Biohydrometallurgy: From the Single Cell to the Environment

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Authors: Djamila Slyemi, Jeanine Ratouchniak, Violaine Bonnefoy
Abstract: At the abandoned mining site of Carnoulès (Gard, France), weathering of the arsenopyrite rich tailings leads to the formation of acidic effluents heavily loaded with arsenite (As(III)). However, further downstream, the As(III) concentration decreases while the sediments are richer in arsenate (As(V)). A Thiomonas sp. able to oxidize arsenite to arsenate has been isolated and characterized. The aoxA and aoxB genes encoding the two subunits of this enzyme belong to an operon. Analysis of the genome sequence (Genoscope, Ivry, France) shows that this operon encodes also two cytochromes c, which could be the physiological partners of the arsenite oxidase, and a transcriptional regulator belonging to the metalloregulator ArsR/SmtB family, which could control the expression of the aox operon. The expression of this operon is higher in the presence than in the absence of As(III) and appears also to be repressed in the presence of thiosulfate, a more energetic substrate.
Authors: Carol S. Davis-Belmar, James Le C. Nicolle, Paul R. Norris
Abstract: Growth on ferrous iron of a new isolate of the halotolerant acidophile “Thiobacillus prosperus” occurred with a substrate oxidation rate similar to that of Acidithiobacillus ferrooxidans, but with a requirement for salt (NaCl). These observations contrast with the previous description of “T. prosperus” in which a salt requirement was not noted and growth on ferrous iron was described as poor. As well as similar capacities for iron oxidation, these species were shown to possess similar clusters of genes (the rus operon) that encode proteins likely to be involved in transfer of electrons from ferrous iron. There were some differences in the organization of the genes and one of them that encodes a cytochrome c in At. ferrooxidans was absent from the “T. prosperus” cluster.
Authors: Pedro A. Galleguillos, V. Zepeda, F. Galleguillos, Danny Castillo, Nicolas Guiliani, Francisco Remonsellez, E. Ortiz, Cecilia Demergasso
Abstract: During the last decades, microbial bioleaching of metallic sulphide ores has become a very important process in the mining industry. The study of biodiversity in those systems has revealed the occurrence of several acidophilic organisms, forming a dynamic community including Bacteria and Archaea. Despite the importance of the heap bioleaching process, the metabolic behaviour of a microbial community in bioleaching systems remains unknown. The role played by members of the community inhabiting a bioheap environment has been estimated by the phenotypic characteristics of pure cultures, such as iron oxidation, sulphur oxidation, resistance to high ion concentration and metal tolerance, among others. The global genomic expression of the community inhabiting an industrial bioheap of low-grade copper sulphide ore at Escondida mine in Chile was investigated by random arbitrary primed polymerase chain reaction (RAP-PCR). Three random primers were used and twenty differentially expressed bands were cloned and sequenced. Several sequences were related to 16S rRNA of members of the microbial community; two sequences were related to the α subunit of pyruvate dehydrogenase from Acidithiobacillus ferrooxidans. The expression levels of the pyruvate dehydrogenase gene in samples from an industrial bioleaching operation at different stages were checked by quantitative real-time PCR.
Authors: Jorge H. Valdés, Inti Pedroso, Raquel Quatrini, Kevin B. Hallberg, Pablo D.T. Valenzuela, David S. Holmes
Abstract: Draft genome sequences of Acidithiobacillus thiooxidans ATCC 19377 and A. caldus ATCC 51756 have been annotated. Bioinformatic analysis of these two new genomes, together with that of A. ferrooxidans ATCC 23270, allows the prediction of metabolic and regulatory models for each species and has provided a unique opportunity to undertake comparative genomic studies of this group of bioleaching bacteria. In this paper, we report preliminary information on metabolic and electron transfer pathways for ten characteristics of the three acidithiobacilli: CO2 fixation, the TCA cycle, sulfur oxidation, sulfur reduction, iron oxidation, iron assimilation, hydrogen oxidation, flagella formation, Che signaling (chemotaxis) and nitrogen fixation. Predicted transcriptional and metabolic interplay between pathways pinpoints potential coordinated responses to environmental signals such as energy source, oxygen and nutrient limitations. The predicted pathway for nitrogen fixation in A. ferrooxidans will be described as an example of such an integrated response. Several responses appear to be especially characteristic of autotrophic microorganisms and may have direct implications for metabolic processes of critical relevance to the understanding of how these microorganisms survive and proliferate in extreme environments, including industrial bioleaching operations.
Authors: Taher M. Taha, Tadayoshi Kanao, Fumiaki Takeuchi, Tsuyoshi Sugio
Abstract: Growth of A. ferrooxidans ATCC 23270 cells in sulfur medium with 0.005% ferric sulfate for 3, 4, 5, 6, 7 and 10 days gave the maximum growth yield of 45, 58, 76, 86, 90 and 95 mg protein per liter medium, respectively. Iron oxidase activities of 1-, 2- and 3- day-cultured cells on sulfur with 0.005% ferric sulfate (3.4, 3.5 and 0.8 μmol Fe2+ oxidized/mg protein/min) were approximately 68, 70 and 16% of iron-grown ATCC 23270 cells (5.0 μmol/mg protein/min). In contrast iron oxidase activities of 1-, 2- and 3-day cultured cells on sulfur without iron (4.9, 3.8 and 2.7 μmol Fe2+ oxidized/mg protein/min) were approximately 98, 76 and 54% of the iron oxidase activity observed in iron-grown ATCC 23270 cell. SFORase activities of 3 day-cultured cell on sulfur with and without ferric sulfate (0.62 and 0.31 μmol Fe2+ produced/mg protein/min) were approximately 20 and 10 fold higher than that of iron-grown cell (0.03 μmol Fe2+ produced/mg protein/min). Both iron oxidase and SFORase activities increased at early-log phase and decreased at late-lag phase during growth of the strain on sulfur with or without Fe3+. The plasma membranes which had iron oxidase activity were prepared not only from iron-grown cells but also sulfur-grown cells. Iron oxidase activities of the plasma membranes prepared from sulfur- and iron-grown cells were 3.6 and 4.5 nmol Fe2+ oxidized per mg protein per min. These results suggest that iron oxidation enzyme system has a role in part in the energy generation of this bacterium from sulfur.
