星状病毒
诺如病毒
病毒学
轮状病毒
生物
多路复用
多重聚合酶链反应
放大器
微生物学
聚合酶链反应
病毒
基因
生物化学
生物信息学
作者
Jacques Rohayem,Stefanie Berger,Thomas Juretzek,Ottmar Herchenröder,Michael Mögel,Maya Poppe,J Henker,Axel Rethwilm
标识
DOI:10.1016/j.jviromet.2004.01.016
摘要
A single-step multiplex reverse transcription-polymerase chain reaction (RT-PCR) assay that detects and identifies Norovirus, Astrovirus and Adenovirus in clinical stool samples is described. Four hundred sixty stool samples were tested from patients with non-rotavirus acute gastroenteritis, that were either stored at −80 °C and tested retrospectively, or tested immediately after viral nucleic acid extraction in a prospective manner, including outbreaks of gastroenteritis that occurred in Germany during the winter of 2003. The multiplex RT-PCR was validated against simplex RT-PCR with published primers for Norovirus (JV12/JV13 and p289/p290) and Astrovirus (Mon340/348), and against simplex PCR for Adenovirus. In both retrospective and prospective settings, the multiplex RT-PCR was equally sensitive and specific in detecting non-rotavirus infections compared with simplex RT-PCR/PCR. The specificity of the multiplex RT-PCR was assessed by sequencing of the amplicons that showed high nucleotide identities to Norovirus genogroup I/1, I/4, II/2, or II/4 clades, as well as to Astrovirus serotypes 1, 2, 4, or 8. The multiplex RT-PCR was also more sensitive than Astrovirus and Norovirus antigen enzyme immunoassays (IDEIA, Dako), as well as Astrovirus isolation in cell culture. This novel multiplex RT-PCR is an attractive technique for the rapid, specific, and cost-effective laboratory diagnosis of non-rotavirus acute gastroenteritis.
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