A mesophilic anaerobic moving bed biofilm reactor (MBBR) was operated to evaluate the effect of sulfate addition on methane production and sulfate reduction using acetate as the sole carbon source. The results show that at the organic loading rate of 4.0 g TOC/L/day, the TOC removal efficiencies and the biogas production rates achieved over 95 % and 7000 mL/L/day without sulfate, respectively, and slightly decreased with sulfate addition (500-800 mg/L). Methane production capacities were not influenced significantly with the addition of sulfate, while sulfate reduction efficiencies were not stable with 23-87 % in the acetate-fed reactor. Fluorescent in situ hybridization (FISH) was used to analyze the functional microbial compositions of acetate-degrading methane-producing bacteria (MPB) and sulfate-reducing bacteria (SRB) in the reactor. The results found that as the increase of sulfate concentration, the proportion of Methanomicrobiales increased up to 58 ± 2 %, while Methanosaeta and Methanosarcina decreased. The dominant methanogens shifted into hydrogenotrophic methanogens from even distribution of acetoclastic and hydrogenotrophic methanogens. When hydrogenotrophic methanogens were dominant, sulfate reduction efficiency was high, while sulfate reduction efficiency was low as acetoclastic methanogens were dominant.