Department of Clinical Laboratory, Shanghai East Hospital, 150 Ji Mo Road, Shanghai, 200120, People's Republic of China.
Eur J Clin Microbiol Infect Dis. 2021 Dec;40(12):2525-2532. doi: 10.1007/s10096-021-04300-8. Epub 2021 Jul 15.
Due to the highly contagious and spreads quickly of respiratory infectious diseases (ADR), the availability of rapid, sensitive, and reliable diagnostic methods is essential for disease control. Here, we develop an approach based on loop-mediated isothermal amplification (LAMP) for the detection of influenza A virus (Flu A), Flu A subtypes H1N1and H3N2, influenza B virus (Flu B), respiratory syncytial virus (RSV) subtypes A and B, human adenovirus (HAdV), parainfluenza virus (PIV) subtypes 1 and 3, and human rhinovirus (HRV) simultaneously. We designed primers specific to detect respiratory viruses above, optimized the RT-LAMP assay and evaluated it for its sensitivity and specificity of detection using real-time monitoring based on SYBR Green I. We also evaluated the result of our RT-LAMP assay on 638 nasopharyngeal swab specimens with the commercial RT-PCR by Cohen's Kappa. The inconsistent results were verified by Sanger sequencing furtherly. The developed RT-LAMP assay displayed a detection limit of 1 × 10 copies/ml RNA close to that of RT-PCR; no cross-reactivity was observed in the 10 kinds of viruses studied. The results obtained with 638 clinical samples indicate that the developed method has high specificity (0.988-1) and sensitivity (0.863-1) for viruses studied, and the Kappa value of all viruses was more than 0.85 revealed an excellent agreement between the two methods. We developed an RT-LAMP-based method and optimized for the detection of common respiratory viruses. It may be a powerful tool for rapid and reliable clinical diagnosis of ADR in primary hospitals.
由于呼吸道传染病（ADR）具有高度传染性且传播迅速，因此快速、敏感、可靠的诊断方法对于疾病控制至关重要。在这里，我们开发了一种基于环介导等温扩增（LAMP）的方法，用于同时检测甲型流感病毒（Flu A）、Flu A 亚型 H1N1 和 H3N2、乙型流感病毒（Flu B）、呼吸道合胞病毒（RSV）亚型 A 和 B、人腺病毒（HAdV）、副流感病毒（PIV）亚型 1 和 3 以及人鼻病毒（HRV）。我们设计了针对上述呼吸道病毒的特异性引物，优化了 RT-LAMP 检测，并通过基于 SYBR Green I 的实时监测评估了其检测的灵敏度和特异性。我们还通过 Cohen's Kappa 评估了我们的 RT-LAMP 检测方法对 638 份鼻咽拭子标本与商业 RT-PCR 的结果。不一致的结果通过 Sanger 测序进一步验证。开发的 RT-LAMP 检测方法的检测限为 1×10 拷贝/ml RNA，接近 RT-PCR；在研究的 10 种病毒中未观察到交叉反应。对 638 份临床样本的结果表明，该方法对研究的病毒具有高特异性（0.988-1）和灵敏度（0.863-1），两种方法之间的 Kappa 值均大于 0.85，表明两者具有极好的一致性。我们开发了一种基于 RT-LAMP 的方法并对其进行了优化，用于检测常见的呼吸道病毒。它可能是基层医院快速可靠的临床诊断 ADR 的有力工具。
