Direct interspecies electron transfer (DIET) is an alternative to interspecies H2/formate transfer as a mechanism for microbial species to cooperatively exchange electrons during syntrophic metabolism. To understand what specific properties contribute to DIET, studies were conducted with Pelobacter carbinolicus, a close relative of Geobacter metallireducens, which is capable of DIET. P. carbinolicus grew in co-culture with Geobacter sulfurreducens with ethanol as electron donor and fumarate as electron acceptor, conditions under which G. sulfurreducens formed direct electrical connections with G. metallireducens. In contrast to the cell aggregation associated with DIET, P. carbinolicus and G. sulfurreducens did not aggregate. Attempts to initiate co-cultures with a genetically modified strain of G. sulfurreducens incapable of both H2 and formate utilization were unsuccessful, whereas co-cultures readily grew with mutant strains capable of formate but not H2 uptake, or vice-versa. The hydrogenase mutant of G. sulfurreducens compensated, in co-cultures, with significantly increased formate-dehydrogenase gene expression. In contrast, the transcript abundance of a hydrogenase gene was comparable in co-cultures with the formate dehydrogenase mutant of G. sulfurreducens or wild-type, suggesting that H2 was the primary electron carrier in the wild-type co-cultures. Co-cultures were also initiated with strains of G. sulfurreducens that could not produce pili or OmcS, two essential components for DIET. The finding that P. carbinolicus exchanged electrons with G. sulfurreducens via interspecies transfer of H2/formate rather than DIET demonstrates that not all microorganisms that can grow syntrophically are capable of DIET and that closely related microorganisms may use significantly different strategies for interspecies electron exchange.