Microbial fuel cells (MFCs) can allow the treatment of organic pollutants resistant to conventional biological processes by electro-Fenton (EF) process performed in the cathodic compartment. However, EF usually results in a partial mineralization of pollutants. Here, we have studied the possible treatment of such organics in a MFC by a sequential cathodic and anodic treatment. In particular, the treatment of an aqueous solution of Acid Orange (AO7), a largely used azoic dye resistant to conventional biological processes, was performed in the cathodic compartment of a divided MFC by EF. The process allowed the total removal of the color and the partial removal of the TOC, due mainly to the formation of various carboxylic acids resistant to EF. The effluent of the cathodic compartment was fed to the anodic compartment in order to use the by-products of EF as organic sources for microorganisms. It was shown that the MFC could effectively work using such effluents as feed for the anodic compartment. The effect of the nature of the microorganisms on the process was also evaluated. It was shown that both Shewanella putrefaciens and Geobacter sulfurreducens can allow an effective treatment. In addition, an insect gut microbiota was tested for the first time.