Rapid visual detection of Streptococcus agalactiae in tilapia using recombinase polymerase amplification combined with lateral flow strips

Streptococcus agalactiae critically endangers tilapia farming and causes substantial economic losses to aquaculture. In this study, a rapid and efficient recombinase polymerase amplification-lateral flow strip (RPA-LFS) assay targeting the conserved cfb gene was established for the detection of S. agalactiae. Specific RPA primers were designed for the target region of the cfb gene, and an RPA-LFS system was developed based on primer-probe amplification and colloidal-gold visualization. Conventional PCR was performed in parallel to compare detection performance. The results showed that the F3/R primer pair exhibited the highest specificity under the optimized conditions of 28 °C for 15 min. The lowest detectable DNA concentration was 2.054 × 10^-6 ng/uL. Further sample testing showed that the minimum detectable bacterial copy number was 8.55 × 10³ copies/µL for RPA-LFS and 8.55 × 10⁴ copies/µL for conventional PCR. These results confirmed that the developed RPA-LFS assay had significantly higher detection sensitivity than conventional PCR and showed strong potential for the early surveillance and effective control of streptococcosis in tilapia farming.