A feasible and readily available source of hydrogenproducing acetogens (HPAs) was developed by obtaining microflora D83 dominated by butyrate-oxidizing HPAs through enrichment and subculture of anaerobic sludge with the use of butyric acid. The effect of bioaugmentation of D83 both on methane production through glucose fermentation and molasses wastewater treatment was evaluated. At a bacterial number-to-activated sludge ratio of 1:9, inoculation of microflora D83 could enhance methane yield and production rate from glucose by a factor of 2.1 and 2.0, respectively. In treatment of normal molasses wastewater, chemical oxygen demand (COD) removal efficiency improved from 75.6% to 88.2%, and the accumulated methane yield and methane yield obtained through COD removal increased by a factor of 2.3 and 2.0, respectively. Hydrogen-producing acetogenesis is apparently a rate-limiting step in methanogenesis because bioaugmentation of HPAs during methane production improved not only hydrogen-producing acetogenesis but also hydrolysis/acidogenesis and methanogenesis.