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目的 探讨 RT- PCR用于 HFRS临床标本中病毒基因检测的若干影响因素及西安地区近年主要流行汉坦病毒(HV)毒株的分子流行病学特点 .方法 设计合成了两套分别互补于 HV M基因片段和 S基因片段的引物 ,建立了检测HFRS患者临床血清标本中 HV基因的 RT- nested PCR方法 ,并对扩增的部分 HV靶基因进行了测序分析和比较 .结果 119份 HFRS患者血清 (经临床和 IFA确诊 )经用 S基因片段引物的 RT- nested PCR检测 ,阳性率分别为 :6 0 .5 %(<1wk) ,34 .1% (1～ 2 wk) ,4.0 % (2～ 3wk) ,0 % (>3wk ) ,31份 HFRS急性期患者血清用 M基因片段引物的RT- nested PCR检测 ,仅 12份阳性 ,阳性率 40 .0 % .将 PCR扩增的 6个靶基因序列与 76～ 118株 S基因片段相应的核苷酸序列进行分析比较 ,同源性为 87.2 %～ 96 .0 % ;其推导的氨基酸序列与 76～ 118株核衣壳蛋白相应的氨基酸序列比较 ,同源性为 89.0 %～ 96 .0 % .结论 S基因片段引物的PCR扩增效果优于 M基因片段引物 .6份 S基因片段引物扩增产物的测序结果显示 ,西安地区引起 HFRS流行传播的主要毒株仍为汉滩型
Objective To investigate the influencing factors of viral gene detection in clinical specimens of HFRS by RT-PCR and the molecular epidemiological characteristics of the major epidemic Hantavirus (HV) strains in Xi’an area in recent years.Methods Two sets of complementary molecular hybrids RT-nested PCR method for detecting HV gene in clinical serum samples of HFRS patients was established and some of the amplified HV target genes were sequenced and compared.Results The serum level of 119 The results of RT-nested PCR using S gene fragment primers showed that the positive rates were 60.5% (<1wk), 34.1% (1-2wk), 4.0% (2 ~ 3wk), 0% (> 3wk), 31 patients with acute HFRS sera were detected by RT-nested PCR using M gene fragments, only 12 were positive and the positive rate was 40.0% Compared with the corresponding nucleotide sequence of 76-118 S gene fragments, the homology was 87.2% -96.0%. The deduced amino acid sequence was compared with the corresponding amino acid sequence of 76-118 nucleocapsid protein , The homology was 89.0% ~ 96.0% .Conclusion S group PCR amplification fragment sequencing primers effect than M gene fragment primer .6 parts S gene fragment primer amplification product display, Xi’an major cause epidemic spread of strains HFRS still Hantaan