甘肃省鼠疫耶尔森菌多位点可变数目串联重复序列基因分型及地区分布

Genotyping by MLVA reveals the regional distribution of Yersinia pestis in Gansu Province

目的采用多位点可变数目串联重复序列(MLVA)对甘肃省鼠疫耶尔森菌(鼠疫菌)进行基因分型,探讨甘肃省鼠疫菌地区分布特征。方法选取1962-2014年间甘肃省各市县(区)不同生态型的鼠疫菌198株,培养并提取基因组DNA。采用14+12对引物进行PCR扩增,产物进行测序,确定每对引物串联重复序列(VNTR)位点的拷贝数。应用BioNumerics 5.10软件对菌株的VNTR位点拷贝数进行聚类分析。结果甘肃省鼠疫菌分为10个群,群内的菌株依据分离地点继续细化为5-28个分支。10个群分别为:阿拉善黄鼠疫源地菌株聚为1个群;甘南高原疫源地菌株聚成1个群;阿克塞县菌株分成2个群,为当金山群和加尔乌宗村群;肃北县菌株分为3个群,为马场群、党城湾镇群和鱼儿红群;玉门市菌株聚集在肃北县鱼儿红群内;肃南县菌株分为3个群,为大河乡群、马蹄乡群和康乐乡群。结论MLVA可以清晰地区分甘肃省不同鼠疫疫源地的菌株,将疫源地内菌株继续细分为不同分支,直至精确的分离地点。MLVA分型方法可用于鼠疫菌株的溯源分析。

Multiple variable number tandem repeat(VNTR)analysis(MLVA)was used to study the genotypes and distribution of Yersinia pestis in Gansu province.Different ecotypes of Yersinia pestis from 1962 to 2014 were selected for genotyping by MLVA.PCR amplification was carried out at the 14+12 VNTR loci,and the amplicons were analyzed by capillary electrophoresis to determine the repeat number of each VNTR locus.The data were analyzed with BioNumerics 5.10.The strains of Y.pestis in Gansu province were divided into ten genotypes.The strains from different foci clustered into different types,which correlated well with the isolation sites.According to the isolation sites,the strains in the types were further subdivided into 5-28 subbranches.The ten genotypes included strains from the Spermophilus alaschanicus plague focus in one type;those from the Gannan Plateau plague focus in one type;those from Akesai County in two types;i.e.,those from Dangjinshan and Jiaerwuzong village;those from Subei County in three types;i.e.,the types from Machang,Dangchengwan Town and Yuerhong;those from Yumen City,which shared the same type as those from Yuerhong in Subei County;and those from Sunan County in three types;i.e.,types from Dahe Town,Mati Town and Kangle Town.The MLVA(14+12)scheme can be used to distinguish strains from different plague foci in Gansu province,and the genotypes correlate well with the plague bacteria isolation sites.This method can be used for source tracing of Yersinia pestis.