Assessment of loop-mediated isothermal application in rice tungro viruses

Loop-mediated isothermal amplification (LAMP), a recent and simple nucleic acid amplification technique, was optimized and applied to detect the rice tungro bacilliform virus (RTBV) and/or the rice tungro spherical virus (RTSV) in greenhouse inoculated rice seedlings and in field collected rice plants from the Philippine Rice Research Institute branch stations. RTBV was the target virus in the greenhouse trials and field samples taken during the 2014 and 2016 wet season (WS) while RTBV and RTSV in 2015 dry season (DS) cropping. LAMP detected RTBV in rice seedlings a day after inoculation in the greenhouse by the insect vector Nephotetix virescens while symptoms of slight stunting and pale yellowing of the second youngest leaf were evident three days later. Rice plants taken in 3x3 hill arrangement of randomly sampled during the 2014 DS and 2016 DS cropping that were noted with yellow leaves and stunted (Y, St) and yellow leaves with slight stunting (Y, SSt) were infected with yellow leaves but not stunted (Y, NSt) and healthy-looking (H) were uninfected, including those sampled exhibiting uniform plant height and yellowish leaves from the farmer's fields. During the 2015 DS cropping, plants noted as Y, St, Y, SSt, Y, NSt, G, St, G, SSt, and H were infected with either RTSV or RTBV and RTSV or uninfected. During this period, TRSV alone was the prevalent tungro virus infection at 41.67%. The combined infection RTSV alone in H and G, SSt plants with no obvious tungro symptoms of leaf yellowing and plant stunting amounted to 77.15%. When compared to the 80.00% negative reaction in other H plants, these revealed convincing evidence for providing an estimation of the real positives and negatives; a critical criteria for specificity. The above results also point to the benefits of using assay for RTBV alone to detect the early stage of tungro infection when symptoms are not yet fully expressed. In this instance, RTSV is plausibly present because RTBV alone is not yet fully transmissible. On the other hand, RTSV assay will reinforce the information on the presence of infected plants prior to symptom expression while visual diagnosis of leaf yellowing and plants stunting symptoms and that of sporadic disease spread features in the rice field are likely enough to confirm the infection to save assay time and resources. Promising results of this pilot undertaking on the use of molecular tool for tungro field diagnosis were obtained. But more trials and plants samples are suggested so that the predictive values target data will be established to creditably prove that LAMP assay can serve as a high through put support diagnostic tool for field diagnosis for tungro viruses.