Detection and identification of toxigenic Vibrio cholerae O1 strains by a simplified polymerase chain reaction method.
The polymerase chain reaction (PCR) with three oligonucleotide primers was used to amplify both 184 bp and 425 bp DNA fragments of cholera toxin A subunit gene (ctxA) for detection and identification of toxigenic V. cholerae O1 strains in one step. For enhancing the sensitivity, we have used the chemiluminescent detection system for Southern hybridization. A total of 92 Vibrio strains were tested by both PCR and enzyme-linked immunosorbent assay (ELISA). Three E. coli (ETEC) strains, one toxigenic V. mimicus, and one toxigenic V. cholerae non-O1 were also tested for cross reactivities. Our results indicated that the correlation between the immunological ELISA method and the molecular biological PCR method were very well. The lowest detectable amount of the purified serotype Inaba chromosomal DNA by PCR is 2 pg and 0.4 pg by chemiluminescent detection system. In this work we report a PCR assay with a combination of three primers to detect and identify the ctxA of V. cholerae O1 in one step. The simplified and accurate method is useful for toxigenic V. cholerae O1 detection and identification.
Lee
H F
HF
Division of Pathology, National Institute of Preventive Medicine, Department of Health, Taipei, Taiwan, R.O.C.
Yeh
H L
HL
Hsiao
H L
HL
Wang
T K
TK
Liu
C H
CH
D001483:Base Sequence; D002772:Cholera Toxin / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D014734:Vibrio cholerae / Q000235:genetics / Q000302:isolation & purification* / Q000472:pathogenicity
D002772:Cholera Toxin