In this study, pulsed-field gel electrophoresis (PFGE) was applied to determine if the Vibrio cholerae 01 strains which reappeared after being temporarily displaced in Calcutta by the 0139 serogroup were different from those isolated before the advent of the 0139 serogroup. NotI digestion generated a total of 11 different patterns among the 24 strains of V. cholerae randomly selected to represent different time frames. Among the V. cholerae 01 strains isolated after July 1993, 4 PFGE banding designated as H through K were observed with pattern H dominating. Pattern H was distinctly different from all other patterns encountered in this study including patterns A, B and C of V. cholerae 01 E1 Tor, which dominated before November 1992, and pattern F, which was the dominant V. cholerae 0139 pattern. Further, pattern H was also different from the NotI banding patterns of the representative strains of the 4 toxigenic clonal groups of V. cholerae 01 E1 Tor currently prevailing in different parts of the world. NotI fragments of the new clone of V. cholerae 01 did not hybridize with an 0139 specific DNA probe, indicating that there was no 0139 genetic material in the new clone of V. cholerae 01. Hybridization data with an 01-specific DNA probe again differentiated between the clones of V. cholerae 01 existing before the genesis of the 0139 serogroup and the 01 strains currently prevalent.