TY - JOUR
AU1 - Zhou, Shun
AU2 - Zheng, Xujia
AU3 - Yang, Zongrui
AU4 - Huang, Qing
AU5 - Yi, Jingyuan
AU6 - Su, Lin
AU7 - Guo, Baoshan
AU8 - Xiu, Yunji
AB - Aeromonas salmonicida is the pathogen underlying furunculosis, causing a septicemic infection that influences both salmonid and non-salmonid fish. Early diagnosis of these contagions is essential for disease surveillance and prevention, so a rapid and sensitive approach is needed. Herein, a recombinase polymerase amplification EXO (RPA-EXO) assay and RPA with a lateral flow dipstick (RPA-LFD) were produced for testing A. salmonicida. The RPA-EXO and RPA-LFD primer sets were devised based on the conserved fragment sequence of the vapA gene. Then, RPA-EXO and RPA-LFD reaction systems were established, and the reaction temperature and time were optimized. After optimization, the RPA-EXO method was capable of testing A. salmonicida within 10 min, and the RPA-LFD method could detect A. salmonicida in only 5 min. The RPA-EXO and RPA-LFD methods exhibited high specificity with no cross-reaction with other strains. To assess sensitivity, a partial vapA gene was cloned, and serial plasmid dilutions were created ranging from 1 × 106 to 1 × 10−1 copies/μL. The detection limit of RPA-EXO was 1 × 102 copies/μL, and the detection limit of RPA-LFD was 1 copy/μL. For spiked turbot tissue samples, the sensitivity detection of A. salmonicida was 1.2 × 101 CFU/mL and 1.2 CFU/mL by RPA-EXO and RPA-LFD, respectively. In comparative analyses of clinical samples, the diagnostic results of RPA-EXO and RPA-LFD were compared with those of the standard conventional PCR test and showed nearly 100% consistency. Therefore, our RPA-EXO and RPA-LFD assays exhibited excellent specificity and sensitivity, which provided two simple, fast and dependable methods to conduct large-scale field investigations of A. salmonicida in resource-limited settings.
TI - Development of Two Recombinase Polymerase Amplification EXO (RPA-EXO) and Lateral Flow Dipstick (RPA-LFD) Techniques for the Rapid Visual Detection of Aeromonas salmonicida
JF - Marine Biotechnology
DO - 10.1007/s10126-022-10170-8
DA - 2022-12-01
UR - https://www.deepdyve.com/lp/springer-journals/development-of-two-recombinase-polymerase-amplification-exo-rpa-exo-39rxOZaaB7
SP - 1094
EP - 1109
VL - 24
IS - 6
DP - DeepDyve
ER -