Establishment and application of multiplex PCR for diagnosing three food-borne bacterial pathogens

Objective To establish a multiplex polymerase chain reaction(PCR) assay for simultaneous detection of Salmonella spp.,Proteus mirabilis and Staphylococcus aureus.Methods Three pairs of primers have been designed according to the hilA of Salmonella spp.,hemolysin(hpmA) of Proteus mirabilis and Staphylococcus aureus clone pSa-442 Sau3AI fragment and the anticipated PCR products were 580bp,401bp and 256bp.A multiplex PCR method has been developed for dignosing Salmonella spp.,Proteus mirabilis and Staphylococcus aureus,after analysis for specificity and sensitivity and optimization of reaction condition by orthogonal experimental design L16(43).Results The sensitivity of multiplex PCR was 94.07 pg genome DNA for Salmonella spp.,140.85 ng for Proteus mirabilis and 1.41 ng for Staphylococcus aureus.Simulation experiment showed that the minimum detection limit after cultivation for 4 hours was 100 CFU/ml for Salmonella spp.,101 CFU/ml for Proteus mirabilis and 100 CFU/ml for Staphylococcus aureus.Conclusions A rapid,specific and sensitive multiplex PCR system has been established and it is valuable for exploration and appliacation.