Abstract: Cronobacter sakazakii is an opportunistic food-borne pathogen that causes serious diseases in humans of different age groups, especially newborns and immunocompromised infants. In this study, recombinase polymerase isothermal amplification (RPA) cobined with lateral flow test strips (LF) was developed to quickly and reliably detect C. sakazakii. RPA amplification using the primers modified with biotin and digoxin produced large amount of double-stranded DNA products labeled with biotin and digoxin at each end. The products could interact with gold-labeled anti-digoxin antibody and streptavidin fixed on the LF, and finally showed visible results on the strip. RPA-LF method showed detection limit of 1.7×102 CFU/mL for C. sakazakii in pure culture and no cross-reactivity with other common pathogens. Performance of the RPA-LF for atrifically contaminated food samples was evaluated and the results showed that the detection limit was 1.7×100 CFU/g after 4 h or 6 h enrichment for different food types. In order to further evaluate the performance of the method developed in this study, 20 real food samples were detected by the RPA-LF and the national standard method simultaneously. The results showed that test results of the RPA-LF were in agreement with that of national standard method of China.

Key words: recombinase polymerase isothermal amplification (RPA), lateral flow test strips (LF), Cronobacter sakazakii, rapid detection