The volatile fatty acids (VFAs) productions, as well as hydrolases activities, microbial communities, and homoacetogens, of mesophilic and thermophilic sludge anaerobic fermentation were investigated to reveal the microbial responses to different fermentation temperatures. Thermophilic fermentation led to 10-fold more accumulation of VFAs compared to mesophilic fermentation. alpha-glucosidase and protease had much higher activities in thermophilic reactor, especially protease. Illumina sequencing manifested that raising fermentation temperature increased the abundances of Clostridiaceae, Microthrixaceae and Thermotogaceae, which could facilitate either hydrolysis or acidification. Real-time PCR analysis demonstrated that under thermophilic condition the relative abundance of homoacetogens increased in batch tests and reached higher level at stable fermentation, whereas under mesophilic condition it only increased slightly in batch tests. Therefore, higher fermentation temperature increased the activities of key hydrolases, raised the proportions of bacteria involved in hydrolysis and acidification, and promoted the relative abundance of homoacetogens, which all resulted in higher VFAs production. (C) 2014 Elsevier Ltd. All rights reserved.