Extracellular electron transfer (EET) is a mechanism that enables microbes to respire solid-phase electron acceptors. These EET reactions most often occur in the absence of oxygen, since oxygen can act as a competitive electron acceptor for many facultative microbes. However, for Shewanella oneidensis MR-1, oxygen may increase biomass development, which could result in an overall increase in EET activity. Here, we studied the effect of oxygen on S. oneidensis MR-1 EET rates using bioelectrochemical systems (BESs). We utilized optically accessible BESs to monitor real-time biomass growth, and studied the per-cell EET rate as a function of oxygen and riboflavin concentrations in BESs of different design and operational conditions. Our results show that oxygen exposure promotes biomass development on the electrode, but significantly impairs per-cell EET rates even though current production does not always decrease with oxygen exposure. Additionally, our results indicated that oxygen can affect the role of riboflavin in EET. Under anaerobic conditions, both current density and per-cell EET rate increase with the riboflavin concentration. However, as the dissolved oxygen (DO) value increased to 0.42 mg/L, riboflavin showed very limited enhancement on per-cell EET rate and current generation. Since it is known that oxygen can promote flavins secretion in S. oneidensis, the role of riboflavin may change under anaerobic and aerobic conditions. (C) 2016 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.