Rapid PCR with nested primers for direct detection of Campylobacter jejuni in chicken washes.
Rapid detection of Campylobacter jejuni by PCR directly from foods, without prior growth steps, would be beneficial for the poultry industry. We have previously reported a PCR assay that allows detection of this bacterium after 48 h growth on Campy cefex agar. We have now developed a more rapid nested PCR assay that specifically detects C. jejuni in chicken washes that have not undergone any lengthy growth steps prior to PCR. For the nested reaction, an external set of primers, C-1 and C-4, are used for 24 cycles. At this time, 1 microl of the PCR product is removed and added to a second reaction. The second PCR assay is run with C-1 and an internal primer, C-2, for 24 cycles. A single band on a 4% NuSieve agarose gel at 122 bp was apparent with C. jejuni cells at a sensitivity of 10(2) cfu ml-1. With this method chicken carcasses can be washed and C. jejuni identified all within 1 day. We detected C. jejuni in approximately 80% of four groups of chickens using this method. The identifications have been confirmed by standard microbiological techniques.
Winters
D K
DK
Department of Poultry Science and the Arkansas Biotechnology Center, University of Arkansas, Fayetteville, AR 72701, USA.
O'Leary
A E
AE
Slavik
M F
MF
D000362:Agar; D000818:Animals; D001483:Base Sequence; D016123:Campylobacter jejuni / Q000235:genetics / Q000302:isolation & purification*; D002498:Centrifugation; D002645:Chickens / Q000382:microbiology*; D017931:DNA Primers / Q000235:genetics; D005069:Evaluation Studies as Topic; D005516:Food Microbiology; D008460:Meat / Q000382:microbiology; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013048:Specimen Handling
D017931:DNA Primers; D000362:Agar