This work aims to investigate the long-term behavior of interactions of electrochemically active bacteria in bioelectrochemical systems. The electrochemical performance and biofilm characteristics of pure cultures of Geobacter sulfurreducens and Shewanella oneidensis are being compared to a defined mixed culture of both organisms. While S. oneidensis pure cultures did not form cohesive and stable biofilms on graphite anodes and only yielded 0.034 ± 0.011 mA/cm 2 as maximum current density by feeding of each 5 mM lactate and acetate, G. sulfurreducens pure cultures formed 69 μm thick, area-wide biofilms with 10 mM acetate as initial substrate concentration and yielded a current of 0.39 ± 0.09 mA/cm 2. Compared to the latter, a defined mixed culture of both species was able to yield 38% higher maximum current densities of 0.54 ± 0.07 mA/cm 2 with each 5 mM lactate and acetate. This increase in current density was associated with a likewise increased thickness of the anodic biofilm to approximately 93 μm. It was further investigated whether a sessile incorporation of S. oneidensis into the mixed culture biofilm, which has been reported previously for short-term experiments, is long-term stable. The results demonstrate that S. oneidensis was not stably incorporated into the biofilm; rather, the planktonic presence of S. oneidensis has a positive effect on the biofilm growth of G. sulfurreducens and thus on current production.