A two-stage 68degreesC/55degreesC anaerobic degradation process for treatment of cattle manure was studied. In batch experiments, an increase of the specific methane yield, ranging from 24% to 56%, was obtained when cattle manure and its fractions (fibers and liquid) were pretreated at 68degreesC for periods of 36, 108, and 168 h, and subsequently digested at 55degreesC. In a lab-scale experiment, the performance of a two-stage reactor system, consisting of a digester operating at 68degreesC with a hydraulic retention time (HRT) of 3 days, connected to a 55degreesC reactor with 12-day HRT, was compared with a conventional single-stage reactor running at 55degreesC with 15-days HRT. When an organic loading of 3 g volatile solids (VS) per liter per day was applied, the two-stage setup had a 6% to 8% higher specific methane yield and a 9% more effective VS-removal than the conventional single-stage reactor. The 68degreesC reactor generated 7% to 9% of the total amount of methane of the two-stage system and maintained a volatile fatty acids (VFA) concentration of 4.0 to 4.4 g acetate per liter. Population size and activity of aceticlastic methanogens, syntrophic bacteria, and hydrolytic/fermentative bacteria were significantly lower in the 68degreesC reactor than in the 55degreesC reactors. The density levels of methanogens utilizing H-2/CO2 or formate were, however, in the same range for all reactors, although the degradation of these substrates was significantly lower in the 68degreesC reactor than in the 55degreesC reactors. Temporal temperature gradient electrophoresis profiles (TTGE) of the 68degreesC reactor demonstrated a stable bacterial community along with a less divergent community of archaeal species. (C) 2004 Wiley Periodicals, Inc.