The ars Detoxification System Is Advantageous but Not Required for As(V) Respiration by the Genetically Tractable Shewanella Species Strain ANA-3

Abstract

Arsenate [As(V); HAsO 42؊ ] respiration by bacteria is poorly understood at the molecular level largely due to a paucity of genetically tractable organisms with this metabolic capability. We report here the isolation of a new As(V)-respiring strain (ANA-3) that is phylogenetically related to members of the genus Shewanella and that also provides a useful model system with which to explore the molecular basis of As(V) respiration. This gram-negative strain stoichiometrically couples the oxidation of lactate to acetate with the reduction of As(V) to arsenite [As(III); HAsO 2 ]. The generation time and lactate molar growth yield (Y lactate ) are 2.8 h and 10.0 g of cells mol of lactate ؊1 , respectively, when it is grown anaerobically on lactate and As(V). ANA-3 uses a wide variety of terminal electron acceptors, including oxygen, soluble ferric iron, oxides of iron and manganese, nitrate, fumarate, the humic acid functional analog 2,6-anthraquinone disulfonate, and thiosulfate. ANA-3 also reduces As(V) to As(III) in the presence of oxygen and resists high concentrations of As(III) (up to 10 mM) when grown under either aerobic or anaerobic conditions. ANA-3 possesses an ars operon (arsDABC) that allows it to resist high levels of As(III); this operon also confers resistance to the As-sensitive strains Shewanella oneidensis MR-1 and Escherichia coli AW3110. When the gene encoding the As(III) efflux pump, arsB, is inactivated in ANA-3 by a polar mutation that also eliminates the expression of arsC, which encodes an As(V) reductase, the resulting As(III)-sensitive strain still respires As(V); however, the generation time and the Y lactate value are two-and threefold lower, respectively, than those of the wild type. These results suggest that ArsB and ArsC may be useful for As(V)-respiring bacteria in environments where As concentrations are high, but that neither is required for respiration.The contamination of groundwaters and surface waters with arsenic (As) is a major concern to public health in countries such as Bangladesh, China, Taiwan, Argentina, Chile, and the United States (40). Elevated As concentrations typically derive from the weathering of As-bearing minerals and/or from geothermal sources (2, 58). It is now known that a variety of microorganisms, including members of the Eukarya, Archaea, and Bacteria, influence As geochemistry in many locales throughout the world by virtue of their metabolism (31,35,53). These metabolic processes include oxidation (49, 58), reduction (1, 13), and methylation reactions (5) that strongly affect (and in some cases, control) As speciation in the environment. One process that is particularly intriguing is microbial respiration of arsenate [As(V); HAsO 4 Ϫ2 ]. In the absence of oxygen, microorganisms can gain energy by coupling the oxidation of organic material to As(V) reduction, resulting in the production of the highly toxic As compound, arsenite [As(III); HAsO 2 ]. As(V)-respiring organisms can affect water quality by catalyzing the mobilization of As(III) from sediments...

Electrically conductive bacterial nanowires produced by Shewanella oneidensis strain MR-1 and other microorganisms

“…The ligated product was transformed into E. coli EC100D pir-116 (EPICENTRE Biotechnologies) and plated on LB with gentamycin selection (15 g͞ml). The resulting recombinant plasmids were purified and used to transform E. coli ␤-2155 (26) or WM3064 (27). Plasmids were transferred from E. coli ␤-2155 mating strain to S. oneidensis MR-1 by conjugation, and primary integrants were selected by plating on LB medium containing 7.5 g͞ml gentamycin.…”

Directed networks reveal genomic barriers and DNA repair bypasses to lateral gene transfer among prokaryotes

“…ANA-3. This g-proteobacterium was isolated from an arsenate-treated wooden pier located in a brackish estuary (Saltikov et al 2003). The genus Shewanella usually resides in marine habitats, so that their link to this subset is probably due to gene exchange with aquatic Burkholderiales.…”

Genetic identification of a respiratory arsenate reductase

“…This organism contains two systems for reducing As(V). One is similar to the well conserved ars detoxification system from Escherichia coli plasmid R773 (11) and is advantageous, but not required, for respiratory As(V) reduction (8); a separate system appears to be required for As(V) respiration. In this report, we use genetic analysis to identify and describe the operon that encodes that latter system.…”

“…In response to this need, a new As(V)-respiring species, Shewanella strain ANA-3 that is amenable to genetic analysis, was recently isolated (8). This organism contains two systems for reducing As(V).…”

“…Although much of the As contamination derives from natural weathering and dissolution of As-bearing minerals, recognition that microorganisms can alter the mobility of As in natural waters through redox transformations (3) drove the discovery of the first arsenate [As(V)]-respiring bacterium nearly a decade ago (4). Since then, many more microorganisms that can reduce As(V) to arsenite [H 3 AsO 3 , As(III)] have been discovered (5)(6)(7)(8), but a mechanistic understanding of this metabolism has lagged. To date, only three studies have described the biochemistry of arsenate respiration (5,9,10), and detailed biochemical analyses have not been performed.…”