Bioleaching Operations Research Articles

Complete genome sequence for the thermoacidophilic archaeon Sulfuracidifex (f. Sulfolobus) metallicus DSM 6482.

Reported here is the complete genome sequence (2,191,724 bp) for the thermoacidophilic archaeon Sulfuracidifex (f. Sulfolobus) metallicus DSM 6482 (Topt 65°C, pHopt 2.0). This obligately chemolithoautotrophic microorganism is a prolific metal and sulfur oxidizer and has application in metal bioleaching operations. A multi-assembly reconciliation approach enabled closure of the genome.

Augmentation in bioleaching potential of indigenous Bacillus sp. ISO1 for metals recovery from waste computer-printed circuit boards.

The bio-cyanidation process of various cyanogenic microorganisms is found to be a sustainable and effective method for metals recovery from primary and secondary sources. This process has surpassed the limitations of the chemical cyanide treatment process; thus, prioritized as a promising approach for e-waste "urban mining" strategies. The main focus of the study was to enhance the bioleaching capacity of indigenous Bacillus sp. ISO1 and to implement optimized parameters in large-scale bioleaching operations. The assessment of various amino acids unveiled that like other cyanogenic microorganisms Bacillus sp. ISO1 also preferred glycine as a prime precursor for cyanide synthesis, as maximum metal solubilization was achieved with glycine amino acid. Other amino acids influenced the bacterial growth but not significantly affected the biocyanidation process. The evaluation and optimization of methionine as a lixiviant stimulator demonstrated that the addition of 1 mg/L methionine effectively enhance the production of glycine-utilizing cyanide lixiviant, that led to a significant solubilization of Cu (86%), Au (75%), and Ag (63%) metals. Furthermore, the kinetics of metal solubilization and operating conditions were explored at increased volume (i.e., 3 L working volume) of bioleaching medium to assess the industrial scale potential of this potent bacterial strain with optimized parameters such as temperature, pH, pulp density, and inoculum size. The significant recovery of Cu (˃ 60%) and other metals at this substantial volume suggested the implementation of a bioleaching process with this potent bacterial strain at industrial scale operations.

Simulating compatible solute biosynthesis using a metabolic flux model of the biomining acidophile, Acidithiobacillus ferrooxidans ATCC 23270

Halotolerant, acidophilic, bioleaching microorganisms are crucial to biomining operations that utilize saline water. Compatible solutes play an important role in the adaptation of these microorganisms to saline environments. Acidithiobacillus ferrooxidans ATCC 23270, an iron- and sulfur-oxidizing acidophilic bacterium, synthesizes trehalose as its native compatible solute but is still sensitive to salinity. Recently, halotolerant bioleaching bacteria were found to use ectoine as their key compatible solute. Previously, bioleaching bacteria were recalcitrant to genetic manipulation; however, recent advancements in genetic tools and techniques allow successful genetic modification of A. ferrooxidans ATCC 23270. Therefore, this study aimed to test, in silico, the effect of native and synthetic compatible solute biosynthesis by A. ferrooxidans ATCC 23270 on its growth and metabolism. Metabolic network flux modelling was used to provide a computational framework for the prediction of metabolic fluxes during production of native and synthetic compatible solutes by A. ferrooxidans ATCC 23270, in silico. Complete pathways for trehalose biosynthesis by the bacterium are proposed and captured in the updated metabolic model including a newly discovered UDP-dependent trehalose synthesis pathway. Finally, the effect of nitrogen sources on compatible solute production was simulated and showed that using nitrogen gas as the sole nitrogen source enables the ectoine-producing ‘engineered’ microbe to oxidize up to 20% more ferrous iron in comparison to the native microbe that only produces trehalose. Therefore, the predictive outcomes of the model have the potential to guide the design and optimization of a halotolerant strain of A. ferrooxidans ATCC 23270 for saline bioleaching operations.

