Rapid and sensitive detection of large yellow croaker iridovirus by real‐time RPA and RPA‐LFD

Abstract

Large yellow croaker (Larimichthys crocea) is a vital marine‐cultured species in China. Large yellow croaker iridovirus (LYCIV) can cause a high mortality rate in L. crocea. Rapid and convenient detection of LYCIV is an urgent demand for diagnosis. In this study, rapid and simple recombinase polymerase amplification (RPA), real‐time RPA and RPA combined with lateral flow dipstick (RPA‐LFD) methods were developed for the detection of LYCIV based on the conserved sequence of the LYCIV major capsid protein (MCP) gene. With these optimized RPA analyses, LYCIV detection could be completed within 20 min at 40°C. Both RPA and real‐time RPA could detect viral DNA as low as 102 copies/μL, while the detection limit of RPA‐LFD was 101 copies/μL, and there was no cross‐reaction with other aquatic pathogens (KHV, CyHV‐2, GCRV‐JX01, SVCV, LCDV and LMBV). In practical evaluation of RPA, real‐time RPA and RPA‐LFD methods, the results showed consistency with the general PCR detection. In short, the developed RPA, real‐time RPA and RPA‐LFD analyses could be simple, rapid, sensitive and reliable methods for field diagnosis of LYCIV infection and have significant potential in the protection of LYCIV infection.

Establishment and application of a rapid diagnostic method for BVDV and IBRV using recombinase polymerase amplification-lateral flow device

“…Both RPA and real-time RPA demonstrated the capability to detect viral DNA down to 102 copies/μLL, while RPA-LFD had a slightly lower limit of detection at 101 copies/μL comparable to that of RPA-LFD. Importantly, RPA-LFD showed no cross-reactivity with other waterborne pathogens ( 18 ). Similarly, for the visual detection of Porcine Epidemic Diarrhea Virus (PEDV), a reverse transcription (RT)-RPA assay combined with lateral flow dipstrip (LFD) was established, targeting the N gene.…”