2023 Volume 65 Pages 16-21
We developed a method for detecting pear chlorotic leaf spot-associated virus (PCLSaV) from pear (Pyrus spp.) leaves using a reverse transcription loop-mediated isothermal amplification (RT-LAMP) method. Of the primers we designed, the primer set R2LP4 demonstrated the fastest amplification, enabling the detection of PCLSaV within 15 min using total RNA extracted from symptomatic leaves. In the diagnosis of leaves of the Japanese and European pear cultivars collected from 12 prefectures, the RT-LAMP assay demonstrated similar or superior sensitivity and accuracy to that of reverse transcription polymerase chain reaction. Moreover, PCLSaV could be detected by this simple RT-LAMP assay using sap collected with a syringe needle. Comparison of the nucleotide sequence also indicated that some of the designed primers would be compatible with Chinese isolates, which indicates that the RT-LAMP method developed here is valuable in Japan and other countries.
