Excessive nitrogen fertilizer use has resulted in growing nitrate contamination of groundwater. In this study, an in situ bioelectrochemical reactor (isBER) reinforced with woodchips was developed for the treatment of actual nitrate-contaminated groundwater. During the 75-day experiment, the denitrification performance, grid permeability, and microbial community structure were investigated under different flow rates and current densities. The reactor achieved a remarkable nitrate removal efficiency of 97.6% ± 0.4% and a rate of 2.09 ± 0.14 mg-N/(L·h). These results were obtained at a temperature of 18.5 ± 0.8 °C, a current density of 350 mA/m², and a flow rate of 10 cm/d. Notably, the reactor can adapt to a wide flow-rate range of 5~20 cm/d and the operation proceeded smoothly without any blockages. Furthermore, the cathode module demonstrated enrichment of hydrogen autotrophic denitrifying bacteria (Pseudomonas, Stenotrophomonas) and heterotrophic denitrifying bacteria (Brucella, Enterobacteriaceae). Conversely, the anode module exhibited relatively high enrichment levels of aerobic microorganisms and lignin-degrading bacteria (Cellvibrio). The research results can provide novel insights and technical support for in situ remediation of groundwater nitrate contamination.