Rapid Detection of Vibrio parahaemolyticus in Seafood Using Loop-mediated Isothermal Amplification (LAMP) Method

Vibrio parahaemolyticus has been considered as one of the most important food-borne bacterial pathogens. The loop-mediated isothermal amplification ( LAMP ) that amplifies DNA with high specificity and rapidity under isothermal conditions was applied for rapid detection of V. parahaemolyticus for the first time. A set of four primers, two outer and two inner primers, was designed specifically to recognize the thermolabile hemolysin gene ( tlh ) of V. parahaemolyticus. Genomic DNAs from 28 bacterial strains including 14 V. parahaemolyticus strains were amplified using LAMP, and no amplicon was observed in the other 14 bacterial strains. The detection limits of LAMP assay and pure cultures were 90 fg/LAMP mixture of V. parahaemolyticus genomic DNA and 24 cfu/mL, respectively. And for directly detecting V. parahaemolyticus in artificially contaminated food samples, the detection limit was 89 cfu/g. In addition, 40 seafood samples were tested by LAMP, and 8 of them were V. parahaemolyticus positive. Among the tested positive samples, 6 samples were also detected to be positive by conventional microbiological methods. These results suggest that detection of V. parahaemolyticus by LAMP is an effective and low-cost procedure with high specificity and sensitivity, and no specialized equipment required.