Rapid detection of RSV, RBSDV, and SRBSDV in plants and insect vectors by the RT-RPA-LF assay

Rapid and accurate detection of viral pathogens in plants and insect vectors is critical for effective disease management. Here, we developed a reverse transcription recombinase polymerase amplification combined with lateral flow strip (RT-RPA-LF) assay for on-site detection of rice stripe virus (RSV), rice black-streaked dwarf virus (RBSDV), and southern rice black-streaked dwarf virus (SRBSDV). Virus-specific primers targeting conserved genomic regions of these three viruses were designed, and the specificity of these primes was tested, showing no cross-reactivity or false positives. Sensitivity tests revealed detection limits at 109- and 108-fold dilutions for SRBSDV and RSV/RBSDV in infected rice plants, respectively. For the insect vector samples, the RT-RPA-LF assay could identify a single viruliferous planthopper among a population of 1000 individuals. Then, the RT-RPA-LF assay was shown capable of field detection by integrating with a paper-based RNA rapid extraction method, and its accuracy was also proved to be comparable to that of conventional RT-PCR. In conclusion, the RT-RPA-LF method in this study enables rapid, simple, specific and visualizable on-site detection of rice viruses in both plant tissues and insect vectors, facilitating early diagnosis and timely implementation of disease management measures.