Novel insights into the kinetics and mechanism of arsenopyrite bio-dissolution enhanced by pyrite.

Arsenopyrite and pyrite often coexist in metal deposits and tailings, thus simultaneous bioleaching of both sulfides has economic (as well as environmental) significance. Important targets in bio-oxidation operations are high solubilization rates and minimized accumulation of Fe(III)/As-bearing secondary products. This study investigated the role of pyrite bioleaching in the enhancement of arsenopyrite dissolution. At a pyrite to arsenopyrite mass ratio of 1:1, 93.6% of As and 93.0% of Fe were solubilized. The results show that pyrite bio-oxidation can promote arsenopyrite dissolution, enhance S0 bio-oxidation, and inhibit the formation of jarosites, tooeleite, and amorphous ferric arsenate. The dry weight of the pyrite & arsenopyrite residue was reduced by 95.1% after bioleaching, compared to the initial load, while only 5% weight loss was observed when pyrite was absent. A biofilm was formed on the arsenopyrite surface in the presence of pyrite, while a dense passivation layer was observed in the absence of pyrite. As(III) (as As2O3) was a dominant As species in the pyrite & arsenopyrite residue. Novel and detailed findings are presented on arsenopyrite bio-dissolution in the presence of pyrite, and the presented approach could contribute to the development of novel cost-effective extractive bioprocesses. ENVIRONMENTAL IMPLICATION: The oxidation of arsenopyrite presents significant environmental hazards, as it can contribute to acid mine drainage generation and arsenic mobilization from sulfidic mine wastes. Bioleaching is a proven cost-effective and environmentally friendly extractive technology, which has been applied for decades in metal recovery from minerals or tailings. In this work, efficient extraction of arsenic from arsenopyrite bioleaching was presented through coupling the process with bio-oxidation of pyrite, resulting in lowered accumulation of hazardous and metastable Fe(III)/As-bearing secondary phases. The results could help improve current biomining operations and/or contribute to the development of novel cost-effective bioprocesses for metal extraction.

Metal recovery from spent lithium-ion batteries via two-step bioleaching using adapted chemolithotrophs from an acidic mine pit lake.

The demand for lithium-ion batteries (LIBs) has dramatically increased in recent years due to their application in various electronic devices and electric vehicles (EVs). Great amount of LIB waste is generated, most of which ends up in landfills. LIB wastes contain substantial amounts of critical metals (such as Li, Co, Ni, Mn, and Cu) and can therefore serve as valuable secondary sources of these metals. Metal recovery from the black mass (shredded spent LIBs) can be achieved via bioleaching, a microbiology-based technology that is considered to be environmentally friendly, due to its lower costs and energy consumption compared to conventional pyrometallurgy or hydrometallurgy. However, the growth and metabolism of bioleaching microorganisms can be inhibited by dissolved metals. In this study, the indigenous acidophilic chemolithotrophs in a sediment from a highly acidic and metal-contaminated mine pit lake were enriched in a selective medium containing iron, sulfur, or both electron donors. The enriched culture with the highest growth and oxidation rate and the lowest microbial diversity (dominated by Acidithiobacillus and Alicyclobacillus spp. utilizing both electron donors) was then gradually adapted to increasing concentrations of Li+, Co2+, Ni2+, Mn2+, and Cu2+. Finally, up to 100% recovery rates of Li, Co, Ni, Mn, and Al were achieved via two-step bioleaching using the adapted culture, resulting in more effective metal extraction compared to bioleaching with a non-adapted culture and abiotic control.

Microbiological and geochemical characterization of As-bearing tailings and underlying sediments.

