Ontario Agency for Health Protection and Promotion, 81 Resources Road, Toronto, ON M9P 3T1, Canada.
J Clin Microbiol. 2011 Sep;49(9):3154-62. doi: 10.1128/JCM.00599-11. Epub 2011 Jul 20.
Acute viral gastroenteritis is an intestinal infection that can be caused by several different viruses. Here we describe the evaluation and verification of Seeplex Diarrhea-V ACE (Seeplex DV), a novel commercial multiplex reverse transcription-PCR (RT-PCR) assay that detects 5 diarrheal pathogens, including adenovirus, rotavirus, norovirus genogroup I (GI) and GII, and astrovirus. We describe a retrospective study of 200 clinical specimens of which 177 were stool specimens previously tested for the presence of gastrointestinal viruses by electron microscopy (EM) and/or real-time RT-PCR (rRT-PCR). The remaining 23 specimens comprised other human pathogens of viral or bacterial origin. Discordant norovirus GI and GII results were resolved using a commercial kit; discordant adenovirus and rotavirus results were resolved using a home brew multiplex rRT-PCR assay. Diagnostic sensitivities and specificities were calculated before and after discordant analysis. After discordant analysis, estimated diagnostic sensitivities were 100% for adenovirus, rotavirus, and norovirus GI and 97% for norovirus GII. Diagnostic specificities after discordant analysis were 100% for adenovirus, rotavirus, and norovirus GI and 99.4% for norovirus GII. The 95% limits of detection were 31, 10, 2, and 1 genome equivalent per reaction for adenovirus, rotavirus, and norovirus GI and GII, respectively. The results demonstrate that the Seeplex DV assay is sensitive, specific, convenient, and reliable for the simultaneous detection of several viral pathogens directly in specimens from patients with gastroenteritis. Importantly, this novel multiplex PCR assay enabled the identification of viral coinfections in 12 (6.8%) stool specimens.
急性病毒性胃肠炎是一种肠道感染，可由多种不同的病毒引起。在这里，我们描述了 Seeplex Diarrhea-V ACE（Seeplex DV）的评估和验证，这是一种新的商业多重逆转录聚合酶链反应（RT-PCR）检测方法，可检测 5 种腹泻病原体，包括腺病毒、轮状病毒、诺如病毒基因组 I（GI）和 GII，以及星状病毒。我们描述了一项回顾性研究，共 200 例临床标本，其中 177 例为粪便标本，此前曾通过电子显微镜（EM）和/或实时 RT-PCR（rRT-PCR）检测胃肠道病毒。其余 23 个标本包括其他病毒或细菌来源的人类病原体。用商业试剂盒解决了诺如病毒 GI 和 GII 的不一致结果；用自制的多重 rRT-PCR 检测方法解决了腺病毒和轮状病毒的不一致结果。在不一致分析之前和之后计算了诊断灵敏度和特异性。在不一致分析后，腺病毒、轮状病毒和诺如病毒 GI 的估计诊断灵敏度为 100%，诺如病毒 GII 的诊断灵敏度为 97%。不一致分析后，腺病毒、轮状病毒和诺如病毒 GI 的诊断特异性为 100%，诺如病毒 GII 的诊断特异性为 99.4%。腺病毒、轮状病毒和诺如病毒 GI 和 GII 的 95%检测限分别为 31、10、2 和 1 个基因组当量/反应。结果表明，Seeplex DV 检测方法灵敏、特异、方便、可靠，可直接从胃肠炎患者的标本中同时检测几种病毒病原体。重要的是，这种新的多重 PCR 检测方法可在 12（6.8%）份粪便标本中鉴定出病毒混合感染。
