采用pcr-测序法对试验动物汉坦病毒的检测和基因分型.docVIP

PCR-测序法对啮齿类动物汉坦病毒的检测和基因分型戴方伟1，宋晓明2，1，周莎桑1，萨晓婴11 (1 浙江省医学科学院实验动物中心, 杭州 310013; 2 杭州师范大学实验动物中心, 杭州 310036) 【摘要】目的 探讨PCR-测序法在啮齿类动物汉坦病毒临床检测中的应用价值。方法 本文以Genebank7种血清亚型24株汉坦病毒代表性毒株为基础，从病毒基因S片段序列设计引物，采用邻位相连法进行系统进化分析，以该方法对浙江省近年从野生啮齿类动物中临床分离的汉坦病毒毒株进行分型鉴定。结果 所建系统发育树将所分析的毒株分为5个区域，引起HFRS的四种血清型具有较稳定的拓扑结构，且能与引起HPS的血清型进行区分。11株实验毒株进行PCR扩增和测序，结果表明该引物具有高度的敏感性和特异性，其中9株浙江省分离的血清型未知毒株的系统发育分析发现其包含引起HFRS的3株HTN和1株SEO血清型，其它5株属两种未知血清型。讨论 提示本文所建立的PCR-测序法具有用于临床检测汉坦病毒的价值。 【关键词】汉坦病毒；啮齿动物；测序；分型 Diagnosis and genetyping of hantan virus separated from rodent animals by direct PCR sequencing DAI Fang-wei1, SONG Xiao-ming2, LU Ling-qun1,ZHOU Sa-sang1, SA Xiao-yin1*,LV Yu1 (1 Center of Laboratory Animals, Zhejiang Academy of Medical Sciences, Hangzhou 310013, China; 2 Center of Laboratory Animals, Hangzhou Normal University, Hangzhou 310036, China)( [Abstract] Objective To evaluate the application value of PCR- sequencing in clinical detection of Hantan virus in rodent animals. Methods Based on 7 subtype 24 strains of hantavirus representative strains downloaded from Genebank, the virus S gene fragment were used for primer design and the neighbor joining method was applied for phylogenetic analysis. Thereafter, we identified clinical isolates from wild rodents of Hantan virus in Zhejiang province by this method. Results 24 analyzed strains were divided into 5 regions from the phylogenetic tree, four of them with topology structure, more stable. Then eleven strains of the virus was amplified by PCR and sequenced, and the results showed the primers with high sensitivity and specificity. Three HTN strains and 1 strain of serotype SEO was distinguished from 9 strains of unknown strains isolated in Zhejiang province, we also found 5 strains of Hantan virus belonging to two unknown serotype. Discussion the results suggest that PCR- sequencing method proposed in this paper could be used for clinical detection of Hantan virus. [Key word] Hantan virus; rodent animal; sequencing; genetyping 汉坦病毒（HV）属布尼亚病毒科，是一种有包