Quantification and Differentiation of Campylobacter jejuni and Campylobacter coli in Raw Chicken Meats Using a Real-Time PCR Method

Abstract

Campylobacter species are one of the most common causes of bacterial diarrhea in humans worldwide. The consumption of foods contaminated with two Campylobacter species, C. jejuni and C. coli, is usually associated with most of the infections in humans. In this study, a rapid, reliable, and sensitive multiplex real-time quantitative PCR was developed for the simultaneous detection, identification, and quantification of C. jejuni and C. coli. In addition, the developed method was applied to the 50 samples of raw chicken meat collected from retail stores in Korea. C. jejuni and C. coli were detected in 88 and 86% of the samples by real-time quantitative PCR and the conventional microbiological method, respectively. The specificity of the primer and probe sets was confirmed with 30 C. jejuni, 20 C. coli, and 35 strains of other microbial species. C. jejuni and C. coli could be detected with high specificity in less than 4 h, with a detection limit of 1 log CFU/ml by the developed real-time PCR. The average counts (log CFU per milliliter) of C. jejuni or C. coli obtained by the conventional methods and by the real-time PCR assay were statistically correlated with a correlation coefficient (R2) between 0.73 and 0.78. The real-time PCR assay developed in this study is useful for screening for the presence and simultaneous differential quantification of C. jejuni and C. coli.

Rapid Quantification of Viable Campylobacter Bacteria on Chicken Carcasses, Using Real-Time PCR and Propidium Monoazide Treatment, as a Tool for Quantitative Risk Assessment

“…It has been demonstrated in several studies comparing Q-PCR with culture-based enumeration that higher counts are produced by Q-PCR, which has been explained by the detection of DNA from dead and VBNC cells (11,15,46). This was also the case for the untreated samples in the present study; however, when PMA sample treatment was applied, a Q-PCRbased count lower than the culture-based count was often observed.…”

“…The introduction of real-time quantitative PCR (Q-PCR) has enabled faster, more sensitive, and less labor-intensive quantitative detection. Q-PCR methods for food-borne Campylobacter jejuni and C. coli in poultry, which is recognized as an important source of human Campylobacter infections, have been published (11,12,15,38,46). However, since control strategies mostly focus on reduction of the number of bacterial cells on the chicken carcass, the usefulness of these Q-PCR methods for risk assessment could be limited, since they detect all of the Campylobacter bacteria present in a sample, including the dead cells.…”

Simultaneous Detection and Differentiation of Campylobacter jejuni, C. coli, and C. lari in Chickens Using a Multiplex Real-Time PCR Assay

“…Multiplex real-time PCR has been used to discriminate C. jejuni and C. coli in clinical samples of human feces (LaGier et al 2004). However, there have been very limited studies on the detection of low levels of C. jejuni, C. coli, and C. lari in complex food matrices (Hong et al 2007;Bonjoch et al 2010).…”

Detection of Escherichia coli O157:H7 in Food Using Real-Time Multiplex PCR Assays Targeting the stx 1, stx 2, wzy O157, and the fliC h7 or eae Genes

“…Advantages of multiplex PCR assays, in which two or more DNA regions are co-amplified in one reaction, are lower cost and less time to obtain results. Real-time multiplex assays are performed using multiple dyes with distinct emission wavelengths, and a number of multiplex real-time PCR assays have been described for the detection and quantification of pathogens in food (Hong et al 2007;Wang et al 2007;Omiccioli et al 2009). In the present study, real-time PCR assays using two combinations of primers and probes were designed for the detection of E. coli O157:H7 in apple cider, raw ground beef, lettuce, and raw milk.…”