [2]军事医学科学院病原微生物流行病研究所、病原微生物生物安全国家重点实验室,北京100071

出　　处：《重庆医科大学学报》2010年第4期594-597,共4页Journal of Chongqing Medical University

基　　金：病原微生物生物安全国家重点实验室开放基金(编号:SKLPBS0919)

摘　　要：目的：对2009年2-4月重庆地区流行的手足口病（Hand-food-and-mouth disease,HFMD）病原进行分离鉴定,以调查本地区HFMD流行病原学情况。方法：采集47例手足口患儿脑脊液、疱疹液、粪便、咽拭子、肛拭子共100份标本,分别接种于人横纹肌肉瘤细胞（Human rhabdomyosarcoma,RD）中进行病毒分离培养,对出现细胞病变（Cytopathic effect,CPE）的细胞上清,分别用肠道病毒通用引物、EV71属特异性引物和柯萨奇A16属特异性引物进行RT-PCR检测,并对PCR产物进行回收克隆测序鉴定,以确定本次疫情中肠道病毒（Enteroviruses,EV）的病原,将测序结果用生物学软件Bioedit同国际代表BrCr株进行序列比对及同源性相关分析,以鉴定此毒株遗传变异情况。结果：在处理后的100份HFMD患儿临床标本的细胞培养中,有份观察到明显的细胞病变,通过RT-PCR检测有7份呈肠道通用引物阳性,EV71特异性引物检测有3份呈阳性结果,而CA1特异性引物检测结果均为阴性,这3份EV71阳性临床标本来自2例门诊患儿和1例临床诊断疑似病例患儿,其PCR产物测序结果也证实为EV71病毒。结论：重庆地区2-4月发生的HFMD疫情主要是由EV71型引起的。Objective：Toinvestigate the etiological agents of the outbreak ofhand-foot-mouth disease（HFMD）in ChongQing area from April to September in 2009.Methods：100 specimens（including cerebral spinal fluid,vesicle fluid,stool,throat swabs,anal swab） were collected from 47 patients with HFMD during the epidemic season.The specimens were inoculated into RD cells for isolation of enteroviruses（EV）,The supernatant of cytopathic effect（CPE）were detected by RT-PCR with universal primers of enterovirus, genus-specific primers of EV71 and genus-specific primers of Coxsackie A16,and the amplified products were sequenced to determine the outbreak of the pathogen enteroviruses（EV）,The sequencing results will be compared with international representatives BrCr strain to identify the enteroviruses（EV）by Bioedit biological software.Results：In the 100 cell cultures of clinical specimens of children HFMD, apparent Cytopathic were observed in 7.The results of RT-PCR showed that seven specimens were positive with intestinal universal primers,and three were positive results with EV71-specific primers,which were from two cases of out-patient children and one with clinical diagnosis of suspected case,while the negative results were obtained with CA16-specific primers for all the samples.The recovery of the PCR products were cloned and the sequencing identification confirmed as EV71 virus,while sequencing analysis of homology-related in the results.Conclusion：EV71 were the major etiological agents of the HFMD outbreak in Chongqing from February to April in 2009.

关 键 词：手足口病 肠道病毒71型 分离鉴定 

分 类 号：R72[医药卫生—儿科]