In the present work two novel two-stage hydrogen production processes from olive mill wastewater (OMW) have been introduced. The first two-stage process involved dark-fermentation followed by a photofermentation process. Dark-fermentation by activated sludge cultures and photofermentation by Rhodobacter sphaeroides O.U.001 were both performed in 55 ml glass vessels, under anaerobic conditions. In some cases of dark-fermentation, activated sludge was initially acclimatized to the OMW to provide the adaptation of microorganisms to the extreme conditions of OMW. The highest hydrogen production potential obtained was 291H(2)/l(OMW) after photofermentation with 50% (v/v) effluent of dark fermentation with activated sludge. Photofermentation with 50% (v/v) effluent of dark fermentation with acclimated activated sludge had the highest hydrogen production rate (0.00811(-1) h(-1)). The second two-stage process involved a clay treatment step followed by photofermentation by R. sphaeroides O.U.001. Photofermentation with the effluent of the clay pretreatment process (4% (v/v)) gives the highest hydrogen production potential (351H(2)/l(OMW)), light conversion efficiency (0.42%) and COD conversion efficiency (52%). It was concluded that both pretreatment processes enhanced the photofermentative hydrogen production process. Moreover, hydrogen could be produced with highly concentrated OMW. Two-stage processes developed in the present investigation have a high potential for solving the environmental problems caused by OMW. (c) 2006 International Association for Hydrogen Energy. Published by Elsevier Ltd. All rights reserved.