The ␤-proteobacterial strain ULPAs1, isolated from an arsenic-contaminated environment, is able to efficiently oxidize arsenite [As(III)] to arsenate [As(V)]. Mutagenesis with a lacZ-based reporter transposon yielded two knockout derivatives deficient in arsenite oxidation. Sequence analysis of the DNA flanking the transposon insertions in the two mutants identified two adjacent open reading frames, named aoxA and aoxB, as well as a putative promoter upstream of the aoxA gene. Reverse transcription-PCR data indicated that these genes are organized in an operonic structure. The proteins encoded by aoxA and aoxB share 64 and 72% identity with the small Rieske subunit and the large subunit of the purified and crystallized arsenite oxidase of Alcaligenes faecalis, respectively (P. J. Ellis, T. Conrads, R. Hille, and P. Kuhn, Structure [Cambridge] 9:125-132, 2001). Importantly, almost all amino acids involved in cofactor interactions in both subunits of the A. faecalis enzyme were conserved in the corresponding sequences of strain ULPAs1. An additional Tat (twin-arginine translocation) signal peptide sequence was detected at the N terminus of the protein encoded by aoxA, strongly suggesting that the Tat pathway is involved in the translocation of the arsenite oxidase to its known periplasmic location.Arsenic is present in various environments, is released either by natural weathering of rocks or by anthropogenic sources (e.g., mining industries and agricultural practices), and is found in the oxidation states ϩ5 (arsenate), ϩ3 (arsenite), 0 (elemental arsenic), and Ϫ3 (arsine). Contamination of drinkingwater supplies with the inorganic soluble forms arsenite and arsenate has often been reported, and arsenic has been identified as a major risk for human health in different parts of the world (northeast India, Bangladesh, northwest United States) (31). The biogeochemical cycle of this element strongly depends on microbial transformation, which affects the mobility and the distribution of arsenic species in the environment (33, 41). Several bacteria involved in transformation processes comprising reduction, oxidation, and methylation of arsenic species have been described (8,11,26,36,40).The toxicological effects of arsenic are related to its chemical form and oxidation state; the organic forms are the less toxic. Among inorganic forms, As(III) is reported to be on average 100 times more toxic than the less mobile As(V) (25). Several remediation processes have been described for arsenic removal (19) based on chemical oxidation of arsenite to arsenate followed by alkaline precipitation (5, 15-17, 24). The major drawbacks of these processes are that they generate additional pollution and are expensive. This has led to the exploration of alternative methods for arsenic remediation based on its biological oxidation. Several arsenite-oxidizing bacteria have been isolated, starting with an Achromobacter strain in 1918 (14). Since then, different arsenite-oxidizing bacteria, including several Pseudomonas strains (18, 42-44), A...

Microbial biotransformations have a major impact on contamination by toxic elements, which threatens public health in developing and industrial countries. Finding a means of preserving natural environments—including ground and surface waters—from arsenic constitutes a major challenge facing modern society. Although this metalloid is ubiquitous on Earth, thus far no bacterium thriving in arsenic-contaminated environments has been fully characterized. In-depth exploration of the genome of the β-proteobacterium Herminiimonas arsenicoxydans with regard to physiology, genetics, and proteomics, revealed that it possesses heretofore unsuspected mechanisms for coping with arsenic. Aside from multiple biochemical processes such as arsenic oxidation, reduction, and efflux, H. arsenicoxydans also exhibits positive chemotaxis and motility towards arsenic and metalloid scavenging by exopolysaccharides. These observations demonstrate the existence of a novel strategy to efficiently colonize arsenic-rich environments, which extends beyond oxidoreduction reactions. Such a microbial mechanism of detoxification, which is possibly exploitable for bioremediation applications of contaminated sites, may have played a crucial role in the occupation of ancient ecological niches on earth.

A new primer set was designed to specifically amplify ca. 1,100 bp of aoxB genes encoding the As(III) oxidase catalytic subunit from taxonomically diverse aerobic As(III)-oxidizing bacteria. Comparative analysis of AoxB protein sequences showed variable conservation levels and highlighted the conservation of essential amino acids and structural motifs. AoxB phylogeny of pure strains showed well-discriminated taxonomic groups and was similar to 16S rRNA phylogeny. Alphaproteobacteria-, Betaproteobacteria-, and Gammaproteobacteria-related sequences were retrieved from environmental surveys, demonstrating their prevalence in mesophilic Ascontaminated soils. Our study underlines the usefulness of the aoxB gene as a functional marker of aerobic As(III) oxidizers.Arsenic (As) exists mainly in two toxic soluble forms, arsenite, As(III), and arsenate, As(V), with the latter tending to associate with some oxyhydroxides and clay minerals. The bacterial oxidation of As(III) can thus contribute to a natural attenuation of As contamination by decreasing As bioavailability. These properties have recently been used to develop a bioprocess for removing As from a mining effluent by using the activity of As-metabolizing bacteria indigenous to the contaminated site (4). The feasibility of such a process depends on a good knowledge of the ability of the indigenous microflora to oxidize As(III) and requires reliable methods for detecting, identifying, and monitoring As(III) oxidizers in the environment.More than 50 phylogenetically diverse As(III)-oxidizing strains distributed among 25 genera have been isolated from various environments so far. Bacterial aerobic As(III) oxidation is performed by a dedicated enzyme, the As(III) oxidase (1,36,40), which belongs to the dimethyl sulfoxide (DMSO) reductase of the molybdenum family (9). In Alcaligenes faecalis, it is an ␣ 1 ␤ 1 heterodimer comprising a large subunit incorporating a molybdenum center and a [3Fe-4S] cluster and a small subunit incorporating a Rieske-type [2Fe-2S] cluster (9). Genes encoding these subunits are cotranscribed as an operon and have been successively characterized in Herminiimonas arsenicoxydans (26), Rhizobium sp. strain , and Agrobacterium tumefaciens (21). They have also been found in the genome of Chloroflexus aurantiacus, on a plasmid in Thermus thermophilus, in two aerobic thermophilic As(III) oxidizers, and in the genome of strains for which the ability to oxidize As(III) has not been experimentally proven (27).Due to the polyphyly of As(III)-oxidizing bacteria, the aoxB gene encoding the catalytic subunit of the enzyme seems to be a valuable molecular marker for investigating its ecology and the potential of As(III) oxidation in the environment. To this end, a recent study described primers targeting the first quarter of the aoxB gene to detect its presence and expression in the environment and suggested that the gene is widely distributed among the Bacteria and also is widespread in soil-water systems containing As (16).In our present study, we designed...