Vibrio cholerae non-O1 was found throughout the Apalachicola, FL, estuary. V. cholerae O1 was isolated primarily at the City of Apalachicola sewage treatment plant, Scipio Creek and the north shore of St. George Island. Highest concentrations of both serogroups occurred in August and November. Concentrations were lowest in February and increased substantially in May. A cholera toxin-like toxin was not detected in any of the V. cholerae cultures by the Y-1 mouse adrenal cell assay or the enzyme-linked immunosorbent assay (ELISA). However, 35% of the V. cholerae O1 and 22% of the V. cholerae non-O1 cultures selected for testing caused diarrhea in infant rabbits. The proportion of V. cholerae O1 and non-O1 isolates pathogenic to infant rabbits increased as water temperature decreased. Fecal coliforms appeared to be more useful than total coliforms as indicators of the numbers of V. cholerae in water, but neither of those National Shellfish Sanitation Program indicator groups ensured against the presence of pathogenic V. cholerae in shellfish-growing areas.