Reflecting on gold geomicrobiology research: thoughts and considerations for future endeavors

Research in gold (Au) geomicrobiology has developed extensively over the last ten years, as more Au-bearing materials from around the world point towards a consistent story: That microbes interact with Au. In weathering environments, Au is mobile, taking the form of oxidized, soluble complexes or reduced, elemental Au nanoparticles. The transition of Au between aqueous and solid states is attributed to varying geochemical conditions, catalyzed in part by the biosphere. Hence, a global Au-biogeochemical-cycle was proposed. The primary focus of this mini-review is to reflect upon the biogeochemical processes that contribute to what we currently know about Au cycling. In general, the global Au-biogeochemical-cycle begins with the liberation of gold-silver particles from a primary host rock, by physical weathering. Through oxidative-complexation, inorganic and organic soluble-Au complexes are produced. However, in the presence of microbes or other reductants—e.g., clays and Fe-oxides—these Au complexes can be destabilized. The reduction of soluble Au ultimately leads to the bioprecipitation and biomineralization of Au, the product of which can aggregate into larger structures, thereby completing the Au cycle. Evidence of these processes have been “recorded” in the preservation of secondary Au structures that have been observed on Au particles from around the world. These structures—i.e., nanometer-size to micrometer-size Au dissolution and reprecipitation features—are “snap shots” of biogeochemical influences on Au, during its journey in Earth-surface environments. Therefore, microbes can have a profound effect on the occurrence of Au in natural environments, given the nutrients necessary for microbial metabolism are sustained and Au is in the system.Jeremiah Shuster and Frank Reit

Editorial for special issue "geomicrobiology and biogeochemistry of precious metals"

n/aFrank Reith and Jeremiah Shuste

A whole-cell biosensor for the detection of gold

Geochemical exploration for gold (Au) is becoming increasingly important to the mining industry. Current processes for Au analyses require sampling materials to be taken from often remote localities. Samples are then transported to a laboratory equipped with suitable analytical facilities, such as Inductively Coupled Plasma-Mass Spectrometry (ICP-MS) or Instrumental Neutron Activation Analysis (INAA). Determining the concentration of Au in samples may take several weeks, leading to long delays in exploration campaigns. Hence, a method for the on-site analysis of Au, such as a biosensor, will greatly benefit the exploration industry. The golTSB genes from Salmonella enterica serovar typhimurium are selectively induced by Au(I/III)-complexes. In the present study, the golTSB operon with a reporter gene, lacZ, was introduced into Escherichia coli. The induction of golTSB::lacZ with Au(I/III)-complexes was tested using a colorimetric β-galactosidase and an electrochemical assay. Measurements of the β-galactosidase activity for concentrations of both Au(I)- and Au(III)-complexes ranging from 0.1 to 5 µM (equivalent to 20 to 1000 ng g⁻¹ or parts-per-billion (ppb)) were accurately quantified. When testing the ability of the biosensor to detect Au(I/III)-complexes(aq) in the presence of other metal ions (Ag(I), Cu(II), Fe(III), Ni(II), Co(II), Zn, As(III), Pb(II), Sb(III) or Bi(III)), cross-reactivity was observed, i.e. the amount of Au measured was either under- or over-estimated. To assess if the biosensor would work with natural samples, soils with different physiochemical properties were spiked with Au-complexes. Subsequently, a selective extraction using 1 M thiosulfate was applied to extract the Au. The results showed that Au could be measured in these extracts with the same accuracy as ICP-MS (P<0.05). This demonstrates that by combining selective extraction with the biosensor system the concentration of Au can be accurately measured, down to a quantification limit of 20 ppb (0.1 µM) and a detection limit of 2 ppb (0.01 µM).Carla M. Zammit, Davide Quaranta, Shane Gibson, Anita J. Zaitouna, Christine Ta, Joël Brugger, Rebecca Y. Lai, Gregor Grass, Frank Reit