Over the past 100 years, extensive oxidation of As-bearing sulfide-rich tailings from the abandoned Long Lake Gold Mine (Canada) has resulted in the formation of acid mine drainage (pH 2.0-3.9) containing high concentrations of dissolved As (∼400 mg L-1), SO42-, Fe and other metals. Dissolved As is predominantly present as As(III), with increased As(V) near the tailings surface. Pore-gas O2 is depleted to < 1 vol% in the upper 30-80 cm of the tailings profile. The primary sulfides, pyrite and arsenopyrite, are highly oxidized in the upper portions of the tailings. Elevated proportions of sulfide-oxidizing prokaryotes are present in this zone (mean 32.3% of total reads). The tailings are underlain by sediments rich in organic C. Enrichment in δ34S-SO4 in pore-water samples in the organic C-rich zone is consistent with dissimilatory sulfate reduction. Synchrotron-based spectroscopy indicates an abundance of ferric arsenate phases near the impoundment surface and the presence of secondary arsenic sulfides in the organic-C beneath the tailings. The persistence of elevated As concentrations beneath the tailings indicates precipitation of secondary As sulfides is not sufficient to completely remove dissolved As. The oxidation of sulfides and release of As is expected to continue for decades. The findings will inform future remediation efforts and provide a foundation for the long-term monitoring of the effectiveness of the remediation program.

Microbial processes with the potential to mobilize As from a circumneutral-pH mixture of flotation and roaster tailings.

The Northwest Tailings Containment Area at the inactive Giant Mine (Canada) contains a complex mixture of arsenic-containing substances, including flotation tailings (84.8 wt%; with 0.4 wt% residual S), roaster calcine wastes (14.4 wt% Fe oxides), and arsenic trioxide (0.8 wt%) derived from an electrostatic precipitator as well as As-containing water (21.3 ± 4.1 mg L-1 As) derived from the underground mine workings. In the vadose zone the tailings pore water has a pH of 7.6 and contains elevated metal(loid)s (2.37 ± 5.90 mg L-1 As); mineral oxidizers account for 2.5% of total 16S rRNA reads in solid samples. In the underlying saturated tailings, dissolved Fe and As concentrations increase with depth (up to 72 and 20 mg L-1, respectively), and the mean relative abundance of Fe(III)-reducers is 0.54% of total reads. The potential for As mobilization via both reductive and oxidative (bio)processes should be considered in Giant Mine remediation activities. The current remediation plan includes installation of an engineered cover that incorporates a geosynthetic barrier layer.

Mineralogy affects prokaryotic community composition in an acidic metal mine.

The microbial ecology of acidic mine and sulfide cave ecosystems is well characterised with respect to aquatic communities, typically revealing low taxonomic complexity and dominance by a relatively limited number of cosmopolitan acidophilic bacterial and archaeal taxa. Whilst pH, temperature, and geochemistry are recognised drivers of diversity in these ecosystems, the specific question of a possible influence of substratum mineralogy on microbial community composition remains unanswered. Here we address this void, using 81 subterranean mineral samples from a low temperature abandoned, acidic, sulfide ore mine system at Mynydd Parys (Parys Mountain in English), Wales, UK. Four primary and 15 secondary minerals were identified via x-ray diffraction, each sample containing a maximum of five and an average of two minerals. The mineralogy of primary (e.g. pyrite and quartz) and secondary (e.g. melanterite and pisanite) minerals was significantly correlated with prokaryotic community structure at multiple taxonomic levels, implying that the mineralosphere effect reported in less extreme terrestrial environments is also implicated in driving prokaryotic community composition in extremely acidic, base metal-bearing sulfide mineralisation at Mynydd Parys. Twenty phyla were identified, nine of which were abundant (mean relative abundance >1%). While taxa characteristic of acidic mines were detected, for example Leptospirillum (phylum Nitrospirae), Acidithiobacillus (phylum Proteobacteria), Sulfobacillus (phylum Firmicutes) and Ferroplasma (phylum Euryarchaeota), their abundance in individual samples was highly variable. Indeed, in the majority of the 81 samples investigated the abundance of these and other typical acidic mine taxa was low, with 25% of samples devoid of sequences from recognised acidic mine taxa. Most notable amongst the bacterial taxa not previously reported in such environments were the recently cultivated Muribaculaceae family (phylum Bacteroidetes), which often dominated Mynydd Parys samples regardless of their mineralogical content. Our results pose further questions regarding the mechanisms by which taxa not previously reported in such extreme environments appear to survive in Mynydd Parys, opening up research pathways for exploring the biodiversity drivers underlying microbial community composition and function in extremely acidic mine environments.

