Genotypic and phenotypic characterizations of 50 clinical Vibrio cholerae isolates obtained during 4 consecutive y from 2004 to 2007 in Iran were studied. Antimicrobial susceptibility test, biochemical fingerprinting with Phene Plate system (PhP-RV) and pulsed-field gel electrophoresis (PFGE) were performed. Antibiotic susceptibility test of all isolates showed 12 different profiles. The predominant antimicrobial resistance profile (62%) was simultaneous resistance to SXT, streptomycin, chloramphenicol and erythromycin. PFGE revealed a total of 9 pulsotypes with a single dominant type in each y under study. PhP-RV revealed 6 common and 10 single types constituting 80% and 20% of the isolates, respectively. Diversity index of PhP-RV was 0.853, indicative of homogeneity among the isolates. Data obtained from PhP-RV was in close agreement with the results of PFGE genotyping. A comparison of the published PFGE patterns performed using the PulseNet protocol revealed the presence of similar patterns between some of our isolates and the isolates from Pakistan, Nepal and India, suggestive of dissemination of common V. cholerae clones in this region of the world. This could, in part, be due to human travel or occurrence of analogous DNA rearrangements, resulting in the emergence of similar V. cholerae genotypes in regional countries.