A Critical Review on the Recovery of Base and Critical Elements from Electronic Waste-Contaminated Streams Using Microbial Biotechnology.

Pollution by end-of-life electronics is a rapid ever-increasing threat and is a universal concern with production of million metric tons of these wastes per annum. Electronic wastes (E-waste) are rejected electric or electronic equipment which have no other applications. The aggrandized unproper land filling of E-waste may generate hazardous effects on living organisms and ecosystem. At present, millions of tons of E-waste await the advancement of more efficient and worthwhile recycling techniques. Recovery of base and critical elements from electronic scraps will not only reduce the mining of these elements from natural resources but also reduces the contamination caused by the hazardous chemicals (mostly organic micropollutants) released from these wastes when unproperly disposed of. Bioleaching is reported to be the most eco-friendly process for metal recycling from spent electronic goods. A detailed investigation of microbial biodiversity and a molecular understanding of the metabolic pathways of bioleaching microorganisms will play a vital function in extraction of valuable minerals from the end-of-life scraps. Bioleaching technique as an economic and green technology costs around 7 USD per kg for effective reusing of E-waste as compared to other physical and chemical techniques. This review provides a summary of worldwide scenario of electronic pollutants; generation, composition and hazardous components of electronic waste; recycling of valuable elements through bioleaching; mechanism of bioleaching; microorganisms involved in base and critical element recovery from E-waste; commercial bioleaching operations; and upcoming aspects of this eco-friendly technique.

Thermoacidophilic Bioleaching of Industrial Metallic Steel Waste Product.

The continuous deposition of hazardous metalliferous wastes derived from industrial steelmaking processes will lead to space shortages while valuable raw metals are being depleted. Currently, these landfilled waste products pose a rich resource for microbial thermoacidophilic bioleaching processes. Six thermoacidophilic archaea (Sulfolobus metallicus, Sulfolobus acidocaldarius, Metallosphaera hakonensis, Metallosphaera sedula, Acidianus brierleyi, and Acidianus manzaensis) were cultivated on metal waste product derived from a steelmaking process to assess microbial proliferation and bioleaching potential. While all six strains were capable of growth and bioleaching of different elements, A. manzaensis outperformed other strains and its bioleaching potential was further studied in detail. The ability of A. manzaensis cells to break down and solubilize the mineral matrix of the metal waste product was observed via scanning and transmission electron microscopy. Refinement of bioleaching operation parameters shows that changes in pH influence the solubilization of certain elements, which might be considered for element-specific solubilization processes. Slight temperature shifts did not influence the release of metals from the metal waste product, but an increase in dust load in the bioreactors leads to increased element solubilization. The formation of gypsum crystals in course of A. manzaensis cultivation on dust was observed and clarified using single-crystal X-ray diffraction analysis. The results obtained from this study highlight the importance of thermoacidophilic archaea for future small-scale as well as large-scale bioleaching operations and metal recycling processes in regard to circular economies and waste management. A thorough understanding of the bioleaching performance of thermoacidophilic archaea facilitates further environmental biotechnological advancements.

Unraveling the Central Role of Sulfur-Oxidizing Acidiphilium multivorum LMS in Industrial Bioprocessing of Gold-Bearing Sulfide Concentrates.

Acidiphilium multivorum LMS is an acidophile isolated from industrial bioreactors during the processing of the gold-bearing pyrite-arsenopyrite concentrate at 38–42 °C. Most strains of this species are obligate organoheterotrophs that do not use ferrous iron or reduced sulfur compounds as energy sources. However, the LMS strain was identified as one of the predominant sulfur oxidizers in acidophilic microbial consortia. In addition to efficient growth under strictly heterotrophic conditions, the LMS strain proved to be an active sulfur oxidizer both in the presence or absence of organic compounds. Interestingly, Ac. multivorum LMS was able to succeed more common sulfur oxidizers in microbial populations, which indicated a previously underestimated role of this bacterium in industrial bioleaching operations. In this study, the first draft genome of the sulfur-oxidizing Ac. multivorum was sequenced and annotated. Based on the functional genome characterization, sulfur metabolism pathways were reconstructed. The LMS strain possessed a complicated multi-enzyme system to oxidize elemental sulfur, thiosulfate, sulfide, and sulfite to sulfate as the final product. Altogether, the phenotypic description and genome analysis unraveled a crucial role of Ac. multivorum in some biomining processes and revealed unique strain-specific characteristics, including the ars genes conferring arsenic resistance, which are similar to those of phylogenetically distinct microorganisms.

