Development of a reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay for the detection of porcine pegivirus

• Published in: Journal of virological methods 2019-08, Vol.270, p.59-65
• Main Authors: Li, Hao, Li, Kai, Bi, Zhen, Gu, Jun, Song, Deping, Lei, Dan, Luo, Suoxian, Huang, Dongyan, Wu, Qiong, Ding, Zhen, Wang, Leyi, Ye, Yu, Tang, Yuxin
• Format: Article
• Language: English
• Subjects: Nested RT-PCR; Porcine pegivirus; qRT-PCR; Reverse transcription-loop-mediated isothermal amplification
• ISSN: 0166-0934; 1879-0984
• DOI: 10.1016/j.jviromet.2019.04.019

•RT-LAMP assay was established for the simple and rapid detection of PPgV.•RT-LAMP assay showed strong specificity with high sensitivity.•RT-LAMP assay provides an extremely valuable screening tool for the surveillance of PPgV in field conditions. A simple and accurate reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay was developed and evaluated for the detection of porcine pegivirus (PPgV). The specific RT-LAMP primers targeting the conserved regions of NS5A genes were designed and used to detect PPgV. The optimal reaction parameter for RT-LAMP assay was 63℃ for 60 min. The detection limit of the RT-LAMP assay was 10 copies of PPgV genome, which was 100 times more sensitive than that of the conventional RT-PCR and comparable to nested RT-PCR and quantitative RT-PCR (qRT-PCR). There was no cross amplification with other related RNA viruses. In the clinical evaluation, the RT-LAMP assay exhibited a similar sensitivity with nested RT-PCR and qRT-PCR. The results indicated that RT-LAMP assay developed in this study could be a highly specific, sensitive, and cost-effective alternative for a rapid detection of PPgV in field settings.