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Development and testing of a nonradioactive DNA oligonucleotide probe that is specific for Vibrio cholerae cholera
A C Wright1, Y Guo, J A Johnson
1Department of Medicine, University of Maryland School of Medicine, Baltimore 21201.
Abstract:
An alkaline phosphatase-labeled oligonucleotide DNA probe (CTAP) that was specific for the cholera toxin gene (ctxA) was identified. All cholera toxin-producing strains of Vibrio cholerae, regardless of serotype, hybridized with the CTAP probe, while nontoxigenic strains from either environmental sources or from deletion or substitution mutations did not hybridize. Unlike the whole-gene probes for either ctxA or for the heat-labile toxin or Escherichia coli (eltA), this 23-base sequence did not hybridize with E. coli or with vibrios other than V. cholerae that produce related toxins. By using CTAP to identify colonies grown on nonselective medium, V. cholerae was enumerated at concentrations of 10(3) to 10(7)/g from stool samples of volunteers who had ingested V. cholerae O1 strain 569B. CTAP provides a specific and sensitive tool for diagnosis and environmental monitoring of cholera toxin-producing V. cholerae.
Insights
A new DNA probe, cholera toxin alkaline phosphatase (CTAP), specifically detects cholera toxin genes in Vibrio cholerae. This tool enables accurate diagnosis and environmental monitoring of cholera-causing bacteria.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Cholera remains a significant public health concern, necessitating rapid and accurate diagnostic tools.
- Existing methods for detecting cholera toxin-producing Vibrio cholerae can be time-consuming or lack specificity.
Purpose of the Study:
- To develop and validate a specific DNA probe for the cholera toxin gene (ctxA).
- To assess the probe's utility in identifying and enumerating toxigenic Vibrio cholerae.
Main Methods:
- Development of an alkaline phosphatase-labeled oligonucleotide DNA probe (CTAP) targeting the ctxA gene.
- Hybridization assays with various Vibrio cholerae strains (toxigenic and non-toxigenic) and other bacterial species.
- Enumeration of Vibrio cholerae in stool samples using the CTAP probe.
Main Results:
- The CTAP probe specifically hybridized with all cholera toxin-producing Vibrio cholerae strains, irrespective of serotype.
- Non-toxigenic strains and other bacterial species, including E. coli, did not hybridize with the CTAP probe.
- Accurate enumeration of Vibrio cholerae in stool samples was achieved using CTAP.
Conclusions:
- CTAP is a highly specific and sensitive tool for detecting cholera toxin-producing Vibrio cholerae.
- This probe facilitates rapid diagnosis of cholera.
- CTAP is valuable for environmental monitoring of toxigenic Vibrio cholerae.