A microbial fuel cell (MFC) is a very promising way to remove organic pollutants. Hexachlorobenzene (HCB) is a widely used agricultural pesticide. In this study, single-chamber and membrane-less soil MFCs were constructed. The HCB was degraded to pentachlorobenzene (PeCB), tetrachlorobenzene (TeCB), and trichlorobenzene (TCB) in sequence by a reductive dechlorination process in soil MFCs. The influences of the external resistance, concentration of phosphate buffer, and electrode spacing in soil MFCs on the degradation rate and removal efficiency of HCB were analyzed. The results showed that the degradation rate and removal efficiency of HCB were increased when the external resistance decreased from 2000 to 20 (1, and also when the concentration of phosphate buffer increased. The anode area played a significant role in dechlorination of HCB. Altering the spacing of the reducing electrode resulted in a lower ohmic resistance in the soil MFCs. The ohmic resistance was negatively correlated with the removal efficiency and degradation rate (P < 0.05). In conclusion, HCB removal efficiency could be enhanced by soil MFCs, the performance of which was improved by a decrease in external resistance and internal resistance, and an increase in phosphate buffer concentration, rather than just by shortening the electrode spacing.