Deciphering the Role of Multiple Thioredoxin Fold Proteins of Leptospirillum sp. in Oxidative Stress Tolerance.

Thioredoxin fold proteins (TFPs) form a family of diverse proteins involved in thiol/disulfide exchange in cells from all domains of life. Leptospirillum spp. are bioleaching bacteria naturally exposed to extreme conditions like acidic pH and high concentrations of metals that can contribute to the generation of reactive oxygen species (ROS) and consequently the induction of thiol oxidative damage. Bioinformatic studies have predicted 13 genes that encode for TFP proteins in Leptospirillum spp. We analyzed the participation of individual tfp genes from Leptospirillum sp. CF-1 in the response to oxidative conditions. Genomic context analysis predicted the involvement of these genes in the general thiol-reducing system, cofactor biosynthesis, carbon fixation, cytochrome c biogenesis, signal transduction, and pilus and fimbria assembly. All tfp genes identified were transcriptionally active, although they responded differentially to ferric sulfate and diamide stress. Some of these genes confer oxidative protection to a thioredoxin-deficient Escherichia coli strain by restoring the wild-type phenotype under oxidative stress conditions. These findings contribute to our understanding of the diversity and complexity of thiol/disulfide systems, and of adaptations that emerge in acidophilic microorganisms that allow them to thrive in highly oxidative environments. These findings also give new insights into the physiology of these microorganisms during industrial bioleaching operations.

Microbial assisted galvanic leaching of chalcopyrite concentrate in continuously stirred bioreactors

The objective of this study is to combine two proven technologies, namely galvanic and microbial assisted leaching of chalcopyrite in a novel way. Using a commercial chalcopyrite-rich flotation concentrate and pure natural pyrite samples from an Iranian copper mining complex as well as mixed cultures of moderately thermophilic microorganisms, continuous electrochemical bioleaching tests were performed in stirred bioreactors at 50 °C as a first step towards a commercial application of microbially assisted galvanic leaching of chalcopyrite.From the results of this study, the importance of oxidation reduction potential (ORP) on the catalytic interaction between chalcopyrite and pyrite could be pointed out as a main parameter for successful bioprocessing of chalcopyrite concentrates. Optimization of the particle size of feed (D80) and adjusting the ORP in the range between 400 and 500 mV are important criteria, governing the electrochemical bioleaching rate of chalcopyrite concentrates. It is believed that the main reason for increased copper recovery could be the control of chalcopyrite passivation resulting from improved galvanic interaction between chalcopyrite and pyrite in the selected ORP range and the bacterial assisted oxidation of Fe2+ to Fe3+. Under optimum conditions of this study (T = 50 ± 2 °C, pH = 1.1 ± 0.2, ORP = 400–500 mV, solid load = 10%, stirring rate = 300 rpm) the copper extraction from chalcopyrite flotation concentrate during continuous electrochemical bioleaching operations in stirred bioreactors was >95%, which should be high enough to justify the process economically.

