Acetylacetoin synthase (AACSase) and acetylacetoin reductase (AACRase) are representative enzymes of the 2,3-butanediol cycle. After examining their induction conditions in various bacteria, the former was induced by acetoin and the latter by glucose. All strains carrying AACSase also had AACRase, but the reverse was not true. Therefore, AACSase indicates the existence of the cycle. Acetylacetoin (AAC) accumulation or the ratio of 2,3-butanediol isomer formed also indicated the presence of the cycle in bacteria. This cycle is present in some strains and not in others even for those belonging to the same species. The cycle was not always associated with the representative 2,3-butanediol-producing bacteria or bacterial sporogenesis as reported previously.