Genes encoding enzymes that catalyze butyryl-CoA formation from acetyl-CoA in a type II strain of Butyrivibrio fibrisolvens were analyzed. The genes encoding thiolase, beta-hydroxybutyryl-CoA dehydrogenase, butyryl-CoA dehydrogenase, and electron transfer flavoproteins were clustered, but the crotonase gene was not present in this region. The deduced amino acid sequences of these enzymes were similar to those of clostridia. The clustered genes were shown to be cotranscribed. The rate of butyrate production increased with an increase in acetate concentration in the medium up to 5 mm, suggesting that the butyryl-CoA/acetate CoA transferase reaction limits butyrate production. Transcription of the clustered genes was not affected by acetate concentration, suggesting that acetate does not affect the synthesis of enzymes involved in butyryl-CoA formation. These results confirm that acetate stimulates butyrate production by acting as a CoA acceptor in the butyryl-CoA/acetate CoA transferase reaction.