Objective: To develop a polymerase chain reaction(PCR) method for rapid detection of Listeria monocytogenes in oysters without pre-enrichment.
Methods: The combination of β-cyclodextrin and bentonite-coated activated carbon was used to remove PCR inhibitors from oyster samples, and the target gene inlB was used for the PCR subsequently. The specificity, sensitivity, and application of the developed method were verified, and the stability and application of the reagents stored under cryopreservation conditions were evaluated.
Results: The specificity of the developed PCR method was 100% for the detection of 130 target bacterial strains and 63 non-target bacterial strains. The method reduced the time required for Listeria monocytogenes detection to 4 h without pre-enrichment, and the detection limit was 10 CFU/25 g. The method was consistent with the conventional culture method on the detection rate and viable bacteria detection rate of Listeria monocytogenes in natural oyster samples(the coincidence rate was 100%). Additionally, the reagents could be used normally after storing at-20 ℃ for at least one year.
Conclusion: The PCR method developed in this study has high specificity and sensitivity, and can be used for rapid, accurate detection of Listeria monocytogenes in oysters.
Keywords: Listeria monocytogenes; bentonite-coated activated carbon; oysters; polymerase chain reaction; β-cyclodextrin.