BACKGROUND: Olive oil mill wastewaters (OMW) from a three-phase system were treated in a laboratory-scale continuous stirred tank reactor (CSTR) for hydrogen production. Microbiological and physicochemical characterizations of raw and anaerobically treated olive oil mill effluents were performed. Toxicity assessment was also conducted in order to determine the impact of residual waters on the aquatic environment. RESULTS: Pseudomonas aeruginosa that has the ability to utilize phenolic compounds was isolated in raw OMW, along with several gram-negative bacteria (Serratia odorifera, Enterobacter cloacae and Aeromonas hydrophila). The dominant species in the untreated effluent appears to be Enterobacter cloacae, whereas in the treated effluent it is Citrobacter braakii. Phenols, pH, NO2-, total dissolved solids (TDS) and conductivity were the main parameters related to the observed toxicity. CONCLUSION: A significant differentiation in the microbiological flora was detected between treated and untreated effluents whereas the microbial communities were correlated to the toxicity test results, pointing to the need for further research regarding identification of the microbial flora. This can improve the treatment processes to optimize management of OMW for environmentally safe discharge. (C) 2010 Society of Chemical Industry