ESTABLISHMENT OF A METHOD FOR RAPID DETECTION OF AEROMONAS HYDROPHILA BY RECOMBINASE POLYMERASE AMPLIFICATION (RPA) COMBINED WITH LATERAL FLOW STRIPS (LFD)

In order to establish a rapid detection method for Aeromonas hydrophila utilizing recombinase polymerase amplification (RPA) technology, we designed specific RPA primers based on the conserved sequence of the gyrB gene of Aeromonas hydrophila. The research included the optimization of reaction conditions and an assessment of specificity and sensitivity by both agarose gel electrophoresis (AGE) and lateral flow dipstick (LFD) chromatography. The experimental results showed that the established rapid detection for Aeromonas hydrophila could detect Aeromonas hydrophila within 30min at the optimal temperature of 38℃. The minimum limit of detection (LOD) for Aeromonas hydrophila pure culture and genomic DNA using the RPA-LFD method was determined to be 10³ cfu/mL and 100 pg/μL, respectively. Furthermore, the simultaneous detection of clinical samples from hybridized sturgeon, treated using our established RPA-LFD method, exhibited consistent results when compared to conventional PCR. In conclusion, the recombinase polymerase amplification combined with lateral flow test strip (RPA-LFD) method of Aeromonas hydrophila was successfully established by exploring and optimizing RPA reaction conditions. The method was characterized by its simplicity, rapid reaction time, and lack of reliance on expensive instrumenttion compared with conventional PCR. Importantly, this method provides effective technical support for the early diagnosis of sturgeon suffering from bacterial diseases caused by Aeromonas hydrophila in the future.