RT-nestedPCR和限制性酶切分析用于HV的检测和基因分型

EARLY DIAGNOSIS AND TYPING OF HV BY TRANSCRIPTION POLYMERASE CHAIN REACTION AND RESTRICTION ENZYME ANALYSIS

为建立检测肾综合征出血热 (HFRS)患者血清中汉坦病毒 (HV)基因的RT nestedPCR方法 ,并进一步进行限制性酶切分型 ,设计并合成互补于HVS基因片段的引物 ,对 78例HFRS患者血清进行RT nestedPCR检测 ,并对PCR产物进行酶切分析。结果显示 ,阳性率分别为 76 % (≤ 7天 )、6 7% (8～ 14天 )、43% (15～2 1天 )和 2 9% (>2 1天 )。 48例检测阳性患者PCR产物酶切分型 ,47例为Ⅰ型 ,1例为Ⅱ型。提示RT nestedPCR用于早期HFRS患者的HV基因检测 ,具有直接、敏感、特异等优点 。

To establish the technique of RT nested PCR for diagnosis of hemorrhagic fever with renal syndrome(HFRS), and typing of the amplified product was carried out by restriction enzyme digestion. Designed and synthesized the primers complemented to S segment of HV, the sera from 78 HFRS cases were tested by RT nested PCR and the amplified product was analysed by restriction enzyme digestion. The results showed that the positive rates of RT nested PCR were 76% (≤7 days), 67% (8~14 days), 43% (15~21 days),and 29% (>21 days). 48 positive amplified products from PCR were typed by restriction enzyme digestion,and it was found that 47 cases was type Ⅰand 1 case was type Ⅱ.It suggested RT nested PCR is a direct, sensitive and specific method, and typing by restriction enzyme digestion may be a rapid and accurate technique for differentiating dominating types of HV virus in China.