Development of species-specific DNA probes for Campylobacter jejuni, Campylobacter coli, and Campylobacter lari by polymerase chain reaction fingerprinting.
The application of polymerase chain reaction (PCR) fingerprinting assays enables discrimination between species and strains of microorganisms. PCR primers aiming at arbitrary sequences in combination with primers directed against the repetitive extragenic palindrome (REP) or enterobacterial repetitive intergenic consensus (ERIC) motifs generate isolate-specific DNA banding patterns. Analysis of these PCR fingerprints obtained for 33 isolates of Campylobacter jejuni, 30 isolates of Campylobacter coli, and 8 isolates of Campylobacter lari revealed that besides generation of isolate-specific fragments, species-specific DNA fragments of identical size were synthesized. It appeared that these DNA fragments could be used as species-specific probes, since they are unique for the pattern which they are deriving from. The probes do not cross-react with amplified DNA originating from a large panel of nonrelated microorganisms. Moreover, these probes displayed species specificity, as they reacted with a single restriction fragment on Southern blots containing DNA from C. jejuni, C. coli, and C. lari and other Campylobacter species. This combination of PCR fingerprinting and probe hybridization results in a highly specific identification assay and provides an example of specific test development without the prior need for DNA sequence information. The principle of the procedure holds great promise for the rapid isolation of DNA probes which, in combination with a general PCR assay, may lead to efficient typing and detection procedures for a multitude of medically important nonviral microorganisms.
Giesendorf
B A
BA
Department of Molecular Biology, Diagnostic Center SSDZ, Delft, The Netherlands.
van Belkum
A
A
Koeken
A
A
Stegeman
H
H
Henkens
M H
MH
van der Plas
J
J
Goossens
H
H
Niesters
H G
HG
Quint
W G
WG
D001483:Base Sequence; D002167:Campylobacter / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D002169:Campylobacter Infections / Q000175:diagnosis; D017000:Campylobacter coli / Q000235:genetics; D016123:Campylobacter jejuni / Q000235:genetics; D016172:DNA Fingerprinting; D015342:DNA Probes / Q000235:genetics* / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000235:genetics; D005069:Evaluation Studies as Topic; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D013045:Species Specificity
D015342:DNA Probes; D004269:DNA, Bacterial