A delve into the exploration of potential bacterial extremophiles used for metal recovery

A multitude of microbes are involved in the solubilisation of minerals and metals as this approach offers numerous advantages over traditional methods. This strategy is preferred as it is eco-friendly and economical, thus overcoming the drawbacks of the traditional approach of pyrometallurgy. Many different types of bacteria are employed in the process of Bioleaching, which are collectively grouped under chemolithotrophs, as they derive their energy from inorganic compounds. Bioleaching is the mobilization of metal cations from insoluble ores by microorganisms. All chemolithotropic bacteria are extremophiles since they have the ability to survive in extreme conditions. They carry out the process of Bioleaching through three mechanisms: Indirect, contact/ direct and cooperative bioleaching. This review gives a sneak peek into the different strains of chemolithotrophs which are used in bioleaching, and some recent work in the field. It also gives an insight into the general process and mechanism of Bioleaching, the study of which will pave way for developing better and efficient industrial bioleaching operations.

From Knowledge to Best Practices in Bioleaching

The huge industrial data recorded by several years in copper bioleaching operations represents an opportunity for the technology improvement. A systematic approach is being developed to get insights from the field data from an industrial process and to deliver the obtained knowledge with the aim to serve as the foundation for optimal industrial decision making even in presence of inherent process variations. The development of this Decision Support System (DSS) considers a Q-PCR array, a database for data logging and storage, the application of suitable statistical and computational tools for data analyses and knowledge acquiring and finally the creation of a system of knowledge translation to transform it into action (operational suggestions). The user can accurately retrieve data and design similar matches to the historic operation to get e.g. the expected metallurgical performance of a strip based on its mineralogical parameters. In addition, the user can get computed information and recommendations that should be analyzed. We will discuss the process followed to construct the base of knowledge of the DSS.

Transcription Dynamics of CBB-Pathway Genes in <i>Acidithiobacillus thiooxidans</i> Growing under Different CO<sub>2</sub> Levels

Acidithiobacillus thiooxidans is one of the most stable components in the consortium of microorganisms inhabiting the copper bioleaching heap of Minera Escondida Limitada. CalvinBenson-Bassham (CBB) is the CO2 fixation pathway most used by aerobic bacteria, and is the exclusive pathway used by A. thiooxidans. In this work, the population dynamics and the transcriptomic dynamics of five key genes involved in the A. thiooxidans CBB pathway were studied in bioleaching column and pure culture tests at different concentrations of CO2 availability. Association between CO2 level, population dynamics and relative expression of CBB-genes was discovered. Differences between CBB pathways from closely related species were reported. The transcription profile modelling could provide useful knowledge for improving industrial bioleaching operations.

Bioelectrochemical Leaching of Copper Sulfide Minerals

The objective of this study is to improve the understanding of copper sulfides dissolution and to use this knowledge for optimization of process parameters for commercial application of electrochemical bioleaching of chalcopyrite concentrates in stirred bioreactors. From the results of this study, the importance of the oxidation reduction potential (ORP) on the catalytic interaction between chalcopyrite and pyrite can be pointed out as the main parameters for successful bioprocessing of chalcopyrite concentrates. Under these conditions, the optimization of the average particle size of feed (D80) and adjusting the ORP in the range between 400-450 mV are important criteria for increasing the electrochemical bioleaching rate of chalcopyrite concentrates. It seems that the main reason for the increased copper recovery could be the control and prevention of chalcopyrite passivation resulting from improved galvanic interaction between copper sulfide minerals, here especially chalcopyrite and pyrite in the selected ORP range and the right particle size distribution of feed. At optimum conditions, the copper extraction from chalcopyrite flotation concentrate during 7 days of continuous electrochemical bioleaching operations in stirred tanks was about 95%, which should be high enough to justify the process economically.

