Solid State Phenomena Vol. 262

Abstract: Eukaryotic diversity in Río Tinto turns out to be unexpectedly high when compared to the prokaryotic one. Unlike the prokaryotic community, little is known about the role of the most abundant eukaryotes, mainly algae and fungi, in this ecosystem. Previous studies using acidophilic fungi isolated from the Tinto basin have shown their ability to specifically sequester toxic metals. We have also been able to demonstrate their direct implication in the geochemical cycles through biomineralization processes. Although the role that fungi may play in the Tinto basin is still poorly understood, is becoming clear that they participate very actively in the geological conformation of the environment, generating minerals of possible economical interest.

Abstract: Acidiphilium sp. DSM 27270 (Yenapatur) forms part of a microbial consortium isolated from copper mines in Chile, that is currently used in state of the art industrial-scale biotechnology. Its taxonomic assignment is still tentative and is metal resistance and homeostatic responses poorly characterized. Here we report the genomic taxonomy evaluation of the Yenapatur strain and the preliminary characterization of its metal resistance and homeostatic responses. The genome of Yenapatur was re-assembled, annotated and compared to other sequenced strains of the genus. Genomic signatures were derived to better define the taxonomy of the strain. The minimum inhibitory concentrations of diverse cations and anions (Cu (II), Fe (II), Mg (II), Mn (II), Zn (II), Al (III), As (III), Fe (III), chloride and nitrate) were determined. Known resistance determinants were profiled in the genome of Yenapatur and publically available sequenced Acidiphilium strains. Results are presented and discussed under the light of the operational conditions in which Yenapatur thrives.

Abstract: Acidophilic leaching microorganisms have been reported to be in general intolerant to high salinity, namely high concentrations of chloride. At present this restriction hampers the use of sea water for bioleaching technology. Enrichment cultures obtained in this study from a former ore deposit near the Spanish coast oxidize ferrous iron in the presence of up to 50 gL-1 NaCl at pH 2.5 and 37°C. The presence of at least 5 gL-1 NaCl was shown to be an obligate requirement for iron oxidation. The major microbial groups comprise Alicyclobacillus and Arthrobacter. The findings may be of biotechnological relevance.

Abstract: The identification of genes involved in cold adaptations of psychrotolerant bacteria Acidithiobacillus ferrivorans is important for biomining processes that take place at low temperatures like Andean mining installations in Peru. We have performed relative quantification RT-qPCR on candidate genes to have a role in adaptations at low temperature (5°C). The candidate genes analyzed were six: Two trehalose synthesis pathway genes, trehalose synthase (treS) and malto-oligosiltrehalose trehalohydrolase (treZ) showing no overexpression at 5°C. Two diguanylate cyclases genes related to exopolymer synthesis and biofilm formation (designated as dgc-I and dgc-II in this paper) were overexpressed at 21°C. The rusA and rusB genes involved in iron oxidation showed no significant change for rusA and no expression for rusB gene in any of both conditions. Genes rpoC, gyrB and alaS were validated as reference genes. These results show congruency with trancriptomics studies about gene expression of A. ferrivorans. Furthermore, the trehalose synthesis genes show no overexpression at low temperatures suggesting that other cold adaptation mechanisms are involved.

Abstract: After 24h of exposure to acidic media, pyrite generates reactive oxygen species (ROS). Freshly-crushed pyrite with grain sizes between 50-100 μm at a 5 % (w/v), pulp density generated 0.17 ± 0.01 mM H₂O₂, while 10% pyrite generated 0.29 ± 0.01 mM and 30 % pyrite generated approximately 0.83 ± 0.06 mM. These levels of H₂O₂ inhibit iron oxidation in iron-grown cells of Acidithiobacillus ferrooxidans^T but not in pyrite-grown cells. ROS originating from pyrite, which was incubated for 24 h in acidic medium, prohibited pyrite dissolution by iron-grown cells, while pyrite-grown cells were adapted to these concentrations of ROS. Periodical addition of 100 μM H₂O₂ to pyrite cultures inoculated with pyrite-grown cells did not lower iron dissolution as it was observed with iron-grown cells. By high throughput proteomics analysis, an increased expression of proteins related to oxidative stress management, iron-and sulfur oxidation systems, carbon fixation and biofilm formation was observed in biofilm cells grown on pyrite compared to iron-grown cells.

Abstract: 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.

