An aciduric high-yielding hydrogen yeast named Candida maltosa HY-35 was screened, which can grow and produce hydrogen at pH 1.3. Further research was carried out batchwise to measure the hydrogen-producing ability of a mixed culture of C maltosa HY-35 and a facultative anaerobe Enterobacter aerogenes W-23. In this method, with the mixed culture of these two strains at 35 degrees C for 28 h, the hydrogen yield was 1735 ml/l, which was 17.15% and 119.90% higher than those of the monoculture of E. aerogenes W-23 and C. maltosa HY-35, respectively. Meanwhile, the average hydrogen production rate with the mixed culture was 261.1 ml/h/l, which was 7.85% and 146.23% higher than those of the monoculture of E aerogenes W-23 and C. maltosa HY-35, respectively. The results suggested that mixed culture of these two strains had the synergistic effect on hydrogen production. The optimum fermentation medium and operation conditions for hydrogen production with mixed culture were also investigated. (C) 2006 International Association for Hydrogen Energy. Published by Elsevier Ltd. All rights reserved.