An improved reverse transcription loop-mediated isothermal amplification assay for sensitive and specific detection of serotype O foot-and-mouth disease virus

Publication date: Available online 30 June 2018
Source:Journal of Virological Methods
Author(s): Da-Rae Lim, Hye-Ryung Kim, Min-Ji Park, Ha-Gyeong Chae, Bok-Kyung Ku, Jin-Ju Nah, So-Yoon Ryoo, Sung-Hwan Wee, Yu-Ri Park, Hyo-Sung Jeon, Ji-Jeong Kim, Bo-Young Jeon, Hyeong-Woo Lee, Sang-Geon Yeo, Choi-Kyu Park
A sensitive and specific swarm primer-based reverse transcription loop-mediated isothermal amplification (sRT-LAMP) assay for the detection of serotype O foot-and-mouth disease virus (FMDV) was developed and evaluated. The assay specifically amplified the VP3 gene of serotype O FMDV, but did not amplify the VP3 gene of other serotype FMDVs or any other viruses. The limit of detection of the assay was 102 TCID50/mL or 103 RNA copies/µL, which is 100 times lower than that of the RT-LAMP assay without swarm primers. The new assay is 10 times more sensitive than reverse transcription-polymerase chain reaction (RT-PCR) and is comparable to the sensitivity of real time RT-PCR (qRT-PCR). Evaluation of the assay using different serotypes of FMDV strains showed 100% agreement with the RT-PCR results. The previously reported serotype O FMDV-specific RT-LAMP assay did not detect 20 out of 22 strains of serotype O FMDVs, probably due to multiple mismatches between the primer and template sequences, showing that it is not suitable for detecting the serotype O FMDVs circulating in Pool 1 region countries, including Korea. In contrast, the developed sRT-LAMP assay with improved primers can rapidly and accurately diagnose serotype O FMDVs circulating in Pool 1 region countries and will be a useful alternative to RT-PCR and qRT-PCR.