Sulfoacidibacillus ferrooxidans, gen. nov., sp. nov., Sulfoacidibacillus thermotolerans, gen. nov., sp. nov., and Ferroacidibacillus organovorans, gen. nov., sp. nov.: Extremely acidophilic chemolitho-heterotrophic Firmicutes.

Ten strains of extremely acidophilic bacteria, isolated from different environments form a distinct monophyletic clade within the phylum Firmicutes. Comparison of complete genomes of the proposed type strains confirm that they comprise two genera (proposed names Sulfoacidibacillus and Ferroacidibacillus), and at least three species (Sulfoacidibacillus ferrooxidans, Sulfoacidibacillus thermotolerans and Ferroacidibacillus organovorans). The bacterial strains share some physiological traits, including catalysing the dissimilatory oxidation and reduction of iron, and in being obligately heterotrophic. Both species of Sulfoacidibacillus are also able to oxidise elemental sulfur and tetrathionate. Both S. ferrooxidans and Ferroacidibacillus spp. are mesophilic, while S. thermotolerans isolates are moderate thermophiles. The isolates display different degrees of acid-tolerance: Ferroacidibacillus spp. are the most acid-sensitive while the type strain of S. ferrooxidans grows at pH 0.9. MK7 was detected as the sole menaquinone present in all three nominated type strains, and their peptidoglycans all contain meso-2,6 diaminopimelic acid type A1γ. The chromosomal DNA of the strains examined contain between 44 and 52 mol% G + C. The nominated type strains of the new species are S. ferrooxidans S0ABT (= DSM 105355T = JCM 33225T); S. thermotolerans Y002T (= ATCC TSD-104T = JCM 31946T); F. organovorans SLC66T (= ATCC TSD-103T = JCM 31945T).

Efficient dealkalization of red mud and recovery of valuable metals by a sulfur-oxidizing bacterium.

Red mud (RM) is a highly alkaline polymetallic waste generated via the Bayer process during alumina production. It contains metals that are critical for a sustainable development of modern society. Due to a shortage of global resources of many metals, efficient large-scale processing of RM has been receiving increasing attention from both researchers and industry. This study investigated the solubilization of metals from RM, together with RM dealkalization, via sulfur (S0) oxidation catalyzed by the moderately thermophilic bacterium Sulfobacillus thermosulfidooxidans. Optimization of the bioleaching process was conducted in shake flasks and 5-L bioreactors, with varying S0:RM mass ratios and aeration rates. The ICP analysis was used to monitor the concentrations of dissolved elements from RM, and solid residues were analyzed for surface morphology, phase composition, and Na distribution using the SEM, XRD, and STXM techniques, respectively. The results show that highest metal recoveries (89% of Al, 84% of Ce, and 91% of Y) were achieved with the S0:RM mass ratio of 2:1 and aeration rate of 1 L/min. Additionally, effective dealkalization of RM was achieved under the above conditions, based on the high rates (>95%) of Na, K, and Ca dissolution. This study proves the feasibility of using bacterially catalyzed S0 oxidation to simultaneously dealkalize RM and efficiently extract valuable metals from the amassing industrial waste.

Release and fate of As mobilized via bio-oxidation of arsenopyrite in acid mine drainage: Importance of As/Fe/S speciation and As(III) immobilization.

