Arsenite-Oxidizing Hydrogenobaculum Strain Isolated from an Acid-Sulfate-Chloride Geothermal Spring in Yellowstone National Park

Abstract

An arsenite-oxidizing Hydrogenobaculum strain was isolated from a geothermal spring in Yellowstone National Park, Wyo., that was previously shown to contain microbial populations engaged in arsenite oxidation. The isolate was sensitive to both arsenite and arsenate and behaved as an obligate chemolithoautotroph that used H 2 as its sole energy source and had an optimum temperature of 55 to 60°C and an optimum pH of 3.0. The arsenite oxidation in this organism displayed saturation kinetics and was strongly inhibited by H 2 S.Arsenite [As(III)] is often the predominant valence of inorganic arsenic in geothermal source waters, although arsenate [As(V)] can also be present, with As(V)/As(III) ratios varying among different springs due to mixing with meteoric surface waters prior to discharge (3,12,19). However, subsequent to discharge, As(V)/As(III) ratios in the spring water can also be significantly influenced by redox transformations (10, 12), which are well documented for microorganisms (2,4,5,6,10,11,16,17). As(V) reduction is widespread among prokaryotes, occurring when As(V) is utilized as an electron acceptor for anaerobic or microaerobic respiration (13) or as part of a detoxification strategy (8). As(III) oxidation has likewise been observed in various organisms, where it has also been viewed as an apparent detoxification mechanism (1, 14, 15) or as a source of energy to support chemolithoautotrophic growth (1, 17).We previously documented rapid microbial oxidation of As(III) in an acid-sulfate-chloride-type geothermal spring in Norris Geyser Basin, Yellowstone National Park (9). This shallow spring is fed by a nearly constant geothermal source water (63°C, pH 3.1) containing ϳ35 M As(III). The prokaryote microbial community in this spring forms visually and chemically distinguishable mats. A filamentous yellow microbial mat containing visible amounts of S 0 (63 to 60°C) is present 0 to ϳ3.5 m from the spring source and changes to a brown, Fe(III) oxyhydroxide filamentous microbial mat (51 to 55°C) at ϳ3.5 to 5 m from the spring source (9). Chemical analysis of the aqueous and solid phases documented high rates of As(III) oxidation in the brown mat region, and the role of microorganisms in As(III) oxidation was confirmed in assays that showed no As(III) oxidation in the formaldehyde-killed samples (9). The PCR-generated 16S ribosomal DNA clone libraries representing the yellow and brown mat regions were dominated by Hydrogenobaculum-and Desulfurella-like sequences (7). However, since the phylogenetic data could not predict which population(s) was involved in the As(III) oxidation, the present study was conducted to initiate isolation and characterization of the As(III)-oxidizing microorganism(s) in this spring for use in modeling important and dominant biogeochemical features found in this spring type.Sampling, enrichment, and isolation. Brown microbial mat material was aseptically sampled and transferred to sterile 70-ml serum bottles and submerged with 35 ml of spring water sampled from above the mat. Th...

Arsenite Oxidase from Ralstonia sp. 22

“…Are these compounds true enzymatic inhibitors? A concentration of 60 M sulfide has been observed to completely stop As III oxidation in whole cells of Hydrogenobaculum at low pH (48), whereas sulfide has been reported to strongly enhance the As III oxidation in Mono Lake samples at high pH (58). However, did sulfide act on the Aro directly in these cases?…”

“…63, 64 for original works). More recently, the interaction between As III oxidation and sulfur metabolism was addressed (48,58,65), but it has not been studied at the molecular level prior to this work. Although ability of Aro to use cyt c 552 as electron acceptor may be interpreted as a possible cross-talk between sulfur and arsenic metabolisms, this merging of electron transfer pathways is not obligatory because Aro possesses a second, dedicated, acceptor, i.e.…”

Involvement of RpoN in Regulating Bacterial Arsenite Oxidation

“…Subsequent follow-up characterizations of this organism and this process failed to materialize; however, approximately 2 decades later, Santini et al (52) described the isolation and initial characterization of a Rhizobiumlike bacterium (strain NT-26) that could grow chemolithoautotrophically with As III as a sole electron donor for energy generation and with CO 2 as a sole carbon source. Soon thereafter, and in part stimulated by the massive arsenic poisoning disaster in Bangladesh (2), a series of studies initiated the characterization of microbial As III oxidation in natural environments, including geothermal springs (9,11,12,17,19,24,25,35,51) and soils (41); in mining-contaminated environments (6, 13, 40); and, most recently, in anoxic photosynthesis (21, 33). Likewise, progress has been made in the understanding of the biochemistry of the As III oxidase enzyme (1,14,37 oxidase structural genes were later cloned from the above-mentioned Rhizobium NT-26 organism (53).…”

“…Subsequent follow-up characterizations of this organism and this process failed to materialize; however, approximately 2 decades later, Santini et al (52) described the isolation and initial characterization of a Rhizobiumlike bacterium (strain NT-26) that could grow chemolithoautotrophically with As III as a sole electron donor for energy generation and with CO 2 as a sole carbon source. Soon thereafter, and in part stimulated by the massive arsenic poisoning disaster in Bangladesh (2), a series of studies initiated the characterization of microbial As III oxidation in natural environments, including geothermal springs (9,11,12,17,19,24,25,35,51) and soils (41); in mining-contaminated environments (6,13,40); and, most recently, in anoxic photosynthesis (21,33 (28,31) indicated the role and importance of the sensor kinase AioS and its putative regulatory partner AioR (a bacterial enhancer binding protein), direct proof of these two proteins working together as part of a putative As III signal perception and transduction cascade was just recently provided by Sardiwal et al (54), who demonstrated the autophosphorylation of an AioS component and the AioS-specific phosphorylation of AioR. Recently, our work has expanded this regulatory model to now include a third component, AioX, which is a periplasmic As III binding protein that is also essential for aioBA expression (39 …”

A Na + :H + Antiporter and a Molybdate Transporter Are Essential for Arsenite Oxidation in Agrobacterium tumefaciens

“…Analytical As chemistries were determined using techniques previously described (10). Briefly, cell suspensions were centrifuged and supernatants were filtered (0.22-m pore size) into two separate 15-ml bottles (5 ml each).…”