Papaya ringspot virus (PRSV) and Papaya leaf distortion mosaic virus (PLDMV), which causes disease symptoms similar to PRSV, threaten commercial production of both non-transgenic-papaya and PRSV-resistant transgenic papaya in China.A reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay to detect PLDMV was developed previously.In this study, the development of another RT-LAMP assay to distinguish among transgenic, PRSV-infected and PLDMV-infected papaya by detection of PRSV is reported.A set of four RT-LAMP primers was designed based on the highly conserved region of the P3 gene of PRSV.The RT-LAMP method was specific and sensitive in detecting PRSV, with a detection limit of 1.15× 10-6 μg of total RNA per reaction.Indeed, the reaction was 10 times more sensitive than one-step RT-PCR.Field application of the RT-LAMP assay demonstrated that samples positive for PRSV were detected only in non-transgenic papaya, whereas samples positive for PLDMV were detected only in commercialized PRSV-resistant transgenic papaya.This suggests that PRSV remains the major limiting factor for non-transgenic-papaya production, and the emergence of PLDMV threatens the commercial transgenic cultivar in China.However, this study, combined with the earlier development of an RT-LAMP assay for PLDMV, will provide a rapid, sensitive and cost-effective diagnostic power to distinguish virus infections in papaya.