Leptothrix discophora SP-6, a type of manganese(Mn)-oxidizing bacteria, has been known to accumulate Mn oxides from the aqueous environment and thus play a key role in microbiologically influenced corrosion by increasing the electrochemical potential of steel and other metals. Similarly, this bacterium was found to modify the surface of glassy carbon in aqueous solution and increase its potential (i.e. ennoblement). In the latter case, biomineralized Mn oxides can be used as cathodic reactants for a new generation of microbial fuel cells featuring a bio-cathode. In this preliminary study, factors affecting the biofilm formation and biomineralization processes were examined. The inflow of air into the culture medium was found essential to sustain the ennoblement of substrate electrodes. The OCP and FESEM/EDS data indicated that a smoother initial substrate surface generally led to better ennoblement. Polarizing the carbon electrode at +500mVSCE for 15 minutes was found to facilitate the ennoblement on carbon electrodes, and so did the coating with a poly(L-lysine) layer. Independent of substrate material, initial surface roughness and pretreatment, there were three parameters in the EIS equivalent circuit that correlated well with the OCP indicating the level of ennoblement by L. discophora SP-6, i.e., electrolyte resistance, double-layer capacitance, and low-frequencies capacitance. These fascinating findings merit further investigation as they may shed light on the fundamental bacteria/substrate interactions and help advance the knowledge base needed for the engineering applications.