Development of a PCR ELISA assay for the identification of Campylobacter jejuni and Campylobacter coli.
A polymerase chain reaction (PCR) assay was developed based on a solution-hybridization colorimetric end-point detection format (PCR ELISA) for the identification of Campylobacter jejuni and Campylobacter coli. PCR primers were designed to target a gene sequence with species-specific motifs. Five biotin-labelled probes targeted to the species-specific motifs were investigated for the detection of digoxygenin-labelled PCR products from C. jejuni and C. coli using the PCR ELISA format. Two probes were identified, one which reacts with both the C. jejuni and C. coli target sequences (probe CC2) and one probe which reacts with the C. jejuni target sequence only (probe CJ2). The specificity of the assay with the CJ2 and CC2 probes was investigated with a range of Campylobacter spp., Arcobacter spp., Helicobacter spp. and a range of unrelated organisms. The PCR ELISA assay and probes were demonstrated to be specific for C. jejuni and C. coli. The sensitivity of the PCR ELISA assay was demonstrated to be 10-100-fold more sensitive than a gel-based PCR method using the same primers. This PCR ELISA assay is sensitive, specific and significantly reduces the time needed for the identification of C. jejuni and C. coli and has the potential to facilitate early detection of these important gastro-intestinal pathogens.
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Preston Public Health Laboratory, Royal Preston Hospital, PO Box 202, Sharoe Green Lane, Fulwood, Preston, Lancs., PR2 9HG, UK.
Fox
A J
AJ
Bolton
F J
FJ
Wareing
D R
DR
Greenway
D L
DL
Borrow
R
R
D002169:Campylobacter Infections / Q000175:diagnosis*; D017000:Campylobacter coli / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D016123:Campylobacter jejuni / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D009693:Nucleic Acid Hybridization; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D015345:Oligonucleotide Probes