Rapid identification of Campylobacter jejuni from poultry carcasses and slaughtering environment samples by real-time PCR

Abstract

The objective of this study was to develop a real-time PCR assay for rapid identification of Campylobacter jejuni and to apply the method in analyzing samples from poultry processing. A C. jejuni-specific primer set targeting a portion of the C. jejuni hippuricase gene was developed. The specificity of the newly designed primer pair was verified using 5 C. jejuni strains and 20 other bacterial strains. Sensitivity was determined to be as low as 1 genome copy per reaction. A total of 73 samples were collected at different sites along the processing line during 2 visits to a poultry slaughterhouse and were examined by direct plating onto modified charcoal cefoperazone deoxycholate agar or after enrichment in Bolton broth followed by plating on modified charcoal cefoperazone deoxycholate agar. The newly developed real-time PCR assay was used to identify the presumptive colonies as belonging to C. jejuni. A real-time PCR assay targeting 16S ribosomal RNA was also applied to determine Campylobacter spp. prevalence. Results from the real-time PCR analysis indicated considerable variability in Campylobacter contamination, with incidence rates of 72.7 and 27.6% for sampling days A and B, respectively. Campylobacter was isolated from 100% of prescalded and preeviscerated carcasses on sampling day A. In contrast, on sampling day B, the highest number of Campylobacter-positive carcasses was recovered after evisceration (60%). The chilling process significantly reduced (P < 0.05) Campylobacter population, but the percentage of positive samples on sampling day A increased to 80%. All samples collected from the processing environment, except scalding tank 3 and the prechiller and chiller tanks, were 100% positive on day A, whereas no campylobacters were isolated from machinery on sampling day B. Our results revealed the widespread of C. jejuni in poultry processing and proved that the newly developed real-time PCR assay is a simple, specific, and inexpensive method for rapid C. jejuni identification. The newly developed PCR method can be easily used in laboratories for reliable and unambiguous identification of C. jejuni in poultry samples.

Characterization and Prevalence of Campylobacter spp. From Broiler Chicken Rearing Period to the Slaughtering Process in Eastern China

“…Chlorine has been known to have bactericidal effect, and has been applied on microbial contamination during chicken processing (25). A previous study from South California also showed a significant decrease of the Campylobacter contamination load for 0.97 logCFU/g after the water cooling process (7,26). In this study, the Campylobacter positive rate continuously kept at a low level (13.64%) during partition Campylobacter contamination in the environment during the slaughter process was relatively high indicating the possibility of cross contamination during slaughter.…”

Comparison of the Filtration Culture and Multiple Real-Time PCR Examination for Campylobacter spp. From Stool Specimens in Diarrheal Patients

“…Stool antigen tests for Campylobacter from the stool samples was rapid but still with the problems of specificity and positive predictive value (Giltner et al, 2013). The method of multiple real-time PCR is widely used in detecting and identifying Campylobacter (Rudi et al, 2004; Ivanova et al, 2014). In this study, we used quadruple real-time PCR method which can detect the major foodborne Campylobacter spp.…”

Unravelling the reservoirs for colonisation of infants with Campylobacter spp. in rural Ethiopia: protocol for a longitudinal study during a global pandemic and political tensions

“…Detection and quantification of Campylobacter is performed using genus-specific Taqman real-time PCR 32 and species-specific Sybr Green real-time PCR. [32][33][34][35][36] Open access…”