Multiplex PCR assay for detection of virulence genes in Avian pathogenic Escherichia coli

Abstract:

Abstract:［Objective］To efficiently study the virulence genes distribution of Avian pathogenic Escherichia coli (APEC)，we developed four multiplex PCＲ to detect adhesin-associated genes，invasin and toxin-associated genes，serum resistanceassociated genes and iron acquisition-associated genes． ［Methods］According to gene sequences published in GenBank，we designed and synthesized 18 specific primer pairs，which were used in the four multiplex PCR．Then，we determined the sensitivity of multiplex PCR using diluted bacterial or DNA templates．To verify the feasibility of these multiplex PCR， we determined the distribution of virulence genes in 100 APEC isolates using these multiplex PCR．［Results］According to the results of PCR，we can conclude that each of the 18 genes was exactly and effectively amplified in the four multiplex PCR．The sensitivities of these four multiplex PCR were 103 Colony forming units (CFU)，103 CFU，105 CFU，105 CFU bacteria and 1ng，1ng，10ng and 10ng DNA，respectively．Furthermore，the results multiplex PCR for virulence genes distribution in 100 APEC were same as the single PCR．[Conclusion] These results suggest that multiplex PCR developed in this study could efficiently detect the virulence genes of APEC，which was a useful and rapid technique for epidemiological investigation．