This study examines the performance of a microbial fuel cell (MFC) utilizing Shewanella bacteria through electrochemical impedance spectroscopy (EIS). Exo‐electrogen bacteria are key agents in an MFC. Shewanella sp. as a common exo‐electrogen bacteria can transfer electrons from the cell surface through different electron transfer mechanisms. In this work, EIS was used to probe the effects of biofilms of Shewanella sp. and the solution of 10% V/V Shewanella on the MFC performance. This research investigates the effects of both microbial biofilms and Shewanella bacterial solutions on MFC efficacy. Findings revealed that biofilm formation on the anode surface significantly reduces anode charge transfer resistance, thereby enhancing power generation. Notably, a 10% Shewanella solution resulted in a 25% higher power density compared to the biofilm. Furthermore, the MFC demonstrated up to 80% chemical oxygen demand removal efficiency in treating brewery wastewater. The study underscores the viability of Shewanella bacterial solutions as an efficient alternative to biofilms, emphasizing their role in improving MFC performance and wastewater treatment efficiency.