基于受体捕获诺如病毒检测方法的特异性及应用

摘要

目的 评价新型感染性诺如病毒检测方法原位捕获反转录实时定量PCR(In situ capture realtime quantitative reversetranscription polymerase chain reaction,ISC-RT-qPCR)的特异性,并将其应用于新鲜草莓中诺如病毒的检测.方法 以诺如病毒G Ⅱ组不同基因型和不同浓度的临床粪便标本评价ISC-RT-qPCR方法的特异性.此外,利用ISC-RT-qPCR方法对以G Ⅱ组诺如病毒人工污染新鲜草莓样本进行检测.结果 ISC-RT-qPCR可特异性检测所有G Ⅱ诺如病毒8种基因型临床腹泻样本;与传统RT-qPCR相比,检测的Ct值并未随着诺如病毒滴度的降低而升高.人工污染新鲜草莓的ISC-RT-qPCR检测限为1.36基因组拷贝数.结论 ISC-RT-qPCR是一种可用于临床腹泻样本和草莓中G Ⅱ组感染性诺如病毒的检测方法,为食品中诺如病毒的检测与监测提供了技术储备.%Objective To evaluate of the specificity of a new norovirus (NoV) detection method of in situ capture real-time quantitative reverse transcription polymerase chain reaction (ISC-RT-qPCR) and to apply the method for the detection of NoVs in fresh strawberry.Methods A panel of stool samples with different NoV genotypes and various inoculums were used for the experiments.Results We found that all the tested samples of eight genogroup Ⅱ (G Ⅱ) NoVs could be detected specifically by ISC-RT-qPCR.Moreover,in contrast to the conventional RT-qPCR method,the situation that the Ct value increased as the inoculum of NoV G Ⅱ decreased was not shown using ISC-RT-qPCR.When we tested NoVs in strawberry samples by ISC-RT-qPCR,the minimum test limit could reach 1.36 genocopy/10 g of fresh strawberry.Conclusions ISC-RT-qPCR is an effective and specific technic and it could be applied for the detection of infectious NoVs from stool samples and fresh strawberry samples.