A one-step reverse-transcription loop-mediated isothermal amplification assay optimized for the direct detection of cucumber green mottle mosaic virus in cucurbit seeds

Cucumber green mottle mosaic virus (CGMMV) is a seed-borne virus that causes significant economic losses in farms cultivating cucurbit plants. With the increase in global trade of cucurbit seeds, it is essential to develop a rapid, reliable, and convenient diagnostic method for the direct detection of CGMMV in these seeds for prevention and management of the disease. Here, we developed a one-step reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay for the direct detection of CGMMV in cucurbit seeds. To improve the efficiency of the one-step RT-LAMP assay, six primers were designed to target the most conserved regions of the gene encoding the movement protein of CGMMV. Our one-step RT-LAMP assay was optimized to improve specificity and sensitivity for CGMMV detection in individual seeds. A comparison of the detection sensitivity revealed that our one-step RT-LAMP assay was 100-fold more sensitive than the current reverse transcription-polymerase chain reaction assay used for CGMMV quarantine in Korea. Collectively, the one-step RT-LAMP assay developed in the present study is appropriate for the direct detection of CGMMV in individual cucurbit seeds.