The present study investigates the diversification and dynamic behavior of a multi-population microfluidic microbial fuel cell (MFC) as a biosensor. The cost effective microfluidic MFC coupled to a comprehensive model, presents a novel platform for monitoring chemical and biological phenomena. The importance of competition among different microbial groups, hierarchical biochemical processes, bacterial chemotaxis and different mechanisms of electron transfer were significant considerations in the present model. The validation of the model using experimental data from a microfluidic MFC shows an appropriate match with the hierarchal biodegradation processes of a complex substrate as well as development of bacterial chemotaxis during multi-population biofilm formation under real conditions. Microfluidic MFC performance, including temporal and spatial distribution of different microbial group concentrations in the biofilm and anolyte bulk, the competitive behavior of different species, bacterial transport parameters and bioelectrochemical characteristics are also assessed.