Cycling of biogenic elements drives biogeochemical gold cycling

Available online 13 DecemberMicroorganisms are key-drivers of carbon-, nitrogen-, sulfur- and metal cycling on Earth. Through their metabolic activities they directly and indirectly link element cycles. This leads to the cycling of elements through the Earth’s ecosystems from/to the atmosphere to/from the lithosphere. Gold (Au) is a rare, redox-active, noble transition metal, which is neither essential as a nutrient nor, reputedly, mobile in the environment. Therefore, observations published in recent decades, which have shown that gold is highly mobile and subject to biogeochemical cycling largely driven by microbiota, have surprised many. Questions concerning the fundamental biogeochemical processes mediating gold cycling, the organisms involved and the benefits they may gain have puzzled researchers. In this review we integrate the cycling of the major biogenic elements carbon, nitrogen and sulfur with that of gold. We identify key-processes that drive gold cycling and evaluate how different chemical Au(I/III)-species affect microbiota that form biofilms on gold-bearing minerals and placer gold particles. Additionally, we assess how the cycling of the gold-associated metal(loid)s silver, copper, iron, manganese, mercury and arsenic is linked to that of gold. Microbially produced compounds resulting from carbon, nitrogen, sulfur, iron and manganese cycling (e.g., organic acids, cyanides, (thio)sulfates, ammonium, iron sulfides/oxy-hydroxides and managanese oxides) can each play important roles for the mobilization of gold. Highly toxic, mobile Au(I/III)-complexes affect the phylogenetic and functional composition of microbial communities resident on gold particles. This leads to gold detoxification coupled to active and passive biomineralization, and ultimately the aggregation and (trans)formation of metallic gold particles. The complex interplay between gold, microbiota and physicochemical conditions modified by these organisms (e.g., redox or pH) has throughout the Earth’s history led to the aggregation of gold particles (grains to nuggets), led to the formation of the largest known gold deposit (i.e., Witwatersrand paleo-placer), and the largest gold reservoir in seawater. Today it opens up exciting biotechnological pathways for mineral exploration, processing and remediation.Santonu Kumar Sanyal, Jeremiah Shuster, Frank Reit

A genomic perspective of metal-resistant bacteria from gold particles: Possible survival mechanisms during gold biogeochemical cycling

Advance Access Publication Date: 4 June 2020.A bacterial consortium was enriched from gold particles that 'experienced' ca. 80 years of biotransformation within waste-rock piles (Australia). This bacterial consortium was exposed to 10 µM AuCl3 to obtain Au-tolerant bacteria. From these isolates, Serratia sp. and Stenotrophomonas sp. were the most Au-tolerant and reduced soluble Au as pure gold nanoparticles, indicating that passive mineralisation is a mechanism for mediating the toxic effect of soluble Au produced during particle dissolution. Genome-wide analysis demonstrated that these isolates also possessed various genes that could provide cellular defence enabling survival under heavy-metal stressed condition by mediating the toxicity of heavy metals through active efflux/reduction. Diverse metal-resistant genes or genes clusters (cop, cus, czc, znt, ars) were detected, which could confer resistance to soluble Au. Comparative genome analysis revealed that the majority of detected heavy-metal resistant genes were similar (i.e. orthologous) to those genes of Cupriavidus metallidurans CH34. The detection of heavy-metal resistance, nutrient cycling, and biofilm formation genes (pgaABCD, bsmA, hmpS) may have indirect yet important roles when dealing with soluble Au during particle dissolution. In conclusion, the physiological and genomic results suggest that bacteria living on gold particles would likely use various genes to ensure survival during Au biogeochemical cycling.Santonu Kumar Sanyal, Frank Reith and Jeremiah Shuste

Biogeochemical gold cycling selects metal-resistant bacteria that promote gold particle transformation

Bacteria catalyze the dissolution and re-precipitation of gold, thereby driving the biogeochemical cycle of gold. Dissolution of gold/silver and re-precipitation of gold transforms gold particles by increasing gold purity. While soluble gold complexes are highly cytotoxic, little is known about how gold cycling affects bacterial communities residing on gold particles. Micro-analysis of gold particles obtained from Western Australia revealed porous textures and aggregates of pure gold nanoparticles, attributable to gold dissolution and re-precipitation, respectively. By interpreting structure and chemistry of particles, the kinetics of gold biogeochemical cycling at the site was estimated to be 1.60 × 10−9 M year−1. Bacterial communities residing on particles were composed of Proteobacteria (42.5%), Bacteroidetes (20.1%), Acidobacteria (19.1%), Firmicutes (8.2%), Actinobacteria (3.7%), and Verrucomicrobia (3.6%). A bacterial enrichment culture obtained from particles contained a similar composition. Exposure of enrichments to increasing concentrations of soluble gold decreased community diversity and selected for metal-resistant bacteria. Lower gold concentrations, which corresponded well with the concentration from the kinetic rate, provided a selective pressure for the selection of metal-resistant organisms while retaining the overall diversity. In conclusion, biogeochemical gold cycling directly influences bacterial communities on gold particles, thereby contributing to a continuum of particle transformation.Santonu Kumar Sanyal, Jeremiah Shuster, Frank Reit

Proteomic responses to gold(III)-toxicity in the bacterium Cupriavidus metallidurans CH34

