Establishment of Multiplex PCR Detection Method for Salmonella,Pasteurella and Escherichia coli

A multiplex PCR assay was developed and evaluated for the clinical detection of Salmonella spp,Pasteurella and Escherichia coli.Three pairs of primers have been designed and synthesized based on the highly conserved regions of the Salmonella spp hut gene,Pasteurella kmt1 gene and Escherichia coli 23SrRNA gene.The multiplex PCR reaction condition was optimized and the multiplex PCR method for detecting the co-infection of Salmonella spp,Pasteurella and Escherichia coli was established.The multiplex PCR have been tested for specificity,sensitivity,repeatability and artificial simulation tests.The specificity test showed that a fragment of 286 bp,457 bp and 652 bp were amplified from Salmonella spp,Pasteurella and Escherichia coli,respectively.No amplification was achieved from control groups of other bacteria.The sensitivity test showed that the multiplex PCR could detect 1.77×104 CFU/mL for Salmonella spp,1.99×104 CFU/mL for Pasteurella and 2.01×104 CFU/mL for Escherichia coli,respectively.The artificial simulation test showed that the multiplex PCR could detect three kinds of pathogens from co-infection disease material.The result indicated the multiplex PCR method had advantages of specificity,sensitivity,repetitive,and it could effectively detect the co-infection of Salmonella spp,Pasteurella and Escherichia coli.