Rapid detection of Campylobacter fetus by polymerase chain reaction combined with non-radioactive hybridization using an oligonucleotide covalently bound to microwells

Campylobacter fetus is recognized as a human and animal pathogen. The isolation and differentiation of C. fetus in diagnostic laboratories is hindered by its relatively slow growth and lack of distinguishing biochemical characteristics. We cloned and sequenced a 1581-bp DNA fragment, IG02, isolated from a C. fetus genomic library. This fragment was used as a probe on DNAs extracted from C. fetus strains and other Campylobacter species: IG02 hybridized only with DNAs from C. fetus strains. A PCR-based test was developed for the detection of C. fetus. A pair of oligonucleotide primers was designed to amplify a 141-bp fragment of IG02. The amplified product was analysed by a non-radioactive sandwich hybridization in microtiter plate using a capture oligonucleotide and a biotin-labelled oligonucleotide for the detection. The combination of PCR and non-radioactive microplate hybridization is a convenient method for the rapid detection of C. fetus. (C) 2000 Academic Press.