Mining activities expose sulfidic minerals including arsenopyrite (FeAsS) to acid mine drainage (AMD). The subsequent release of toxic arsenic (As) can have great negative implications for the environment and human health. This study investigated the evolution of secondary products and As speciation transformations during arsenopyrite bio-oxidation in AMD collected from a polymetallic mine. Immobilization of the As solubilized via arsenopyrite bio-oxidation using red mud (RM) was also studied. The results show that the high ionic strength (concentrations of dissolved Fe3+, SO42-, and Ca2+ reached values up to 0.75, 3.38, and 0.35 g/L, respectively) and redox potential (up to +621 mV) of AMD (caused primarily by Fe3+) enhanced the dissolution of arsenopyrite. A high [Fe]aq/[As]aq ratio in the AMD favored the precipitation of tooeleite during arsenopyrite bio-oxidation, and the formation of other poorly crystalline products such as schwertmannite and amorphous ferric arsenate also contributed to As immobilization. Bacterial cells served as important nucleation sites for the precipitation of mineral phases. Arsenopyrite completely dissolved after 12 days of bio-oxidation in AMD and the [As]aq (mainly present as As(III)) reached 1.92 g/L, while a greater [As]aq was observed in a basal salts medium (BSM) assay (reaching 3.02 g/L). An RM addition significantly promoted As(III) immobilization, with final [As(III)]aq decreasing to 0.16 and 1.43 g/L in AMD and BSM assays respectively. No oxidation of As(III) was detected during the immobilization process. These findings can help predict As release from arsenopyrite on contact with AMD and, on a broader scale, assist in designing remediation and treatment strategies to mitigate As contamination in mining.

Draft Genome Sequence of Firmicutes Strain S0AB, a Heterotrophic Iron/Sulfur-Oxidizing Extreme Acidophile.

The draft whole-genome sequence of the extremely acidophilic and novel Firmicutes strain S0AB is reported. The genome comprises 3.3 Mbp and has a GC content of 43.72%. In total, 3,240 protein-coding genes, 56 tRNA genes, and 11 rRNA genes were predicted.

Nickel Isotope Fractionation As an Indicator of Ni Sulfide Precipitation Associated with Microbially Mediated Sulfate Reduction.

Microbially mediated sulfate reduction is a promising cost-effective and sustainable process utilized in permeable reactive barriers (PRB) and constructed wetlands to treat mine wastewater. Laboratory batch experiments were performed to evaluate nickel (Ni) isotope fractionation associated with precipitation of Ni-sulfides in the presence of the sulfate-reducing bacterium (SRB) Desulfovibrio desulfuricansT (DSM-642). Precipitates were collected anaerobically and characterized by synchrotron powder X-ray diffraction (PXRD), scanning electron microscopy combined with energy-dispersive X-ray spectroscopy (SEM-EDS), and transmission electron microscopy (TEM). Solid-phase analyses showed that the precipitates associated with bacteria attached to the serum bottle walls were characterized by enhanced size and crystallinity. Lighter Ni isotopes were preferentially concentrated in the solid phase, whereas the solution was enriched in heavier Ni isotopes compared to the input solution. This fractionation pattern was consistent with closed-system equilibrium isotope fractionation, yielding a fractionation factor of Δ60Nisolid-aq = -1.99‰. The Ni isotope fractionation measured in this study indicates multiple Ni reaction mechanisms occurring in the complex SRB-Ni system. The results from this study offer insights into Ni isotope fractionation during interaction with SRB and provide a foundation for the characterization and development of Ni stable isotopes as tracers in environmental applications.

Removal of arsenic and metals from groundwater impacted by mine waste using zero-valent iron and organic carbon: Laboratory column experiments.

Acid mine drainage and the associated contaminants, including As and metals, are ongoing environmental issues. Passive remediation technologies have the potential to remove As from mine waste effluents. A series of laboratory column experiments was conducted to evaluate the effectiveness of varying mixtures of organic carbon (OC), zero-valent iron (ZVI), and limestone for the treatment of As, metals, SO42-, and acidity in groundwater from an abandoned gold mine. The onset of bacterially-mediated SO42- reduction was indicated by a decrease in Eh, a decline in aqueous SO42- concentrations coupled with enrichment of δ34S, and the presence of sulfate-reducing bacteria and H2S. Removal of As was observed within the first 3 cm of reactive material, to values below 10 µg L-1, representing > 99.9% removal. An increase in pH from 3.5 to circumneutral values and removal of metals including Al, Cu, and Zn was also observed. Synchrotron results suggest As was removed through precipitation of As-crystalline phases such as realgar and orpiment, or through adsorption as As(V) on ferrihydrite. The results indicate the potential for a mixture of OC and ZVI to remove As from acidic, mine-impacted water.