The Microbial Ecology of Moderately Thermophilic Mineral Leaching Reactors: The Effect of Solids Loading and Organic Carbon Supplementation on Reactor Performance

Tank bioleaching of refractory sulphidic gold ores is well established, with potential to expand application to base metal concentrates. With increasing commercial tank bioleaching operations, understanding their microbial consortia is essential for process robustness. Recently, it has been shown that the consortia implicated in tank mineral bioleaching of gold-containing pyritic ores are dynamic, responding to the leaching environment. Factors driving the microbial dynamics of these consortia are under investigation, with emphasis also placed on determining the metabolic role of the key players in these consortia. Here, the combined influence of solids loading and organic carbon availability on microbial community dynamics and performance has been studied in agitated, aerated slurry bioreactors at 45°C. The control (autotrophic) and experimental (organic carbon added) reactors performed comparably at low solids loadings (4%, 7% and 10%). At 20% solids loadings, higher ferric iron concentrations (31 g/L vs 25g/L) and sulphide oxidation (66% vs 45%) were observed in the experimental reactor over the control. Under operating conditions used, a shift from bacterially-dominated cultures with L.ferriphilum as major species towards increasing archaeal abundance was observed. Archaeal abundance was higher in the organic carbon supplemented reactor at all solids loadings. The increased microbial diversity with organic supplementation appears to contribute to increased community robustness and associated leaching with increasing stress.

Effect of physico-chemical and operating conditions on the growth and activity of Acidithiobacillus ferrooxidans in a simulated heap bioleaching environment

Recent understanding of microbial retention within heap bioleaching systems has highlighted the importance of quantifying microbial growth and activity in both the bulk flowing solution and in the ore-associated phases. Typically, industrial heap bioleaching operations report variations in process conditions such as inoculum preparation and concentration and elevated copper concentrations in the recycled irrigation solution. In this paper, a mini-column reactor system containing pre-constructed and agglomerated, low-grade ore samples representing grab samples from a larger heap, were used to investigate the effect of a selection of physico-chemical and operating conditions on microbial growth, colonisation and substrate utilisation kinetics, considering both the planktonic and sessile populations of Acidithiobacillus ferrooxidans. The factors studied included inoculum size, inoculum cultivation conditions, availability of ferrous iron in the bulk flowing solution and copper concentration in the bulk flowing solution. The microbial population in the interstitial phase, i.e. associated with, but not bound to, the ore, remained the most abundant within the heap under all physico-chemical conditions considered. A comparison of the tests with different inoculum sizes found that a smaller inoculum size resulted in an increased delay in microbial growth and ferrous iron oxidation, but similar apparent maximum specific growth rates and iron oxidation rates. In contrast to the microbial culture grown on pyrite, a delay in microbial activity was observed for the culture grown on ferrous iron. However, greater microbial cell densities were reached, in the interstitial and attached phases compared with the pyrite-grown culture. The introduction of 6gL−1 cupric ions into the feed solution containing 0.2gL−1 ferric iron resulted in decreased microbial growth rate in the interstitial phase but not in the attached phase. Where the pyrite culture was pre-exposed to cupric ion, the microbial growth rate in the interstitial and attached phases was significantly enhanced. Nevertheless, the presence of cupric ion in the irrigation solution resulted in a decrease in microbial ferrous iron oxidation rate, irrespective of pre-culture conditioning. This study emphasises the important role played by the stagnant interstitial phase during the colonisation of a low-grade heap, particularly under adverse conditions for microbial growth and activity. It also highlights the role of inoculum culture conditions on the potential trade-off between increased heap colonisation and increased lag periods in microbial activity during heap start-up.

Analysis of Gene Expression in Response to Copper Stress in <i>Acidithiobacillus ferrooxidans</i> Strain D2, Isolated from a Copper Bioleaching Operation

