This study designed integrated constructed wetland–microbial fuel cell (CW–MFC) systems using activated carbon (AC) as both CW substrates and MFC anodes and investigated the structure-activity relationship of six kinds of commercial columnar AC, as well as the organics and nitrogen removal, microbial activity and diversity of CW–MFCs. Results showed that the nitrogen adsorption by AC tended to be a linear process in which physical adsorption played a leading role and micropores made great contributions. A higher specific surface area, developed mesopores, and oxygen functionalities were conducive to the capacitance properties of AC, while a higher specific surface area and developed micropores were conducive to reduce material resistance and improve ion permeability. Coconut-shell-based AC had both excellent nitrogen adsorption capacity and electrochemical properties, making it ideal as both CW substrates and MFC anodes for CW–MFCs. The electricity generation, coulombic efficiency, internal resistance, and organics and nitrogen removal of CW–MFCs were positively correlated with the total depth of AC anodes. The total depth of AC anodes can be determined based on the influent organics/nitrogen loadings and organics/nitrogen removal load of AC, and a relatively smaller depth of a single AC anode (5 cm) was recommended. The MFC effectively improved the enzymatic activity (by 10.33% dehydrogenase, 8.72% catalase, and 7.35% ammonia monooxygenase), nitrification/denitrification intensity (by 9.53%/6.68%), and microbial diversity (by 1.64–4.07%) of AC (MFC anodes) in CW–MFCs, while the depth of a single AC anode barely influenced the microbial activity and diversity. MFCs increased COD and NH3-N removal in CW–MFCs by 11.60% and 3.4%, respectively. The increased total adsorption capacity of AC with the increase of its total depth narrowed the difference in COD removal resulting from the promotion of MFCs on organics degradation. MFCs increased TN removal in CW–MFCs by 5.29% through promoting denitrification in cathodes and enhancing NH3-N assimilation in anodes. The phyla of EAB (Proteobacteria, Bacteroidetes, Firmicutes, and Acidobacteria) and genera of EAB (Citrobacter, Geobacter, and Pseudomonas) accounted for 85–86% and 15.58–16.64% of the microbial community on AC anodes in CW–MFCs, respectively.
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