发热伴血小板减少综合征患者血清中新型布尼亚病毒RNA的检测  

SFTSV RNA detection in sera of patients suffering from fever with thrombocytopenia syndrome

作　　者：迟媛媛[1] 翟慎勇 温红玲[1] 崔峰[2] 宋艳艳[1] 王玲[2] 杜俊[2] 曹海霞[2] 王谦[3] 张寿锋 于学杰[1] 赵丽[1] 

机构地区：[1]山东大学公共卫生学院微生物检验研究所,济南250012 [2]淄博市疾病预防控制中心,山东淄博255026 [3]沂源县疾病预防控制中心,山东沂源256100

出　　处：《山东大学学报（医学版）》2012年第11期118-121,共4页Journal of Shandong University:Health Sciences

基　　金：山东大学公共卫生学院人才队伍建设基金

摘　　要：目的对发热伴血小板减少综合征布尼亚病毒(SFTSV)RNA PCR扩增引物进行优化,提高患者血清中SFTSV核酸检测的灵敏度和特异度。方法自行设计(L1、M1、S1)并参考文献报道(L2、M2、S2)选用引物12对,分成6组,巢式PCR方法分别扩增22例发热伴血小板减少综合征(SFTS)确诊病例和20例正常人血清中SFTSV RNA的L、M和S片段,基因克隆后进行DNA测序,计算不同引物扩增的灵敏度与特异度并进行比较,研究病毒检出率与血清采集时间的关系。结果不同引物RT-PCR扩增SFTS患者血清中SFTSV RNA的L、M和S片段的灵敏度不同。从生物学意义上看,S片段灵敏度最高,其次是L片段,未扩出M片段。血清中病毒RNA检出率,7 d内为64.3%,7～14 d为37.5%,应用确切概率法分析,P>0.05(=0.377),二者差异无统计学意义。正常人血清中未扩增出SFTSV RNA任何片断,所有引物检测的特异度均100%。结论 PCR检测SFTS患者血清中SFTSV RNA时,可优先选择S1引物。Objective PCR primers for amplification of severe fever with thrombocytopenia syndrome virus（SFTSV） were optimized in order to improve the detection sensitivity and specificity in sera from patients suffering from severe fever with thrombocytopenia syndrome（SFTS）.Methods RT-PCR primers were designed from the sequences of L,M,and S segments of SFTSV genome by using Primer Premier 5.0 and Oligo 6.0 softwares（L1,M1,S1） or obtained from references（L2,M2,S2）.12 primers were divided into 6 groups to amplify these segments.SFTSV RNAs in sera from patients with SFTS and normal sera were amplified by PCR and sequenced.SFTSV RNA detection sensitivity and specificity between primers were compared.At the same time,the relationship between the sensitivity and serum acquisition time were also compared.Results RT-PCR results showed that SFTSV detection sensitivity was different between primers.S-segment was most frequently amplified with one pair of primer（S1）,while L-segment was amplified occasionally and M segment was never amplified.Sensitivity for the detection of SFTSV RNA in sera was 64.3% when it was perfomed within 7 d,compared with 37.5% in 7~14 d.The difference between them was not statistically significant[P0.05（=0.377）].No segment of SFTSV was amplified in normal sera.Specificity of all primers was 100%.Conclusion S1 primer pair is the most sensitive one among all primer pairs derived from S-,M-and L-segments of SFTSV for RT-PCR amplification of S-segment.

关 键 词：发热伴血小板减少综合征布尼亚病毒 巢式PCR 引物优化 

分 类 号：R446.1[医药卫生—诊断学]