Development of a multiplex real-time PCR assay with an internal amplification control for the detection of Campylobacter spp. and Salmonella spp. in chicken meat

摘要

Conventional methods for the detection of Campylobacter and Salmonella based on culturing are time consuming and laborious. The aim of this study was to develop a multiplex real-time PCR assay with an internal amplification control for the simultaneous detection of Campylobacter spp. and Salmonella spp. in chicken meat Boiling was used for DNA extraction, followed by nucleic acid purification with phenol chloroform. Assay specificity was 100%, and the detection limit was 10(3) CFU of Campylobacter spp. and 10(6) CFU of Salmonella spp. per milliliter of spiked chicken meat rinse without an enrichment step. After 24 h of the selective enrichment of Campylobacter spp. and the non-selective enrichment of Salmonella spp., the assay sensitivity was 1 CFU of each of these pathogens per milliliter of rinse. To our knowledge, the present study is the first multiplex real-time PCR assay developed for the simultaneous detection of these pathogens with the inclusion of an internal amplification control to monitor PCR inhibitors. The developed assay is a relatively inexpensive and efficient means to detect Campylobacter spp. and Salmonella spp. in chicken meat after enrichment, and can be a useful alternative in food processing to prevent the distribution of contaminated food. (C) 2016 Elsevier Ltd. All rights reserved.