Acidithiobacillus ferrooxidans strain D2 was isolated from a copper bioleaching operation in Atacama Desert, Chile. Copper is widely used as cofactor in proteins but high concentrations of copper are toxic. Cells require certain mechanisms to maintain the copper homeostasis and avoid toxic effects of high intracellular concentration. The molecular response of A. ferrooxidans strain D2 grown in the presence/absence of copper was examined using a A. ferrooxidans whole-genome DNA microarrays. Roughly 23% of 3,147 genes represented on the microarray were differentially expressed; about 9% of them were upregulated in the presence of copper. Among the upregulated genes, those encoding for the copper efflux protein (CusA) and for the copper-translocating P-type ATPase (CopA) were upregulated. The expression of genes encoding proteins related to iron transport was repressed. Similarly, genes related with assimilative metabolism of sulfur (L-cysteine biosynthesis) cysB, cysJ, cysI, CysD-2 and cysN were upregulated. Our results show that when A. ferrooxidans strain D2 was challenged with high copper concentrations, genes related to copper stress response were upregulated as well as others that have not been reported to be related to that mechanism. In addition, some genes related to other metabolic pathways were repressed, probably because of the energy cost of the stress response.

Effect of Contact with Organic Extractant LIX 84IC during Solvent Extraction Process on the Re-Establishment of Growth of Bioleaching Microorganisms

Solvent extraction (SX) used for the recovery of copper is a primary low-cost hydrometallurgical process applied extensively in cases like commercial bioleaching operations. Bioleaching solutions during SX treatment are exposed to important concentrations of organic extractants, a step that has a detrimental impact on microbial activity. The following work studies and determines the effect of this exposure on the re-establishment of growth and activity of different bioleaching microorganisms after contact and phase separation. Different bioleaching iron and sulfur -oxidizing microorganisms were tested. After contact with an industrially utilized organic extractant (LIX 84IC as extractant and Oxform SX12 as solvent), cultures were grown and their substrate utilization monitored.Acidithiobacillus ferrooxidansshowed only a slight increase in doubling time without significant latency period. On the contrary, for bothSulfobacillusspecies studied,S. thermosulfidooxidansandS. thermotolerans, were strongly impacted showing important lag phases before growth could take place. Additionally, the SX reagent showed disruption of cell walls and protein liberation during contact with all the different strains studied.

Thermophilic bioleaching of heavy metals from waste sludge using response surface methodology

The bioleaching process is considered to be more efficient and environmentally friendly than conventional technologies for removal of heavy metals from waste sludge. The objective of this study was to develop an optimal thermophilic bioleaching process for the treatment of waste sludge containing high concentrations of heavy metals. In this study, two operating parameters, sludge solid content and sulfur (substrate) concentration, were studied based on a central composite design (CCD) for their metal solubilization and solid degradation performances. The optimal bioleaching operation conditions were then determined using the response surface methodology (RSM). The results indicated that an increase in sludge solid content range from 0.5% to 5.0% resulted in a decrease in the pH reduction rate due to the increase in buffering capacity. The rate of acidification corresponded to sulfur concentration until sulfur itself became inhibitory. At sulfur concentration beyond approximately 2.75%, the lower acidification rate was caused by a lower bacteria growth rate. Similar trends were also observed in the variations of ORP and sulfate concentrations during this thermophilic bioleaching process. At the optimum conditions of a sludge solid content of 0.5% and sulfur concentration of 2.5%, the thermophilic bioleaching process achieved the maximum solubilization of 97%, 99%, 99% and 78% for Cu, Zn, Ni and Pb, respectively. At the same time, the maximum SS and VSS destruction efficiency were 69% and 63%, respectively.

Anaerobic Sulfur Metabolism Coupled to Dissimilatory Iron Reduction in the Extremophile Acidithiobacillus ferrooxidans