Abstract: Ferric ion precipitation in a typical bioleach operation is known to be inevitable. However, its management has been emphasized as critical to efficiently maximize the potential that bioleach technique offers, especially in heap bioleach systems. Although there are some studies on ferric ion precipitation, limited studies exist in the context of a cold condition which often prevails in some operations. This study investigated ferrous ion biooxidation and ferric ion precipitation under cold temperature conditions. The results showed that maximum ferrous ion oxidation rates of 1.05, 1.25, 1.51 and 1.60 mmol/L/h at 6, 7, 8 and 10°C respectively were achieved in packed column reactor. The results further showed a simultaneous ferric ion precipitation during the oxidation process. The kinetics of ferric ion precipitation follows a first order kinetics with the rate constants of 0.0066, 0.0074, 0.0080 and 0.011 h^-1_. Activation energies of 68 and 77 kJ/mol were obtained for both ferrous ion biooxidation and ferric ion precipitation respectively. Although slow kinetics are expected for both processes, the kinetics information may be useful for design management consideration at least to predict when bioleach liquor would likely run out of dissolved ferric ion.

Abstract: Chloride leaching is considered a promising alternative method to recover copper from chalcopyrite and other primary copper sulfides, because it favors the leaching kinetics and avoids passivation of minerals. Nevertheless, chloride ions are highly toxic for iron-oxidizing microorganisms that participate in the bioleaching process. A comparative genomic analysis was carried out based on the complete genomes of bacteria belonging to Nitrospirae, Firmicutes, Actinobacteria and Proteobacteria phyla, and of archaea belonging to Euryarchaeota and Crenarchaeota was carried out to identify molecular determinants involved in chloride tolerance of acidophilic iron-oxidizing microorganisms. The results obtained showed that representative Nitrospirae and Firmicutes harbor genes for the biosynthesis and uptake of compatible solutes such as ectoine, trehalose and potassium, which have been shown to have a role in salt tolerance. Microorganisms belonging to other phyla harbor genes for potassium transporters, but no genes for compatible solutes were detected. In agreement with the bioinformatic results, minimum inhibitory concentration (MIC) determinations and growth kinetics experiments showed that Leptospirillum ferriphilum (Nitrospirae) was more tolerant to NaCl than Acidithiobacillus ferrooxidans (Proteobacteria). Furthermore, it was observed that the addition of 0.5 mM ectoine to the L. ferriphilum culture stimulated growth in the presence of 100 mM NaCl. On the contrary, ectoine had no effect on the growth of At. ferrooxidans. These results suggest that ectoine and likely trehalose could play a key role in chloride tolerance in L. ferriphilum. conferring adaptative advantages compared to A. ferrooxidans and possibly other iron-oxidizing microorganisms.

Abstract: Most bacteria and all archaea possess as outermost cell envelope so called surface-layers (S-layers). These layers were formed by self-assembling proteins having a number of habitat depending interesting intrinsic properties. As example, S-layers from bacterial isolates recovered from heavy metal contaminated environments have outstanding metal binding properties and are highly stable. Thus they selectively bind several metals with different affinity. For using S-layer proteins for metal bioremediation and recycling three aspects of the metal-interactions with S-layer proteins must be taken into account. First, S-layers possess different functionalities, e.g. carboxyl, phosphoryl groups, binding toxic metals and metalloids, like U(VI) and As(V), nonspecifically depending on pH. Second, precious metals like Au and Pd are likewise nonspecifically bound to functional groups, but presumably covalently, making the binding irreversible. Third, intrinsic specifically bound metals, e.g. Ca²⁺, are needed for native protein folding, self-assembly, and the formation of highly-ordered lattices. Their binding sites also allow selective binding of chemical-equal elements including the trivalent rare earth elements, possessing comparable ionic radii. Thus this study combines older and recently generated results regarding the metal dependent binding behavior of the S-layer proteins. It enables the development of biohybrid materials for the separation, removal or recovery of strategic relevant metals from natural occurring or industrial waste waters using pH-value as regulating parameter for selective metal binding and also conceivably release.

Abstract: Bacteria of the genus Sulfobacillus are successfully used in biotechnologies of treatment of sulfide ore materials. However, energy processes are still poorly understood in sulfobacilli. Several enzymes encoded in the genomes of sulfobacilli are putatively involved in the initial phase of Fe²⁺ oxidation. Cytochromes c, sulfocyanins, and rusticyanin-like proteins were revealed in genomes of these bacteria. Thus, a membrane bound cytochrome c can be reduced directly by Fe²⁺, and electrons can be further transferred to sulfocyanin, rusticyanin or some other components of the transport chain. HPLC identification of prosthetic groups of cytochromes predicts the possible presence of heme c in the cells of thermotolerant sulfobacillus. Determination of metal content in the cells shows that sulfobacillus harbours higher levels of copper ions under specific conditions. This can be explained by higher expression of copper-containing proteins, including cupredoxins.