Microbiology of a multi-layer biosolid/desulfurized tailings cover on a mill tailings impoundment.

The Strathcona Waste Water Treatment System (SWWTS; Sudbury, ON, Canada) has received mill tailings from Ni/Cu ore processing from 1970 to present. Demonstration-scale, multi-layer cover systems were installed on selected tailings deposition cells at the SWWTS. The cover systems are comprised of an upper layer of organic carbon-rich material, composed of a layer biosolids fertilizer along with composted municipal food and yard waste, then a layer of desulfurized, fine-grained tailings. Organic carbon components used in these covers promote microbial communities that consume O2, thus decreasing sulfide oxidation rates in the underlying tailings. The aim of this study was to investigate the microbiology of the cover systems and the underlying tailings, using a combination of culture-dependent (most probable number) and culture-independent (16S rRNA gene amplicon sequencing) techniques, and assess the impact of the organic component of the cover system four to six years after implementation. Most tailings samples were characterized by circumneutral bulk pH and low concentrations of dissolved metals. The presence of the organic cover resulted in elevated counts of sulfate-reducers (by two orders of magnitude, compared to control samples) immediately below the organic cover, as well as an increased abundance of heterotrophic species (∼108 cells g-1) at greater depth (∼4 m) in the tailings profile. Mineral-oxidizing microorganisms were also present in the tailings, with neutrophilic sulfur-oxidizers dominating the samples (mean ∼106 cells g-1). Relative abundances of sulfur- and/or iron-oxidizers determined by sequencing ranged from 0.5 to 18.3% of total reads (mean ∼5.6% in amended tailings) and indicated the presence of local microenvironments with ongoing sulfide oxidation. This work provides a detailed characterization of the microbiology of a tailings impoundment with an organic cover, highlighting the opportunities associated with monitoring microbial processes in such remediation systems.

A Model of Aerobic and Anaerobic Metabolism of Hydrogen in the Extremophile Acidithiobacillus ferrooxidans.

Hydrogen can serve as an electron donor for chemolithotrophic acidophiles, especially in the deep terrestrial subsurface and geothermal ecosystems. Nevertheless, the current knowledge of hydrogen utilization by mesophilic acidophiles is minimal. A multi-omics analysis was applied on Acidithiobacillus ferrooxidans growing on hydrogen, and a respiratory model was proposed. In the model, [NiFe] hydrogenases oxidize hydrogen to two protons and two electrons. The electrons are used to reduce membrane-soluble ubiquinone to ubiquinol. Genetically associated iron-sulfur proteins mediate electron relay from the hydrogenases to the ubiquinone pool. Under aerobic conditions, reduced ubiquinol transfers electrons to either cytochrome aa 3 oxidase via cytochrome bc 1 complex and cytochrome c 4 or the alternate directly to cytochrome bd oxidase, resulting in proton efflux and reduction of oxygen. Under anaerobic conditions, reduced ubiquinol transfers electrons to outer membrane cytochrome c (ferrireductase) via cytochrome bc 1 complex and a cascade of electron transporters (cytochrome c 4, cytochrome c 552, rusticyanin, and high potential iron-sulfur protein), resulting in proton efflux and reduction of ferric iron. The proton gradient generated by hydrogen oxidation maintains the membrane potential and allows the generation of ATP and NADH. These results further clarify the role of extremophiles in biogeochemical processes and their impact on the composition of the deep terrestrial subsurface.

Performance of a Geosynthetic-Clay-Liner Cover System at a Cu/Zn Mine Tailings Impoundment.

