Sluggish fermentation is known to occur in the industrial acetone-butanol fermentation of Clostridium acetobutylicum. The endogenous cause of sluggish fermentation was not clearly elucidated, though exogeneous causes, for example bacteria or temperate phage contamination were studied in detail. In this work, we indicated the sluggish fermentation would be occurred by the increase of degenrative strain, that is a spontaneous mutant defected in NADH generation and fail to onset of solventogenesis, during serial subculturings. A degenerative strain co-cultivated with wild type strain increased up to 40-50% in co-cultivated population after 15-20 times of subculturing, and caused drastic decrease of solvent productivity.