At the anode of a microbial fuel cell (MFC), denitrifying bacteria can coexist with exoelectrogens, enabling an MFC to achieve simultaneous nitrate reduction and electricity generation by the oxidization of organic matter. In this study, an anodic denitrifying MFC (MFC-D) was constructed by incorporating heterotrophic denitrification. This was compared to a control, MFC without denitrification (MFC-C). The results showed that MFC-D exhibited higher chemical oxygen demand (COD) removal and power density than MFC-C. Microbial community analysis showed that the proportion of bacteria of the genus Geobacter significantly decreased from 72.8% in MFC-C to 29.4% in MFC-D, and that denitrifiers in MFC-D increased by 36.8% in comparison with those in MFC-C. The COD used for denitrification was calculated to be 3.7 +/- 0.3 g COD/g NO3--N when MFC-D was fed with different COD/N ratios. The optimal proportion of COD/N in the MFC-D system was 5:1, at which the highest coulombic efficiency (CE) of electricity generation and anodic denitrification were obtained. The maximum voltage output was not inhibited at a low COD/N, but the cycle duration was decreased with decreasing COD/N. Additionally, the bacterial community was more diverse when complex organics were used as an electron donor or in a single-chamber MFC, which subsequently altered the electricity recovery and denitrification performance. This study provides a new strategy to improve the performance of MFCs in actual applications by the addition of denitrifiers to the anodic biofilms.