Intermediary sulfur compounds in pyrite oxidation: implications for bioleaching and biodepyritization of coal

Accumulation of elemental sulfur during pyrite oxidation lowers the efficiency of coal desulfurization and bioleaching. In the case of pyrite bioleaching by Leptospirillum ferrooxidans, an iron(II)-ion-oxidizing organism without sulfur-oxidizing capacity, from the pyritic sulfur moiety about 10% elemental sulfur, 2% pentathionate, and 1% tetrathionate accumulated by a recently described cyclic pyrite oxidation mechanism. In the case of pure cultures of Thiobacillus ferrooxidans and mixed cultures of L. ferrooxidans and T. thiooxidans, pyrite was nearly completely oxidized to sulfate because of the capacity of these cultures to oxidize both iron(II) ions and sulfur compounds. Pyrite oxidation in acidic solutions, mediated chemically by iron(III) ion, resulted in an accumulation of similar amounts of sulfur compounds as obtained with L. ferrooxidans. Changes of pH to values below 2 or in the iron ion concentration are not decisive for diverting the flux of sulfur compounds. The literature on pyrite bioleaching is in agreement with the findings indicating that the chemistry of direct and indirect pyrite leaching is identical. Received: 20 April 1998 / Received revision: 27 August 1998 / Accepted: 3 September 1998     

AHL signaling molecules with a large acyl chain enhance biofilm formation on sulfur and metal sulfides by the bioleaching bacterium Acidithiobacillus ferrooxidans

Biofilm formation plays a pivotal role in bioleaching activities of bacteria in both industrial and natural environments. Here, by visualizing attached bacterial cells on energetic substrates with different microscopy techniques, we obtained the first direct evidence that it is possible to positively modulate biofilm formation of the extremophilic bacterium Acidithiobacillus ferrooxidans on sulfur and pyrite surfaces by using Quorum Sensing molecules of the N-acylhomoserine lactone type (AHLs). Our results revealed that AHL-signaling molecules with a long acyl chain (12 or 14 carbons) increased the adhesion of A. ferrooxidans cells to these substrates. In addition, Card-Fish experiments demonstrated that C14-AHL improved the adhesion of indigenous A. ferrooxidans cells from a mixed bioleaching community to pyrite. Finally, we demonstrated that this improvement of cell adhesion is correlated with an increased production of extracellular polymeric substances. Our results open up a promising means to develop new strategies for the improvement of bioleaching efficiency and metal recovery, which could also be used to control environmental damage caused by acid mine/rock drainage.     

Leaching of Pyrites of Various Reactivities by Thiobacillus ferrooxidans

Wide variations were found in the rate of chemical and microbiological leaching of iron from pyritic materials from various sources. Thiobacillus ferrooxidans accelerated leaching of iron from all of the pyritic materials tested in shake flask suspensions at loadings of 0.4% (wt/vol) pulp density. The most chemically reactive pyrites exhibited the fastest bioleaching rates. However, at 2.0% pulp density, a delay in onset of bioleaching occurred with two of the pyrites derived from coal sources. T. ferrooxidans was unable to oxidize the most chemically reactive pyrite at 2.0% pulp density. No inhibition of pyrite oxidation by T. ferrooxidans occurred with mineral pyrite at 2.0% pulp density. Experiments with the most chemically reactive pyrite indicated that the leachates from the material were not inhibitory to iron oxidation by T. ferrooxidans.     

The extremophile Acidithiobacillus ferrooxidans possesses a c-di-GMP signalling pathway that could play a significant role during bioleaching of minerals

Aims: The primary goal of this study was to characterize the existence of a functional c‐di‐GMP pathway in the bioleaching bacterium Acidithiobacillus ferrooxidans. Methods and Results: A bioinformatic search revealed that the genome sequence of At. ferrooxidans ATCC 23270 codes for several proteins involved in the c‐di‐GMP pathway, including diguanylate cyclases (DGC), phosphodiesterases and PilZ effector proteins. Overexpression in Escherichia coli demonstrated that four At. ferrooxidans genes code for proteins containing GGDEF/EAL domains with functional DGC activity. MS/MS analysis allowed the identification of c‐di‐GMP in nucleotide preparations obtained from At. ferrooxidans cells. In addition, c‐di‐GMP levels in cells grown on the surface of solid energetic substrates such as sulfur prills or pyrite were higher than those measured in ferrous iron planktonic cells. Conclusions: At. ferrooxidans possesses a functional c‐di‐GMP pathway that could play a key role in At. ferrooxidans biofilm formation during bioleaching processes. Significance and Impact of the Study: This is the first global study about the c‐di‐GMP pathway in an acidophilic bacterium of great interest for the biomining industry. It opens a new way to explore the regulation of biofilm formation by biomining micro‐organisms during the bioleaching process.     

