Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
8737310 | Diagnostic Microbiology and Infectious Disease | 2018 | 5 Pages |
Abstract
The sensitivity of qRT-PCR assay is not adequate for the detection of the samples with lower viral load, particularly in the cerebrospinal fluid (CSF) of patients. Here, we present the development of a highly sensitive real-time nested RT-PCR (RTN RT-PCR) assay in a single closed tube for detection of human enterovirus (HEV). The clinical performance of both RTN RT-PCR and qRT-PCR was also tested and compared using 140 CSF and fecal specimens. The sensitivities of RTN RT-PCR assay for EV71, Coxsackievirus A (CVA)16, CVA6 and CVA10 achieved 10â8 dilution with a corresponding Ct value of 38.20, 36.45, 36.75, and 36.45, respectively, which is equal to traditional two-step nested RT-PCR assay and approximately 2-10-fold lower than that of qRT-PCR assay. The specificity of RTN RT-PCR assay was extensively analyzed insilico and subsequently verified using the reference isolates and clinical samples. Sixteen qRT-PCR-negative samples were detected by RTN RT-PCR and a variety of enterovirus serotypes was identified by sequencing of inner PCR products. We conclude RTN RT-PCR is more sensitive than qRT-PCR for the detection of HEV in clinical samples.
Keywords
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Applied Microbiology and Biotechnology
Authors
Xin-xin Shen, Fang-zhou Qiu, Huai-long Zhao, Meng-jie Yang, Liu Hong, Song-tao Xu, Shuai-feng Zhou, Gui-xia Li, Zhi-shan Feng, Xue-jun Ma,