Accepted 11th October 2016The metal-resistant β-proteobacterium Cupriavidus metallidurans drives gold (Au) biomineralisation and the (trans)formation of Au nuggets largely via unknown biochemical processes, ultimately leading to the reductive precipitation of mobile, toxic Au(i/iii)-complexes. In this study proteomic responses of C. metallidurans CH34 to mobile, toxic Au(iii)-chloride are investigated. Cells were grown in the presence of 10 and 50 μM Au(iii)-chloride, 50 μM Cu(ii)-chloride and without additional metals. Differentially expressed proteins were detected by difference gel electrophoresis and identified by liquid chromatography coupled mass spectrometry. Proteins that were more abundant in the presence of Au(iii)-chloride are involved in a range of important cellular functions, e.g., metabolic activities, transcriptional regulation, efflux and metal transport. To identify Au-binding proteins, protein extracts were separated by native 2D gel electrophoresis and Au in protein spots was detected by laser absorption inductively coupled plasma mass spectrometry. A chaperon protein commonly understood to bind copper (Cu), CupC, was identified and shown to bind Au. This indicates that it forms part of a multi-metal detoxification system and suggests that similar/shared detoxification pathways for Au and Cu exist. Overall, this means that C. metallidurans CH34 is able to mollify the toxic effects of cytoplasmic Au(iii) by sequestering this Au-species. This effect may in the future be used to develop CupC-based biosensing capabilities for the in-field detection of Au in exploration samples.Carla M. Zammit, Florian Weiland, Joël Brugger, Benjamin Wade, Lyron Juan Winderbaum, Dietrich H. Nies, Gordon Southam, Peter Hoffmann and Frank Reit

From biomolecules to biogeochemistry: Exploring the interaction of an indigenous bacterium with gold

Specialised microbial communities colonise the surface of gold particles in soils/sediments, and catalyse gold dissolution and re-precipitation, thereby contributing to the environmental mobility and toxicity of this ‘inert’ precious metal. We assessed the proteomic and physiological response of Serratia proteamaculans, the first metabolically active bacterium enriched and isolated directly from natural gold particles, when exposed to toxic levels of soluble Au3+ (10 μM). The results were compared to a metal-free blank, and to cultures exposed to similarly toxic levels of soluble Cu2+ (0.1 mM); Cu was chosen for comparison because it is closely associated with Au in nature due to similar geochemical properties. A total of 273 proteins were detected from the cells that experienced the oxidative effects of soluble Au, of which 139 (51%) were upregulated with either sole expression (31%) or had synthesis levels greater than the Au-free control (20%). The majority (54%) of upregulated proteins were functionally different from up-regulated proteins in the bacteria-copper treatment. These proteins were related to broad functions involving metabolism and biogenesis, followed by cellular process and signalling, indicating significant specificity for Au. This proteomic study revealed that the bacterium upregulates the synthesis of various proteins related to oxidative stress response (e.g., Monothiol-Glutaredoxin, Thiol Peroxidase, etc.) and cellular damage repair, which leads to the formation of metallic gold nanoparticles less toxic than ionic gold. Therefore, indigenous bacteria may mediate the toxicity of Au through two different yet simultaneous processes: i) repairing cellular components by replenishing damaged proteins and ii) neutralising reactive oxygen species (ROS) by up-regulating the synthesis of antioxidants. By connecting the fields of molecular bacteriology and environmental biogeochemistry, this study is the first step towards the development of biotechnologies based on indigenous bacteria applied to gold bio-recovery and bioremediation of contaminated environments.Santonu K. Sanyal, Tara Pukala, Parul Mittal, Frank Reith, Joël Brugger, Barbara Etschmann, Jeremiah Shuste

Biogeochemical cycling of silver in acidic, weathering environments

Under acidic, weathering conditions, silver (Ag) is considered to be highly mobile and can be dispersed within near-surface environments. In this study, a range of regolith materials were sampled from three abandoned open pit mines located in the Iberian Pyrite Belt, Spain. Samples were analyzed for Ag mineralogy, content, and distribution using micro-analytical techniques and high-resolution electron microscopy. While Ag concentrations were variable within these materials, elevated Ag concentrations occurred in gossans. The detection of Ag within younger regolith materials, i.e., terrace iron formations and mine soils, indicated that Ag cycling was a continuous process. Microbial microfossils were observed within crevices of gossan and their presence highlights the preservation of mineralized cells and the potential for biogeochemical processes contributing to metal mobility in the rock record. An acidophilic, iron-oxidizing microbial consortium was enriched from terrace iron formations. When the microbial consortium was exposed to dissolved Ag, more than 90% of Ag precipitated out of solution as argentojarosite. In terms of biogeochemical Ag cycling, this demonstrates that Ag re-precipitation processes may occur rapidly in comparison to Ag dissolution processes. The kinetics of Ag mobility was estimated for each type of regolith material. Gossans represented 0.6–146.7 years of biogeochemical Ag cycling while terrace iron formation and mine soils represented 1.9–42.7 years and 0.7–1.6 years of Ag biogeochemical cycling, respectively. Biogeochemical processes were interpreted from the chemical and structural characterization of regolith material and demonstrated that Ag can be highly dispersed throughout an acidic, weathering environment.Jeremiah Shuster, Frank Reith, Matthew R. M. Izawa, Roberta L. Flemming, Neil R. Banerjee and Gordon Southa