逆转录-聚合酶链反应结合限制性片段长度多态性分析对我国肾综合征出血热病毒分型的研究(英文)

建立准确、快速、灵敏的汉坦病毒基因分型方法，可弥补传统血清学方法的不足，对防治该病毒所致的肾综合征出血热(HFRS)具有重要意义。本试验采用逆转录－聚合酶反应(RT-PCR)和限制性片段长度多态性(RFLP)和限制性片段长度多态性(RFLP)方法，对流行于我国的两型汉坦病毒代表株??汉滩型(HTNV)76－118和汉城型(SEOV)R22株进行基因分析。根据病毒DNA序列的电脑软件分析，不同HFRSV囊膜糖蛋白编码基因M节段1199~1497间核苷酸序列上RsaI、TaqI和HindIII的酶切位点存在差异(Fig.1), 可用于进行限制性内切酶基因多态性分析，以确定HFRV的型别。首先，以一对引物扩增该片段(Fig.2)。然后，分别用这三种内切酶(Fig.3,4,5)进行酶切分型。共分析了从我国不同地区，不同宿主分离的毒株18株，及国际标准毒株2株，酶切图谱显示，这些毒株可以被分为三组(Table.1)：9株可定为HTNV型，8株可定为SEOV型，3株无法确定其型别(X型)。该法分型结果与血清学经典的空斑减数中和试验分型结果基本一致，说明该酶切分型方法具有一定的可行性。

Studies on Genotyping of HFRSV Isolates in China by RT-PCR/RFLP

In order to develop a more accurate and rapid virus genotyping method, we carried out the RT-PCR/RFLP to analyze 18 Hemorrhagic Fever with Renal Syndrome Virus (HFRSV) isolates with different hosts and distribution. The primers were selected from the published sequence of the M segment of Hantaan virus strain 76-118 (HTNV76-118) and Seoul virus strain -R22 (SEOV R22), which made it possible to obtain an amplified product of 299 bp by RT-PCR. The differentiation of the amplified products was carried out by restriction enzyme digestion. With RsaI and TaqI, the RFLP of the RT-PCR products of 20 hantavirus strains exhibited three patterns, which were identical to those of serotyping data except for three samples (15%). These results suggest that the 299 bp region cDNA is virus type-dependent and can be used to differentiate isolates of this virus.