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Foodborne pathogens can cause foodborne illness. In reality, one food sample may carry more than one pathogen. A rapid, sensitive, and multiple target method for bacteria detection is crucial in food safety. For the simultaneous detection of Staphylococcus aureus , Vibrio parahaemolyticus , and Salmonella Enteritidis, multi-objective recombinase polymerase amplification (RPA) combined with a lateral flow dipstick (LFD) was developed in this study. The whole process, including amplification and reading, can be completed in 15 min at 37 °C. The detection limits were 2.6 × 10 1 CFU/mL for Staphylococcus aureus , 7.6 × 10 1 CFU/mL for Vibrio parahaemolyticus , and 1.29 × 10 1 CFU/mL for Salmonella Enteritidis. Moreover, colored signal intensities on test lines were measured by a test strip reader to achieve quantitative detection for Staphylococcus aureus (R 2 = 0.9903), Vibrio parahaemolyticus (R 2 = 0.9928), and Salmonella Enteritidis (R 2 = 0.9945). In addition, the method demonstrated good recoveries (92.00%-107.95%) in the testing of spiked food samples. Therefore, the multiplex LFD-RPA assay is a feasible method for the rapid, sensitive, and quantitative detection of bacterial pathogens in seafood.

Multiplex Recombinase Polymerase Amplification Assay for the Simultaneous Detection of Three Foodborne Pathogens in Seafood