Real-Time PCR Assay Using Light Upon eXtension Fluorogenic Primer for Detection of Campylobacter jejuni/coli in Food after Enrichment Culture

Abstract

The Light Upon eXtension fluorogenic (LUX) primers were designed to detect the hip gene specific for Campylobacter jejuni, the glyA gene specific for C. coli, and the glyA gene specific for both C. jejuni and C. coli. The real-time quantitative PCR assay using LUX primer (LUX-qPCR) was performed with the primer sets of J-hip-FU/RL for detection of C. jejuni, C-glyA-FU / RL for detection of C. coli, and JC-glyA-FL / RU for detection of C. jejuni / coli. Those LUX primers were specific for C. jejuni, C. coli, and C. jejuni/coli, respectively; the detection limit of the LUX-qPCR was 2,200 to 3,800 CFU/ml (11 to 19 CFU/reaction mixture) and coe$cient (r 2) for the correlation between amount of DNA and Ct value was calculated to be more than 0.99. The multiplex LUX-qPCR with J-hip-FU / RL and C-glyA-FU / RL allowed simultaneous detection and di#erentiation of C. jejuni and C. coli. The LUX-qPCR and the multiplex LUX-qPCR were carried out in food after enrichment culture and compared to the cultural method. The results between these LUX-qPCR assays and the cultural method were mostly corresponding: C. jejuni/coli strains were isolated from most of the enrichment cultures which were PCR-positive and not isolated from all of the cultures which QQQ