Aims: To isolate, identify, and characterize heterotrophic bacteria in acid-mine drainage that mediate oxidation of As(III). Methods and Results: Samples of acid-mine drainage were collected over a period of 14 months. Heterotrophic and non-obligatory acidophilic bacteria in the samples were cultured on a solid medium (pH 7.0-7.2), and three strains were isolated. The three different strains belong to the genus Thiomonas , and have more than 99% homology with the group Ynys1. Culturing in mineral media demonstrated that the isolated strains used thiosulphate as an energy source, and oxidized iron in the presence of thiosulphate. However, none of the strains were able to oxidize arsenic in the presence of thiosulphate, nor could they use iron or arsenic alone as an energy source. In vitro experiments demonstrated that two of the Thiomonas strains were able to oxidize more than 90% of the As(III) present in the acid-mine drainage, whereas no abiotic oxidation of arsenic occurred. Conclusions: Two strains of newly identified Thiomonas sp. found in acid-mine drainage are capable of oxidizing arsenic. Significance and Impact of Study: These results represent the first reported oxidation of arsenic by Thiomonas sp. Biologically mediated oxidation and subsequent immobilization of arsenic is of great interest for the remediation of contaminated mine sites.