[Detection of Campylobacter species by using polymerase chain reaction and nonradioactive DNA probes. II. PCR direct sequencing of the Campylobacter DNA].
We have developed a sensitive DNA hybridization assay for the detection and identification of Campylobacter species which are recognized as important pathogens of acute diarrheal disease in humans. This technique utilizes DNA probes complementary to nucleotide sequences present in 16S ribosomal RNA (rRNA) of C. jejuni, C. coli, C. laridis, C. fetus, C. fetus subsp. fetus, C. fetus subsp. venerealis and C. hyointestinalis, and polymerase chain reaction. The partial sequence of DNAs encoding the Campylobacter rRNA was first analyzed by direct solid phase sequencing in order to select suitable DNA probes. Amplified target DNA of 240 base pairs could be resolved on ethidium bromide-stained gels, and hybridized with DNA probes conjugated to alkaline phosphatase. In identification experiments, one of the 10 probes tested here gave a positive hybridization reaction with C. jejuni, C. coli and C. hyointestinalis but not with other Campylobacter species. The other was specifically reactive with C. fetus, C. fetus subsp. fetus and C. fetus subsp. venerealis. When applied to stool specimens, a good correlation was found between the results obtained by the present assay and by biochemical tests. These findings suggest that the nonradioactive probe assay can be used as the practical criterion for differentiating Campylobacter species.
Sato
M
M
Clinical Laboratory, Hyogo College of Medicine, Nishinomiya.
Yamashita
K
K
Muraki
C
C
Takarada
Y
Y
Otsuka
N
N
Kagawa
S
S
Matsuoka
A
A
D001483:Base Sequence; D002167:Campylobacter / Q000235:genetics* / Q000302:isolation & purification*; D015342:DNA Probes; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction
D015342:DNA Probes