This paper evaluated the oxygen reduction reaction (ORR) in a microbial fuel cell (MFC) system by using chemically and physically activated electrospun carbon nanofibers (ACNFs) in an MFC and comparing their performance with that of plain carbon paper. The chemical and physical activation was carried out by KOH reagents and CO(2) gas to increase the electrode surface area and the catalytic activity. As a result, it was found that the MFC with the chemically activated carbon nanofibers (ACNFs) exhibited better catalytic activity than that of the physically activated ACNFs. Chemically ACNFs with 8 M KOH were found to be one of the most promising candidates for the ORR and could generate up to 3.17 times more power than that of the carbon paper. The ACNFs with 8 M KOH exhibited 78% more power generation than that of the physically activated ACNFs and exhibited 16% more power generation than the chemically activated ACNFs with 4 M KOH. The power per cost of ACNFs with 8 M KOH is 2.65 times greater than that of the traditionally used platinum cathode. Thus, ACNFs are a good alternative catalyst to Pt for MFCs. Copyright (C) 2011, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.