Investigation of energy gene expressions and community structures of free and attached acidophilic bacteria in chalcopyrite bioleaching

In order to better understand the bioleaching mechanism, expression of genes involved in energy conservation and community structure of free and attached acidophilic bacteria in chalcopyrite bioleaching were investigated. Using quantitative real-time PCR, we studied the expression of genes involved in energy conservation in free and attached Acidithiobacillus ferrooxidans during bioleaching of chalcopyrite. Sulfur oxidation genes of attached A. ferrooxidans were up-regulated while ferrous iron oxidation genes were down-regulated compared with free A. ferrooxidans in the solution. The up-regulation may be induced by elemental sulfur on the mineral surface. This conclusion was supported by the results of HPLC analysis. Sulfur-oxidizing Acidithiobacillus thiooxidans and ferrous-oxidizing Leptospirillum ferrooxidans were the members of the mixed culture in chalcopyrite bioleaching. Study of the community structure of free and attached bacteria showed that A. thiooxidans dominated the attached bacteria while L. ferrooxidans dominated the free bacteria. With respect to available energy sources during bioleaching of chalcopyrite, sulfur-oxidizers tend to be on the mineral surfaces whereas ferrous iron-oxidizers tend to be suspended in the aqueous phase. Taken together, these results indicate that the main role of attached acidophilic bacteria was to oxidize elemental sulfur and dissolution of chalcopyrite involved chiefly an indirect bioleaching mechanism.     

Corrosion and Electrochemical Oxidation of a Pyrite by Thiobacillus ferrooxidans

The oxidation of a pure pyrite by Thiobacillus ferrooxidans is not really a constant phenomenon; it must be considered to be more like a succession of different steps which need characterization. Electrochemical studies using a combination of a platinum electrode and a specific pyrite electrode (packed-ground-pyrite electrode) revealed four steps in the bioleaching process. Each step can be identified by the electrochemical behavior (redox potentials) of pyrite, which in turn can be related to chemical (leachate content), bacterial (growth), and physical (corrosion patterns) parameters of the leaching process. A comparison of the oxidation rates of iron and sulfur indicated the nonstoichiometric bacterial oxidation of a pure pyrite in which superficial phenomena, aqueous oxidation, and deep crystal dissolution are successively involved.     

Optimization of Lithotrophic Activities of Acidithiobacillus ferrooxidans toward Significant Reduction of Sulfur and Ash from Low Rank Bitumen

In the present study, the potential application of Acidithiobacillus ferrooxidans for elimination of ash and sulfur from bitumen was investigated in batch experiments. A comparison between the bioleaching and abiotic treatments indicated that A. ferrooxidans cells enhanced ash and pyritic sulfur removal by 20 and 59%, respectively. The X-ray diffraction profiles of the samples indicated the precipitation of some mineral elements inside of bitumen decreased the bioleaching performance after 9 days from beginning of the experiments. The effects of bitumen particle size (X₁), agitation speed (X₂) and initial pH (X₃) as interfacial factors each at three levels on the ash removal (Y₁) and pyritic sulfur removal (Y₂) were investigated by response surface methodology as a statistical design of the experiment. On the basis of quadratic models applied to the performance of the bioleaching process, 66.42% of the pyritic sulfur and 50.88% of the ash could be removed after 9 days under optimal conditions, namely a bitumen particle size of 100 µm, an agitation speed of 80 rpm, and initial pH of 2.     

Manipulation of pyrite colonization and leaching by iron-oxidizing <Emphasis Type="Italic">Acidithiobacillus</Emphasis> species

In this study, the process of pyrite colonization and leaching by three iron-oxidizing Acidithiobacillus species was investigated by fluorescence microscopy, bacterial attachment, and leaching assays. Within the first 4–5 days, only the biofilm subpopulation was responsible for pyrite dissolution. Pyrite-grown cells, in contrast to iron-grown cells, were able to oxidize iron(II) ions or pyrite after 24 h iron starvation and incubation with 1 mM H₂O₂, indicating that these cells were adapted to the presence of enhanced levels of reactive oxygen species (ROS), which are generated on metal sulfide surfaces. Acidithiobacillus ferrivorans SS3 and Acidithiobacillus ferrooxidans R1 showed enhanced pyrite colonization and biofilm formation compared to A. ferrooxidans ^T. A broad range of factors influencing the biofilm formation on pyrite were also identified, some of them were strain-specific. Cultivation at non-optimum growth temperatures or increased ionic strength led to a decreased colonization of pyrite. The presence of iron(III) ions increased pyrite colonization, especially when pyrite-grown cells were used, while the addition of 20 mM copper(II) ions resulted in reduced biofilm formation on pyrite. This observation correlated with a different extracellular polymeric substance (EPS) composition of copper-exposed cells. Interestingly, the addition of 1 mM sodium glucuronate in combination with iron(III) ions led to a 5-fold and 7-fold increased cell attachment after 1 and 8 days of incubation, respectively, in A. ferrooxidans ^T. In addition, sodium glucuronate addition enhanced pyrite dissolution by 25 %.     

