Technical Abstract: Hydrogen (H2) production from simulated cheese processing wastewater via anaerobic fermentation was conducted using mixed microbial communities under mesophilic conditions. In batch H2 fermentation experiments H2 yields of 8 and 10 mM/g-COD fed were achieved at food-to-microorganism (F/M) ratios of 1.0 and 1.5, respectively. Butyric, acetic, propionic, and valeric acids were the major volatile fatty acids (VFA) produced in the fermentation process. Continuous H2 fermentation experiments were also performed using a completely mixed reactor (CSTR). The pH of the bioreactor was controlled in a range of 4.0 to 5.0 by the addition of carbonate in the feed material. Maximum H2 yields were between 1.8 and 2.3 mM/g-COD fed for the loading rates (LR) tested with a hydraulic retention time (HRT) of 24 hr. Occasionally CH4 was produced in the biogas with concurrent reductions in H2 production; however, continuous H2 production was achieved for over three weeks at each LR. The 16S rDNA analysis of DNA extracted from the bioreactors during periods of high H2 production revealed that more than 50% of the bacteria present were members of the genus Lactobacillus and about 5% were Clostridia. When H2 production in the bioreactors decreased concurrent reductions in the genus Lactobacillus was also observed. Therefore, the microbial populations in the bioreactors were closely related to the conditions and performance of the bioreactors.