An enrichment medium for increasing a very small number of vibrio parahaemolyticus cells to the detection limit of the loop-mediated isothermal amplification (LAMP) assay

摘要

We developed an enrichment medium for use with the loop-mediated isothermal amplification (LAMP) assay (enrichment media + LAMP assay) to quickly increase a very small number of Vibrio parahaemolyticus cells to the detection limit of the assay. Thirty-nine different enrichment media were prepared based on evaluating 12 potential ingredients. From our assessment of the 39 media, enrichment medium #36, which contained 2% sodium chloride, 1% proteose peptone no. 2, 0.1% trehalose, 0.5% α-ketoglutaric acid, 0.25% pyruvic acid, and 0.5% yeast extract (pH 8.6), was found to be most effective at enhancing the proliferation of V. parahaemolyticus during incubation for 3 h at 40°C. We compared the detection limits of the LAMP assay, the enrichment medium #36 + LAMP assay, and the cultivation method using bacterial cell and spiked shrimp sample tests. The detection limits of the LAMP assay, the medium #36 + LAMP assay, and the cultivation method were 10 3, 10 0-10 -1, and 10 -1 CFU ml -1, respectively. Enrichment medium #36 promoted a 10 3- to 10 4-fold increase in the bacterial population, and the detection limit of the enrichment media + LAMP assay was the same as that of the cultivation method.