The performance of dual chambered mediator-less microbial fuel cell (MFC) operated under batch mode was evaluated under different operating temperatures, ranging between 20 and 55 °C, with step increase in temperature of 5 °C. Synthetic wastewater with sucrose as carbon source having chemical oxygen demand (COD) of 519–555 mg/L was used in the study. Temperature was a crucial factor in the performance of MFCs for both COD removal and electricity production. The MFC demonstrated highest COD removal efficiency of 84% and power density normalized to the anode surface area of 34.38 mW/m2 at operating temperature of 40 °C. Higher VSS to SS ratio was observed at the operating temperature between 35 and 45 °C. Under different operating temperatures the observed sludge yield was in the range of 0.05 to 0.14 g VSS/g COD removed. The maximum Coulombic and energy efficiencies were obtained at 40 °C, with values of 7.39 and 13.14%, respectively. Internal resistance of the MFC decreased with increase in operating temperature. Maximum internal resistance of 1,150 Ω was observed when the MFC was operated at 20 °C; whereas the minimum internal resistance (552 Ω) was observed at 55 °C.