Gene transcription (microarrays) and protein levels (proteomics) were compared in cultures of the acidophilic chemolithotroph Acidithiobacillus ferrooxidans grown on elemental sulfur as the electron donor under aerobic and anaerobic conditions, using either molecular oxygen or ferric iron as the electron acceptor, respectively. No evidence supporting the role of either tetrathionate hydrolase or arsenic reductase in mediating the transfer of electrons to ferric iron (as suggested by previous studies) was obtained. In addition, no novel ferric iron reductase was identified. However, data suggested that sulfur was disproportionated under anaerobic conditions, forming hydrogen sulfide via sulfur reductase and sulfate via heterodisulfide reductase and ATP sulfurylase. Supporting physiological evidence for H2S production came from the observation that soluble Cu(2+) included in anaerobically incubated cultures was precipitated (seemingly as CuS). Since H(2)S reduces ferric iron to ferrous in acidic medium, its production under anaerobic conditions indicates that anaerobic iron reduction is mediated, at least in part, by an indirect mechanism. Evidence was obtained for an alternative model implicating the transfer of electrons from S(0) to Fe(3+) via a respiratory chain that includes a bc(1) complex and a cytochrome c. Central carbon pathways were upregulated under aerobic conditions, correlating with higher growth rates, while many Calvin-Benson-Bassham cycle components were upregulated during anaerobic growth, probably as a result of more limited access to carbon dioxide. These results are important for understanding the role of A. ferrooxidans in environmental biogeochemical metal cycling and in industrial bioleaching operations.

Dynamic microbial populations in heap leaching of zinc sulphide ore

Polymerase chain reaction (PCR) technology was used to monitor the microbial population within two 6-m-high sulphide ore heaps where zinc was bioleached. Genomic DNA was isolated from microorganisms attached to the ore particles. The microbial population varied in diversity at different heap heights and at different stages of the leaching cycle. Nine bioleaching microorganisms were identified, including iron oxidizers, sulphur oxidizers, whether mesophiles, or moderate thermophiles. No bioleaching extreme thermophiles were detected. Moderate thermophiles were present at average heap temperatures of 40 °C or higher. No moderate thermophiles were present in the top 2 m of the heaps. Mesophiles were present throughout the lifetime of the heaps and at all heap depths. Cultures of Acidithiobacillus caldus and Sulfobacillus thermosulfidooxidans originally introduced to inoculate the heaps did not proliferate but were instead displaced by other species; no benefits were obtained from heap inoculation. The most predominant species were Acidithiobacillus thiooxidans and Acidithiobacillus albertensis. Leptospirillum sp. was not predominant in the heaps, in contrast to the dominant role this organism plays in tank bioleaching operations.

Genomic insights into microbial iron oxidation and iron uptake strategies in extremely acidic environments

This minireview presents recent advances in our understanding of iron oxidation and homeostasis in acidophilic Bacteria and Archaea. These processes influence the flux of metals and nutrients in pristine and man-made acidic environments such as acid mine drainage and industrial bioleaching operations. Acidophiles are also being studied to understand life in extreme conditions and their role in the generation of biomarkers used in the search for evidence of existing or past extra-terrestrial life. Iron oxidation in acidophiles is best understood in the model organism Acidithiobacillus ferrooxidans. However, recent functional genomic analysis of acidophiles is leading to a deeper appreciation of the diversity of acidophilic iron-oxidizing pathways. Although it is too early to paint a detailed picture of the role played by lateral gene transfer in the evolution of iron oxidation, emerging evidence tends to support the view that iron oxidation arose independently more than once in evolution. Acidic environments are generally rich in soluble iron and extreme acidophiles (e.g. the Leptospirillum genus) have considerably fewer iron uptake systems compared with neutrophiles. However, some acidophiles have been shown to grow as high as pH 6 and, in the case of the Acidithiobacillus genus, to have multiple iron uptake systems. This could be an adaption allowing them to respond to different iron concentrations via the use of a multiplicity of different siderophores. Both Leptospirillum spp. and Acidithiobacillus spp. are predicted to synthesize the acid stable citrate siderophore for Fe(III) uptake. In addition, both groups have predicted receptors for siderophores produced by other microorganisms, suggesting that competition for iron occurs influencing the ecophysiology of acidic environments. Little is known about the genetic regulation of iron oxidation and iron uptake in acidophiles, especially how the use of iron as an energy source is balanced with its need to take up iron for metabolism. It is anticipated that integrated and complex regulatory networks sensing different environmental signals, such as the energy source and/or the redox state of the cell as well as the oxygen availability, are involved.