Accomplishment of a protocol for simultaneous detection of 14 intestinal bacterial pathogens based on PCR-Reverse Dot Blot

Food poisoning, caused by a bacterial infection, consequently led to a wide range of infections and endanger to public health, has been considered as a big concern in the world. Therefore, it is an urgent demand for the clinical laboratory to exactly identify infectious bacteria. In our previous study, we successfully published a protocol based on the PCR-Reverse Dot Blot (PCR-RDB) to determine simultaneously 12 bacterial intestinal pathogens, including Bacillus cereus, Clostridium botulinum, Clostridium perfringen, Staphylococcus aureus, Listeria monocytogenes, Escherichia coli O157: H7, Salmonella spp., Shigella spp.., Vibrio cholerae, Vibrio parahaemolyticus, Yersinia enterocolitica and Brucella spp. In this study, we continuously developed our published protocol by designing additional probes: positive control probe, negative control probe, color control probe and background signal controller. Moreover, concerning the clinical demand, the supplement of two designed probes, which detected Campylobacter jejuni và Yersinia enterocolitica O:8 caused intestinal infected diseases mainly in children, was necessary. As a result, this completed PCR-RDB protocol can simultaneously detect a total of 14 intestinal bacterial pathogens within high specificity and the sensitivity of 102 copies/ml. For the protocol confirmation, it was tested by 30 fecal samples and the results completely match with the commercial kit PowerCheckTM 20 Pathogen Multiplex Real-time PCR Kit (Korea).