In order to investigate the impact of organic matter type on the efficiency and microbial community structure during anaerobic digestion operation, an expanded granular sludge bed (EGSB) was applied to treat brewery spent grain hydrolysates (BSGH) and pig manure hydrolysates (PMH) that were pre-hydrolyzed by specific enzymes under thermophilic conditions. Results showed that after the organic matter of BSGH was altered by PMH, a series decrease of 40%, 75% and 25% was observed for the bulk COD removal, methane production and organic biomethanation rate, respectively. Meanwhile, the acetic acid concentration in the effluent increased from 50 mg/L to 3 700 mg/L. For the microbial community, the abundance of Firmicutes doubled after the substrate type changing, while Bacteroidetes decreased 50% instead. The quantity of methanogens dropped by 61% and the previously most abundant genus Methanosaeta was replaced by Methanobacterium.