The abandoned Kam Kotia Mine (Canada) is undergoing remediation. A geosynthetic-clay-liner (GCL) cover system was installed in the Northern Impounded Tailings (NIT) area in 2008 to isolate acid-generating tailings from water and oxygen and to mitigate sulfide oxidation. The cover system includes a vegetated uppermost soil layer underlain by a granular protective layer (sand), a clay moisture-retaining layer, a GCL, a granular capillary-break material (cushion sand), and a crushed waste rock-capillary break layer installed above the tailings. The goal of this study was to characterize the microbiology of the covered tailings to assess the performance of the cover system for mitigating sulfide bio-oxidation. Tailings beneath the GCL were characterized by high sulfur and low carbon content. The bulk pH of the tailings pore water was circumneutral (∼5.5 to 7.3). Total genomic DNA was extracted from 36 samples recovered from the constituent layers of the cover system and the underlying tailings and was analyzed in triplicates using high-throughput amplicon sequencing of 16S rRNA genes. Iron-oxidizing, sulfur-oxidizing, sulfate-reducing, and aerobic heterotrophic microorganisms were enumerated by use of most probable number enumeration, which identified heterotrophs as the most numerous group of culturable microorganisms throughout the depth profile. Low relative abundances and viable counts of microorganisms that catalyze transformations of iron and sulfur in the covered tailings, compared to previous studies on unreclaimed tailings, indicate that sulfide oxidation rates have decreased due to the presence of the GCL. Characterization of the microbial community can provide a sensitive indicator for assessing the performance of remediation systems.IMPORTANCE Mining activities are accompanied by significant environmental and financial liabilities, including the release of acid mine drainage (AMD). AMD is caused by accelerated chemical and biological oxidation of sulfide minerals in mine wastes and is characterized by low pH and high concentrations of sulfate and metal(loid)s. Microorganisms assume important roles in the catalysis of redox reactions. Our research elucidates linkages among the biogeochemistry of mine wastes and remediation systems and microbial community and activity. This study assesses the performance and utility of geosynthetic-clay-liner cover systems for management of acid-generating mine wastes. Analyses of the microbial communities in tailings isolated beneath an engineered cover system provide a better understanding of the complex biogeochemical processes involved in the redox cycling of key elements, contribute to the remediation of mine wastes, and provide a valuable tool for assessment of the effectiveness of the remediation system.

Can Sulfate Be the First Dominant Aqueous Sulfur Species Formed in the Oxidation of Pyrite by Acidithiobacillus ferrooxidans?

According to the literature, pyrite (FeS2) oxidation has been previously determined to involve thiosulfate as the first aqueous intermediate sulfur product, which is further oxidized to sulfate. In the present study, pyrite oxidation by Acidithiobacillus ferrooxidans was studied using electrochemical and metabolic approaches in an effort to extend existing knowledge on the oxidation mechanism. Due to the small surface area, the reaction rate of a compact pyrite electrode in the form of polycrystalline pyrite aggregate in A. ferrooxidans suspension was very slow at a spontaneously formed high redox potential. The slow rate made it possible to investigate the oxidation process in detail over a term of 100 days. Using electrochemical parameters from polarization curves and levels of released iron, the number of exchanged electrons per pyrite molecule was estimated. The values close to 14 and 2 electrons were determined for the oxidation with and without bacteria, respectively. These results indicated that sulfate was the dominant first aqueous sulfur species formed in the presence of bacteria and elemental sulfur was predominantly formed without bacteria. The stoichiometric calculations are consistent with high iron-oxidizing activities of bacteria that continually keep the released iron in the ferric form, resulting in a high redox potential. The sulfur entity of pyrite was oxidized to sulfate by Fe3+ without intermediate thiosulfate under these conditions. Cell attachment on the corroded pyrite electrode surface was documented although pyrite surface corrosion by Fe3+ was evident without bacterial participation. Attached cells may be important in initiating the oxidation of the pyrite surface to release iron from the mineral. During the active phase of oxidation of a pyrite concentrate sample, the ATP levels in attached and planktonic bacteria were consistent with previously established ATP content of iron-oxidizing cells. No significant upregulation of three essential genes involved in energy metabolism of sulfur compounds was observed in the planktonic cells, which represented the dominant biomass in the pyrite culture. The study demonstrated the formation of sulfate as the first dissolved sulfur species with iron-oxidizing bacteria under high redox potential conditions. Minor aqueous sulfur intermediates may be formed but as a result of side reactions.

