采用 RT-PCR 技术结合熔解曲线分析，对临床样本中八种不同病毒病原体进行多重检测。
Multiplex detection of eight different viral enteropathogens in clinical samples, combining RT-PCR technology with melting curve analysis.
机构信息
Department of Clinical Laboratory, The Children's Hospital, Zhejiang University School of Medicine, National Clinical Research Center for Child Health, Hangzhou, 310052, People's Republic of China.
Jiangsu Bioperfectus Biotechnology Technologies Co, LTD, Taizhou, People's Republic of China.
出版信息
Virol J. 2022 Apr 7;19(1):61. doi: 10.1186/s12985-022-01789-z.
BACKGROUND
Early and accurate identification of infection viruses among children can benefit the personalized medical treatment and management, and reduce the future occurrence of serious symptoms. Thus, it is critical to develop a high-throughput multiplex real-time RT-PCR method to improve the accuracy and efficiency in routine clinical lab tests.
METHODS
We developed a real time RT-PCR combined with melting curve analysis (RRCMC) method for simultaneous detection of rotavirus A, B, C, norovirus GI and GII, adenovirus, astrovirus and sapovirus.
RESULTS
Stool samples were collected from 160 children with acute diarrhea and tested by RRCMC assay. A total of 71 patients were tested positive with norovirus, adenovirus or rotavirus. The RRCMC assay has high specificity. There is no internal cross-reaction among the 8 diarrhea viruses and no cross-reaction of other commonly intestinal pathogens and human genome. The limit detection was ranged from 1 × 10 to 1 × 10 nucleic acid copies/ml for each diarrhea virus.
CONCLUSION
The RRCMC method is a suitable rapid clinical test for infectious viruses, with the advantages of high-throughput, low cost, high sensitivity and specificity.
背景
早期、准确地识别儿童感染病毒有助于实现个体化医疗和管理，并降低未来出现严重症状的风险。因此，开发高通量多重实时 RT-PCR 方法以提高常规临床实验室检测的准确性和效率至关重要。
方法
我们开发了一种实时 RT-PCR 结合熔解曲线分析（RRCMC）方法，用于同时检测轮状病毒 A、B、C、诺如病毒 GI 和 GII、腺病毒、星状病毒和肠道病毒。
结果
从 160 例急性腹泻儿童中采集粪便样本，并用 RRCMC 检测。共有 71 例患者检测出诺如病毒、腺病毒或轮状病毒阳性。RRCMC 检测方法具有很高的特异性。8 种腹泻病毒之间无内部交叉反应，与其他常见肠道病原体和人类基因组无交叉反应。每种腹泻病毒的最低检测限为 1×10 到 1×10 核酸拷贝/ml。
结论
RRCMC 方法是一种适合用于检测传染性病毒的快速临床检测方法，具有高通量、低成本、高灵敏度和特异性的优点。
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