Cholera is a major public health problem, recently as an E1 Tor pandemic and now emergent as a new threat in the form of the O139 serotype. The toxin-coregulated pilus (TCP) has been shown to be an essential virulence factor capable of eliciting protective antibodies in the infant mouse cholera model. Biotype-specific antigenic differences have been demonstrated between the classical and E1 Tor proteins. Genetic analysis of a range of classical and environmental isolates reveals strict conservation of the tcpA gene in all clinical isolates examined, and shows the O139 (Bengal) pilin to be of E1 Tor type. TcpA -related fragments of various sizes were detected in the genomes of environmental Vibrio chohrae isolates by Southern hybridisation. Five environmental Vibrio cholerae isolates, 2 non-cholera vibrios, and an Escherichia coli strain, as well as 18 clinical O1 and O139 isolates, were tested by polymerase chain reaction (PCR) at high and low stringencies. This readily identified clinical isolates, and O139 and E1 Tor products are easily separated from those of classical isolates by restriction fragment length poly-morphtem. The data presented illustrates the use of PCR to simply distinguish environmental isolates of O1 and non-O1 from the clinical O1 and O139 isolates tested, and for differentiating classical from E1 Tor (and O139) isolates. This information also supports the notion that only two TcpA types are needed in vaccines targeting O1 and O139 (Bengal) cholera strains, and demonstrates the antigenic stability of this important surface-presented epitope over time.