The hydrogenase and formate dehydrogenase levels in Syntrophobacter fumaroxidans and Methanospirillum hungatei were studied in syntrophic propionate-oxidizing cultures and compared to the levels in axenic cultures of both organisms. Cells grown syntrophically were separated from each other by Percoll gradient centrifugation. In S. fumaroxidans both formate dehydrogenase and hydrogenase levels were highest in cells which were grown syntrophically, while the formate-H 2 lyase activities were comparable under the conditions tested. In M. hungatei the formate dehydrogenase and formate-H 2 lyase levels were highest in cells grown syntrophically, while the hydrogenase levels in syntrophically grown cells were comparable to those in cells grown on formate. Reconstituted syntrophic cultures from axenic cultures immediately resumed syntrophic growth, and the calculated growth rates of these cultures were highest for cells which were inoculated from the axenic S. fumaroxidans cultures that exhibited the highest formate dehydrogenase activities. The results suggest that formate is the preferred electron carrier in syntrophic propionate-oxidizing cocultures of S. fumaroxidans and M. hungatei.Methanogenic decomposition of complex organic matter is a widespread process, which accounts for a large fraction of the global methane emission (15). Examples of natural methanogenic habitats are freshwater environments such as wetlands, sediments, and rice paddy fields, as well as intestinal tracts of higher animals and insects (4,6,8,21,28). Methanogenic processes can be applied to treat industrial wastewaters in high-rate anoxic bioreactors (16,19,33). The microorganisms involved in methanogenic decomposition are usually immobilized in granular aggregates or biofilms, which is essential for a high conversion rate (23) and prevents biomass from being washed out of the reactor.The amount of energy available in methanogenic conversions is small, and therefore the microorganisms involved are forced to cooperate syntrophically (24, 28). In particular, oxidation of intermediary reduced organic compounds, such as ethanol, butyrate, and propionate, is energetically unfavorable. Nevertheless, the methanogens involved keep the concentrations of the oxidation products, acetate and H 2 (or formate), low enough to create a situation in which all partners involved gain energy. To dispose of reducing equivalents, acetogens reduce protons or bicarbonate. Both H 2 and formate have been proposed as electron carriers in syntrophic degradation. Several studies have provided evidence for H 2 transfer by demonstrating syntrophic growth with methanogens that oxidize only H 2 . Schmidt and Ahring (25) studied interspecies electron transfer in granules from a mesophilic anaerobic sludge bed reactor, and concluded that formate transfer was not important during propionate and butyrate oxidation in this system.On the other hand, Thiele and Zeikus (29) provided evidence that interspecies formate transfer was the dominant mechanism in a whey-processing digester,...