Rapid detection of Campylobacter coli, C. jejuni, and Salmonella enterica on poultry carcasses by using PCR-enzyme-linked immunosorbent assay.
Contamination of retail poultry by Campylobacter spp. and Salmonella enterica is a significant source of human diarrheal disease. Isolation and identification of these microorganisms require a series of biochemical and serological tests. In this study, Campylobacter ceuE and Salmonella invA genes were used to design probes in PCR-enzyme-linked immunosorbent assay (ELISA), as an alternative to conventional bacteriological methodology, for the rapid detection of Campylobacter jejuni, Campylobacter coli, and S. enterica from poultry samples. With PCR-ELISA (40 cycles), the detection limits for Salmonella and Campylobacter were 2 x 10(2) and 4 x 10(1) CFU/ml, respectively. ELISA increased the sensitivity of the conventional PCR method by 100- to 1,000-fold. DNA was extracted from carcass rinses and tetrathionate enrichments and used in PCR-ELISA for the detection of Campylobacter and S. enterica, respectively. With PCR-ELISA, Salmonella was detected in 20 of 120 (17%) chicken carcass rinses examined, without the inclusion of an enrichment step. Significant correlation was observed between PCR-ELISA and cultural methods (kappa = 0.83; chi-square test, P < 0.001) with only one false negative (1.67%) and four false positives (6.67%) when PCR-ELISA was used to screen 60 tetrathionate enrichment cultures for Salmonella. With PCR-ELISA, we observed a positive correlation between the ELISA absorbance (optical density at 405 nm) and the campylobacter cell number in carcass rinse, as determined by standard culture methods. Overall, PCR-ELISA is a rapid and cost-effective approach for the detection and enumeration of Salmonella and Campylobacter bacteria on poultry.
Hong
Yang
Y
Department of Avian Medicine, College of Veterinary Medicine, The University of Georgia, Athens, Georgia 30602, USA.
Berrang
Mark E
ME
Liu
Tongrui
T
Hofacre
Charles L
CL
Sanchez
Susan
S
Wang
Lihua
L
Maurer
John J
JJ
D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D002169:Campylobacter Infections / Q000382:microbiology / Q000662:veterinary; D017000:Campylobacter coli / Q000235:genetics / Q000302:isolation & purification*; D016123:Campylobacter jejuni / Q000235:genetics / Q000302:isolation & purification*; D002352:Carrier Proteins / Q000235:genetics; D002645:Chickens / Q000382:microbiology*; D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis; D004797:Enzyme-Linked Immunosorbent Assay; D005511:Food Handling / Q000379:methods; D006801:Humans; D033862:Iron-Binding Proteins; D016133:Polymerase Chain Reaction / Q000379:methods*; D011201:Poultry Diseases / Q000382:microbiology; D012481:Salmonella Infections, Animal / Q000382:microbiology; D019779:Salmonella enterica / Q000235:genetics / Q000302:isolation & purification*; D013997:Time Factors
D001426:Bacterial Proteins; D002352:Carrier Proteins; D017931:DNA Primers; D004269:DNA, Bacterial; D033862:Iron-Binding Proteins; C499680:ceuE protein, Campylobacter coli; C433864:ceuE protein, Campylobacter jejuni; C075518:invA protein, Bacteria