Indirect Redox Transformations of Iron, Copper, and Chromium Catalyzed by Extremely Acidophilic Bacteria.

Experiments were carried out to examine redox transformations of copper and chromium by acidophilic bacteria (Acidithiobacillus, Leptospirillum, and Acidiphilium), and also of iron (III) reduction by Acidithiobacillus spp. under aerobic conditions. Reduction of iron (III) was found with all five species of Acidithiobacillus tested, grown aerobically on elemental sulfur. Cultures maintained at pH 1.0 for protracted periods displayed increasing propensity for aerobic iron (III) reduction, which was observed with cell-free culture liquors as well as those containing bacteria. At. caldus grown on hydrogen also reduced iron (III) under aerobic conditions, confirming that the unknown metabolite(s) responsible for iron (III) reduction were not (exclusively) sulfur intermediates. Reduction of copper (II) by aerobic cultures of sulfur-grown Acidithiobacillus spp. showed similar trends to iron (III) reduction in being more pronounced as culture pH declined, and occurring in both the presence and absence of cells. Cultures of Acidithiobacillus grown anaerobically on hydrogen only reduced copper (II) when iron (III) (which was also reduced) was also included; identical results were found with Acidiphilium cryptum grown micro-aerobically on glucose. Harvested biomass of hydrogen-grown At. ferridurans oxidized iron (II) but not copper (I), and copper (I) was only oxidized by growing cultures of Acidithiobacillus spp. when iron (II) was also included. The data confirmed that oxidation and reduction of copper were both mediated by acidophilic bacteria indirectly, via iron (II) and iron (III). No oxidation of chromium (III) by acidophilic bacteria was observed even when, in the case of Leptospirillum spp., the redox potential of oxidized cultures exceeded +900 mV. Cultures of At. ferridurans and A. cryptum reduced chromium (VI), though only when iron (III) was also present, confirming an indirect mechanism and contradicting an earlier report of direct chromium reduction by A. cryptum. Measurements of redox potentials of iron, copper and chromium couples in acidic, sulfate-containing liquors showed that these differed from situations where metals are not complexed by inorganic ligands, and supported the current observations of indirect copper oxido-reduction and chromium reduction mediated by acidophilic bacteria. The implications of these results for both industrial applications of acidophiles and for exobiology are discussed.

Are there multiple mechanisms of anaerobic sulfur oxidation with ferric iron in Acidithiobacillus ferrooxidans?

To clarify the pathway of anaerobic sulfur oxidation coupled with dissimilatory ferric iron reduction in Acidithiobacillus ferrooxidans strain CCM 4253 cells, we monitored their energy metabolism gene transcript profiles. Several genes encoding electron transporters involved in aerobic iron and sulfur respiration were induced during anaerobic growth of ferrous iron-grown cells. Most sulfur metabolism genes were either expressed at the basal level or their expression declined. However, transcript levels of genes assumed to be responsible for processing of elemental sulfur and other sulfur intermediates were elevated at the beginning of the growth period. In contrast, genes with predicted functions in formation of hydrogen sulfide and sulfate were significantly repressed. The main proposed mechanism involves: outer membrane protein Cyc2 (assumed to function as a terminal ferric iron reductase); periplasmic electron shuttle rusticyanin; c4-type cytochrome CycA1; the inner membrane cytochrome bc1 complex I; and the quinone pool providing connection to the sulfur metabolism machinery, consisting of heterodisulfide reductase, thiosulfate:quinone oxidoreductase and tetrathionate hydrolase. However, an alternative mechanism seems to involve a high potential iron-sulfur protein Hip, c4-type cytochrome CycA2 and inner membrane cytochrome bc1 complex II. Our results conflict with findings regarding the type strain, indicating strain- or phenotype-dependent pathway variation.