Authors: Jochen Petersen, Tunde Victor Ojumu
Abstract: In this study the results from a systematic study of the oxidation kinetics of Leptospirillum ferriphilum in continuous culture at total iron concentrations ranging from 2 to12 g/L are reported. In all experiments the steady-state concentrations of ferrous iron were small and comparable, and at least 97% of was as ferric. Surprisingly, the specific ferrous iron utilisation rate decreased with increasing total iron concentration, while yield coefficients increased. It was noted that the biomass concentration in the reactor (as measured by both CO2 uptake rate and cell counts) dramatically increased with increasing total iron concentrations, whereas it stayed more or less the same over a wide range of dilution rates at a given total iron concentration. The experimental data was re-analysed in terms of ferrous iron kinetics using Monod kinetics with a ferric inhibition term. The results confirm that the maximum specific iron utilisation rate is itself a function of ferric iron concentration, declining with increasing concentration. It thus appears that high concentrations of ferric iron stimulate microbial growth while at the same time inhibiting the rate of ferrous iron oxidation. It is postulated that these phenomena are related, i.e. that more growth occurs to reduce the load on the individual cell, possibly by sharing some metabolic functions.
Authors: Sei Choong Yee, Choon Ping Lim, Kim Yong Ng
Abstract: A strain of At. ferrooxidans was isolated from Penjom Goldmine, Kuala Lipis, Pahang, Malaysia. The 16S rRNA partial gene sequence analysis revealed that this rod-shaped bacterium with size of 0.3 x 0.8 -m is a strain of At. ferrooxidans, and was designated as At. ferrooxidans KLipis-3-1. The whole-cell FAMEs profile analysis showed that At. ferrooxidans KLipis-3-1 possesses large amounts of C18:1, C16:0 and C16:1 fatty acids with C18:1 as the predominant fatty acid. The initial specific iron-oxidizing and molybdenum blue-oxidizing activities of this bacterium were 88.85 mMh-1mg-1 and 196.86 mMh-1mg-1, respectively, which were similar to the activities observed in the type strain At. ferrooxidans ATCC 23270. However, At. ferrooxidans KLipis-3-1 has a higher resistance towards Mo6+ at concentration above 1 mM. At. ferrooxidans KLipis-3-1 has the ability to reduce ferric ion with elemental sulfur as electron donor, with the initial specific SFORase activity of 0.47 mMh-1mg-1. The bioleaching capability of At. ferrooxidans KLipis-3-1 and At. ferrooxidans ATCC 23270 with pyrite as substrate at selected parameters operated in a batch bioprocess were studied.
Authors: Carla M. Zammit, L.A. Mutch, Helen R. Watling, Elizabeth L.J. Watkin
Authors: Ana P. Felício, Eliandre de Oliveira, Maria A. Odean, Oswaldo Garcia Jr., Maria C. Bertolini, Lúcio F.C. Ferraz, Fernanda C. Reis, Laura M.M. Ottoboni, Maria T.M. Novo
Abstract: Acidithiobacillus ferrooxidans is used in bioleaching industrial operations to recover metal ions from mineral sulfides. Chalcopyrite and bornite are copper sulfides that have the same elemental composition, but differ in their susceptibility to the bioleaching process. Our objective was to identify differentially expressed proteins in A. ferrooxidans LR cells exposed to chalcopyrite or bornite, as a sole energy source, for 24 hours. Compared to the control (without minerals), proteins were induced or repressed in planktonic cells after contact with chalcopyrite or bornite by 24 hours. These results demonstrated that the time of exposure to the copper minerals was enough to trigger distinct responses in the A. ferrooxidans metabolism.
Authors: Bestamin Özkaya, Pauliina Nurmi, Erkan Sahinkaya, Anna H. Kaksonen, Jaakko A. Puhakka
Abstract: In this study, ferrous iron oxidation rates of a Leptospirillum ferriphilum dominated culture were determined over the temperature range of 2-50oC at pH below one. The results show that at pH 0.9 the culture oxidizes iron within the temperature range of 10°C to 45°C. Using the Arrhenius equation, an Ea value of 89.9 ± 6.75 kJ/mol was calculated. From the data fitted to Ratkowsky Equation, the optimum, minimum and maximum temperatures were 35 ± 1.5, 9.96 ± 1.72 and 42.93 ± 0.64 °C for this culture, respectively. The redox potential of the solution becomes more positive, which was the maximum (650-700 mV) at temperatures between 19-40 oC due to completing biological oxidation and increasing in ferric iron concentration.

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