Rapid detection of Singapore grouper iridovirus by a recombinase polymerase amplification combined with lateral flow dipstick

Singapore grouper iridovirus (SGIV), a novel species of Ranavirus, caused more than 90% mortality in larval and juvenile of grouper. Up to now, there is still a lack of effective prevention and control strategies for SGIV. Therefore, it is essential to develop convenient diagnostic methods for filed detection of SGIV without special equipment. In this study, to establish a rapid, sensitive and visualized method for the detection of SGIV in clinical samples, specific primers and probes was designed by targeting the SGIV ORF014L, and a recombinase polymerase amplification (RPA) technique combined with lateral flow dipstick (LFD) (RPA-LFD) was developed for the detection of SGIV. The results showed that the RPA reaction specially detect target fragment of SGIV within 20 min at 40.1 °C with the lowest detection limit of 10² copies/μL. The RPA-LFD reaction at constant temperature of 42 °C for 8 min was able to visualized the results on the test strips with the lowest detection limit of 10¹ copies/μL, and showed no cross-reaction with other common aquatic pathogens. The coincidence rate of positive test of clinical samples was consistent with between RPA-LFD and PCR methods. Both RPA and RPA-LFD could specifically detect SGIV with lower limit than conventional PCR assay. Taken together, RPA-LFD assay developed in present study provided a convenient, specific, sensitive and visualized method for on-site rapid detection of SGIV without special equipment.