Arsenic is a carcinogenic compound widely distributed in the groundwater around the world. The fate of arsenic in groundwater depends on the activity of microorganisms either by oxidizing arsenite (AsIII), or by reducing arsenate (AsV). Because of the higher toxicity and mobility of AsIII compared to AsV, microbial-catalyzed oxidation of AsIII to AsV can lower the environmental impact of arsenic. Although aerobic AsIII-oxidizing bacteria are well known, anoxic oxidation of AsIII with nitrate as electron acceptor has also been shown to occur. In this study, three AsIII-oxidizing bacterial strains, Azoarcus sp. strain EC1, Azoarcus sp. strain EC3and Diaphorobacter sp. strain MC, have been characterized. Each strain was tested for its ability to oxidize AsIII with four different electron acceptors, nitrate, nitrite, chlorate and oxygen. Complete AsIII oxidation was achieved with both nitrate and oxygen, demonstrating the novel ability of these bacterial strains to oxidize AsIII in either anoxic or aerobic conditions. Nitrate was only reduced to nitrite. Different electron donors were used to study their suitability in supporting nitrate reduction. Hydrogen and acetate were readily utilized by all the cultures. The flexibility of these AsIII-oxidizing bacteria to use oxygen and nitrate to oxidize AsIII as well as organic and inorganic substrates as alternative electron donors explains their presence in non-arsenic-contaminated environments. The findings suggest that at least some AsIII-oxidizing bacteria are flexible with respect to electron-acceptors and electron-donors and that they are potentially widespread in low arsenic concentration environments.