用于检测蚊子中东马脑炎病毒变体的多靶点TaqMan检测法的开发。
Development of a multi-target TaqMan assay to detect eastern equine encephalitis virus variants in mosquitoes.
作者信息
Armstrong Philip M, Prince Nicholanna, Andreadis Theodore G
机构信息
Center for Vector Biology and Zoonotic Diseases, The Connecticut Agricultural Experiment Station, New Haven, Connecticut 06504, USA.
出版信息
Vector Borne Zoonotic Dis. 2012 Oct;12(10):872-6. doi: 10.1089/vbz.2012.1008. Epub 2012 Jul 26.
Abstract
Disease outbreaks caused by eastern equine encephalitis virus (EEEV; Togaviridae, Alphavirus) may be prevented by implementing effective surveillance and intervention strategies directed against the mosquito vector. Methods for EEEV detection in mosquitoes include a real-time reverse transcriptase PCR technique (TaqMan assay), but we report its failure to detect variants isolated in Connecticut in 2011, due to a single base-pair mismatch in the probe-binding site. To improve the molecular detection of EEEV, we developed a multi-target TaqMan assay by adding a second primer/probe set to provide redundant targets for EEEV detection. The multi-target TaqMan assay had similar performance characteristics to the conventional assay, but also detected newly-evolving strains of EEEV. The approach described here increases the reliability of the TaqMan assay by creating back-up targets for virus detection without sacrificing sensitivity or specificity.
摘要
通过实施针对蚊媒的有效监测和干预策略，可以预防由东部马脑炎病毒（EEEV；披膜病毒科，甲病毒属）引起的疾病暴发。在蚊子中检测EEEV的方法包括实时逆转录聚合酶链反应技术（TaqMan检测法），但我们报告称，由于探针结合位点存在单个碱基对错配，该方法未能检测出2011年在康涅狄格州分离出的病毒变体。为了改进EEEV的分子检测方法，我们开发了一种多靶点TaqMan检测法，通过添加第二组引物/探针为EEEV检测提供冗余靶点。多靶点TaqMan检测法具有与传统检测法相似的性能特征，还能检测出新出现的EEEV毒株。本文所述方法通过为病毒检测创建备用靶点，在不牺牲灵敏度或特异性的情况下提高了TaqMan检测法的可靠性。
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