A better understanding of voltage reversal phenomenon and its long-term effects on power generation are crucial to efficiently improve the voltage of microbial fuel cell (MFC) stacks. In this study, six MFCs with imbalanced performances were connected in series. After over 100 h of operation under voltage reversal conditions, increased turbidity and color were observed in the anolyte of the reversed MFC. In addition, the cyclic voltammogram of the anode changed form a typical catalytic current to a capacitance current. The scanning electron microscopy (SEM) showed that biofilm was dissociated from the anode surface, indicating that long-term operation under voltage reversal could damage the biofilm and ultimately resulted in the failure of MFC stacks. X-ray diffraction (XRD) and SEM equipped with energy dispersive X-ray spectrometry (EDX) analyses showed that the black particles in the anolyte was mostly carbon exfoliated from the anode, suggesting that carbon corrosion caused the biofilm failure. (C) 2017 Hydrogen Energy Publications LLC. Published by Elsevier Ltd. All rights reserved.