Amplification of ctxA gene in Vibrio cholerae 0139

Abstract

The direct detection of cholera enterotoxin (ctxA) gene in Vibrio cholerae 0139 was examined using polymerase chain reaction (PCR). A pair of synthetic primers, each of which were 24 bases in length, were used with Taq polymerase to amplify the cholera enterotoxin. Vibrio cholerae 0139 strains isolated from clinical samples during the 1993 cholera outbreak were tested by PCR to detect the presence of the ctxA gene. The amplified PCR produce of 290-bp was detected by either gel electrophoresis or hybridization to a 25-base synthetic oligonucleotide probe labeled with biotin. The fragment was produced following 35 cycles of amplification plus an additional cycle for the extension step. The PCR method did not detect ctxA gene in diarrhoeagenic E coli, Shigella sp., Aeromonas sp. or Plesiomonas shigelloides. This amplification of ctxA gene by PCR provides a highly sensitive and specific tool for detection of toxigenic Vibrio cholerae directly from clinical samples.