Cholera still remains in the top of causes generating global mortality and morbidity, as revealed by the latest WHO reports. In 2000, on CDC/Atlanta website, the cholera agent was mentioned as potential biological weapon for bioterrorism actions in 9460 publications. Considering these aspects, a correct and rapid diagnosis is necessary in order to take appropriate epidemiological measures and to prevent the secondary transmission. Our study evaluated the efficiency of microtest systems API20E, API 20 NE and RapID 20E, by comparison with classical biochemical tests concerning the diagnosis of 150 Vibrio cholerae strains isolated during cholera epidemics of Romania (1990-1995). Our results demonstrated that classical biochemical identification still remains the most secure diagnostic method for Vibrio cholerae strains. The kits API 20NE gave the highest number of results in concordance with the classical tests. For these reasons, the confirmation of cholera infection must be performed in a National Reference Center. The limits of API galleries concerning the diagnosis of Vibrio cholerae strains must be taken into account and in case of suspect clinical symptoms, the isolated strains must be sent to the Cantacuzino Institute, even if the microtest galleries have not identified V. cholerae strains.