Wang Jian-Chang, Liu Li-Bing, Han Qing-An, Wang Jin-Feng, Yuan Wan-Zhe
Center of Inspection and Quarantine, Hebei Entry-Exit Inspection and Quarantine Bureau, Shijiazhuang 050051, China.
Hebei Animal Disease Control Center, Shijiazhuang 050050, China.
J Virol Methods. 2017 Oct;248:145-147. doi: 10.1016/j.jviromet.2017.06.011. Epub 2017 Jul 6.
Recombinase polymerase amplification (RPA), an isothermal amplification technology, has been developed as an alternative to PCR in pathogen detection. A real-time RPA assay (rt-RPA) was developed to detect the porcine parvovirus (PPV) using primers and exo probe specific for the VP2 gene. The amplification was performed at 39°C for 20min. There was no cross-reaction with other pathogens tested. Using the recombinant plasmid pPPV-VP2 as template, the analytical sensitivity was 103 copies. The assay performance was evaluated by testing 115 field samples by rt-RPA and a real-time PCR assay. The diagnostic agreement between assays was 100%, and PPV DNA was detected in 94 samples. The R value of rt-RPA and real-time PCR was 0.909 by linear regression analysis. The developed rt-RPA assay provides a useful alternative tool for rapid, simple and reliable detection of PPV in diagnostic laboratories and at point-of-care, especially in remote and rural areas.
重组酶聚合酶扩增（RPA）是一种等温扩增技术，已被开发作为病原体检测中聚合酶链反应（PCR）的替代方法。开发了一种实时RPA检测方法（rt-RPA），使用针对VP2基因的引物和外切核酸酶探针来检测猪细小病毒（PPV）。扩增在39°C下进行20分钟。与所检测的其他病原体没有交叉反应。以重组质粒pPPV-VP2为模板，分析灵敏度为103个拷贝。通过rt-RPA和实时PCR检测115份现场样本评估该检测方法的性能。两种检测方法之间的诊断一致性为100%，在94份样本中检测到PPV DNA。通过线性回归分析，rt-RPA与实时PCR的R值为0.909。所开发的rt-RPA检测方法为诊断实验室和即时检测中快速、简单且可靠地检测PPV提供了一种有用的替代工具，特别是在偏远和农村地区。
