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To visually and rapidly detect a novel goose astrovirus (N-GoAstV) causing fatal gout in goslings, an isothermal detection method based on one-step reverse transcription loop-mediated isothermal amplification (one-step RT-LAMP) was established. The one-step RT-LAMP assay for N-GoAstV detection, using Bst 3.0 DNA polymerase with strong reverse transcription activity and primer sets targeting the opening reading frame 1b (ORF1b) of N-GoAstV, could be completed in 30 min using a water bath at 61°C; the detection results could be visually observed by adding a pH-sensitive dye containing phenol red and cresol red. The detection limit of the one-step RT-LAMP assay was 57.8 copies/μL, which was similar to that of reverse transcription–quantitative polymerase chain reaction. The assay specifically detected N-GoAstV without any cross-reaction with other reference viruses, and this was further confirmed using enzyme digestion. These results indicated that the newly established RT-LAMP assay could accomplish reverse transcription, amplification, and visual result determination in one step, and the results obtained via this rapid and cost-effective method could be used to support disease control on farms in terms of N-GoAstV infection.

In this study, a one-step isothermal method combining polymerase spiral reaction (PSR) with reverse transcription (RT-PSR) was established for rapid and specific detection of novel astroviruses causing fatal gout in goslings (N-GoAstV). The one-step RT-PSR was accomplished at the optimal temperature of 62°C and time of 40 min and used primers simply designed as conventional PCR primers, and the results of detection were visible to the naked eye. The detection limit of PSR was above 34.7 copies/μL at a 95% probability level according to probit regression analysis. The assay specifically detected N-GoAstV, and no other reference viruses were detected. These results suggest that the newly established RT-PSR assay could, in one step, accomplish reverse-transcription, amplification, and result determination providing a visible, convenient, rapid, and cost-effective test that can be carried out onsite, in order to ensure timely quarantine of N-GoAstV-infected birds, leading to effective disease control.