Development of a multiplex real-time recombinase polymerase amplification (RPA) assay for rapid quantitative detection of Campylobacter coli and jejuni from eggs and chicken products

摘要

Chicken products are known as main vehicles for human infections and foodborne illness with campylobacteriosis. This study describes a rapid and sensitive method for the detection of C. coli and C. jejuni from eggs, chicken meat, and chicken broth using the multiplex real-time recombinase polymerase amplification (Rti-RPA) and its comparison to a culture dependent method. The optimal procedure for the multiplex Rti-RPA assay was set in isothermal condition at 45 degrees C for 20 min and the detection sensitivity was at 1 CFU/reaction in pure culture. In the case of food applications, the detection limit of C. coli and C. jejuni using the RPA assay was 1 CFU/mL from chicken broth, 10(3) CFU/g from egg and chicken meat samples without a pre-enrichment procedure, and 1 CFU/g from 24 h enriched egg and chicken meat samples. Quantifications of Campylobacter spp. in spiked food samples by the RPA assay were not significantly different with the results by the culture method. This multiplex Rti-RPA assay is considerably faster than other DNA amplification methods and exhibits no losses in detection sensitivity and specificity. This multiplex Rti-RPA is a simple, rapid detection and quantification assay that is completed within a 20 min runtime. The assay is applicable in the surveillance of C. coli and C. jejuni contamination in eggs and chicken products. (C) 2016 Elsevier Ltd. All rights reserved.