摘要
目的探讨山东肾综合征出血热重疫区病人血清中HV分子生物学特征,同时寻找准确、简便、迅速的HV检测与分型方法,从而为制定防治决策提供科学根据。方法从山东肾综合征出血热重疫区临沂市费县收集病人早期血清,应用巢式RT-PCR对病人血清中的HV进行基因扩增,采用RFLP、SSCP分型,并进行序列测定与分析。结果48份临床和ELISA检测确诊的HFRS病人血清经巢式RT-PCR扩增后,41份阳性,阳性率85.42%。41份阳性标本巢式RT-PCR产物经HindIII、HinfI酶切后,呈现2种不同的RFLP图谱:33份显示与R22株相似的酶切图谱,应属SEOV型;另外8份显示出与HTN76-118株相似的图谱,属HTNV型,这8份HTNV型标本均为10-12月间收集的病人血清。41份阳性标本巢式RT-PCR产物经SSCP分析,亦呈现2种不同的图谱:33份具有与R22株相似的SSCP图谱,应属SEOV型;而另8份则与HTN76-118株具有相似的SSCP图谱,属HTNV型。对部分扩增产物序列采用系统进化树分析也得出类似的结果:sdp1、sdp2、sdp3与HTN型76-118株亲缘关系相近,属同一簇,而sdp22、sdp37与SEOV型Z37、R22株亲缘关系相近,属另外一簇,这与RFLP和SSCP获得的结果相一致。结论山东肾综合征出血热重疫区病人基因型以SEOV型为主,但在秋冬季节也存在HTNV型。巢式RT-PCR结合RFLP、SSCP法可对HV准确分型,而且简便、迅速,适合于大规模流行病学调查。
In order to identify the characteristics of Hantaan virus （HV） in sera of hemorrhagic fever with renal syndrome （HFRS） patients and to find out the rapid method for its genotyping , the nested RT-PCR was used to amplify the Sgene fragment of HV, integrated with the use of RFLP to analyse their restriction maps of the nested RT-PCR products of Sgene and SSCP for the rapid analysis of genotypes of HV. In the present investigation, out of 48 serum samples of patients with HFRS, 41 samples were found to be positive in the nested PCR assay with a positive rate of 85.42%. In these 41 positive samples, 33 samples showed the similar RFLP maps with the R22 strain of HV, thus belonging to the SEOV type. and the other 8 samples （all were collected during November to December） had the similar RFLP maps with HTN 76-118 strain of HV, belonging to the HT-NV type, Similarly, these 41 samples appeared to have two different SSCP maps, of which 33 samples had the same SSCP maps as that of the reference strain R22, and the other 8 samples showed the same SSCP maps pattern with the reference strain HTN 76-118. Similar results were also obtained as adopted with the Systemic phylogenetic tree analysis for the partial sequences of amplified products ,that sdp1, sdp2 and sdp3 were very close to the HTNV type 76-118 strain, while sdp22 and sdp37 were very close to SEOV type Z37,R22 strain, locating in two separate clustersin the phylogenetic tree. These results were identical to those obtained by RFLP and SSCP. It is concluded that the genotypes of Hantaan virus in sera from HFRS patients in Shan- dong province mainly belong to the SEOV type, while the HTNV type of virus exists mainly in autumn and winter season. It suggests that the methods used in the present study are rapid, simple and accurate approaches for the genotype analysis of Hantaan virus.
出处
《中国人兽共患病学报》
CAS
CSCD
北大核心
2006年第7期624-628,共5页
Chinese Journal of Zoonoses
基金
山东省医药卫生重大创新研究计划课题(No.CX02207)
北京市自然科学基金重点项目(No.7021004)
济南军区军事医学研究所所长基金资助(No.200202)。