Cathodic behaviour of stainless steel in coastal Indian seawater: calcareous deposits overwhelm biofilms

Type-316 stainless steel (SS) was investigated as the cathode in galvanic couples in full-strength seawater from the Gulf of Mannar on the southeast coast of India. Tests were devised to examine the impact of SS cathodes on anode materials with or without the accrual of marine biofilms. Biofilmed SS cathodes significantly enhanced the rate of corrosion of nickel, causing noble shifts in the couple potentials. With mild steel and zinc as the anodes, calcareous deposits developed quite rapidly on the SS cathodes and led to a significant reduction of bacterial numbers. The calcareous deposits also caused substantial reduction of galvanic corrosion rates for mild steel, whereas there was no difference for zinc. The deposits were identified by XRD as essentially carbonates, oxides and hydroxides of calcium and magnesium. Potentiodynamic polarization performed on the actual couples after disconnection and equilibration provided reasonable interpretations of the galvanic corrosion trends. Data from this work suggest that a potential of about ?0.70 V vs. saturated calomel electrode (SCE) should provide optimum protection of SS in warmer, full-strength seawater that supports the precipitation of calcareous deposits. The criterion commonly recommended for temperate conditions of lower water temperature and estuarine waters of lower alkalinity is ?1.0 V (SCE).     

Bulk phase resource ratio alters carbon steel corrosion rates and endogenously produced extracellular electron transfer mediators in a sulfate-reducing biofilm

Desulfovibrio alaskensis G20 biofilms were cultivated on 316 steel, 1018 steel, or borosilicate glass under steady-state conditions in electron-acceptor limiting (EAL) and electron-donor limiting (EDL) conditions with lactate and sulfate in a defined medium. Increased corrosion was observed on 1018 steel under EDL conditions compared to 316 steel, and biofilms on 1018 carbon steel under the EDL condition had at least twofold higher corrosion rates compared to the EAL condition. Protecting the 1018 metal coupon from biofilm colonization significantly reduced corrosion, suggesting that the corrosion mechanism was enhanced through attachment between the material and the biofilm. Metabolomic mass spectrometry analyses demonstrated an increase in a flavin-like molecule under the 1018 EDL condition and sulfonates under the 1018 EAL condition. These data indicate the importance of S-cycling under the EAL condition, and that the EDL is associated with increased biocorrosion via indirect extracellular electron transfer mediated by endogenously produced flavin-like molecules.     

Short communication: Adverse effect of surface-active reagents on the bioleaching of pyrite and chalcopyrite by Thiobacillus ferrooxidans

Oxidation of Fe(II) iron and bioleaching of pyrite and chalcopyrite by Thiobacillus ferrooxidans was adversely affected by isopropylxanthate, a flotation agent, and by LIX 984, a solvent-extraction agent, each at 1 g/l. The reagents/l were adsorbed on the bacterial surface, decreasing the bacteria's development and preventing biooxidation. Both reagents inhibited the bioleaching of pyrite and LIX 984 also inhibited the bioleaching of chalcopyrite.     

Microbial desulfurization of coal and oxidation of pure pyrite byThiobacillus ferrooxidans andAcidianus brierleyi

Summary Thiobacillus ferrooxidans andAcidianus brierleyi were capable of oxidizing pure pyrite as well as oxidizing sulfur in coal. First order reactions were assumed in the kinetic analysis performed. For oxidation of pure pyrite the rate constant was higher forA. brierleyi than forT. ferrooxidans. For sulfur removal from coal the values of the rate constants were comparable for the two microorganisms.     

Evaluation of the Biotechnological Potential of Pure and Mixture of Indigenous Iron-Oxidizing Bacteria to Dissolve Trace Metals from Cu Bearing Ore

Abstract

This study aimed to investigate the ability of pure and consortia of indigenous iron-oxidizing bacteria to enhance the dissolution of trace metals from Cu and Zn-bearing ore. Three bacterial strains Acidithiobacillus ferrooxidans strain WG101, Leptospirillum ferriphilum strain WG102, Leptospirillum ferrooxidans strain WG103 isolated from Baiyin copper mine, China were used in this study. The biotechnological potential of these indigenous isolates was evaluated both in pure and in consortia to extract cobalt, chromium, and lead from the copper and zinc bearing ore. The sulfur and iron-oxidizing bacterial isolate Acidithiobacillus ferrooxidans strain WG101 exhibited efficient dissolution compared to sole iron-oxidizing Leptospirillum ferriphilum strain WG102, and Leptospirillum ferrooxidans strain WG103. Initial medium pH, pulp density, and temperature were studied as influential parameters in bioleaching carried out by bacterial consortia. The achieved optimum conditions were; initial pH of 1.5, 10% of pulp density, and temperature 30?°C with 68.7?±?3.9% cobalt, 56.6?±?3.9% chromium, and 36?±?3.7% lead recovery. Analytical study of oxidation-reduction potential and pH fluctuation were observed during this whole process that shows the metal dissolution efficiency of bacterial consortia. Alterations in spectral bands of processed residues were reported through FTIR analysis compared with control ore sample. Mössbauer spectroscopy analysis showed the influence of bacterial consortia on iron speciation in bioleached samples. The findings confirm that the indigenous acidophilic iron-oxidizing bacterial strains are highly effective in the dissolution of trace elements present in ore samples. This study not only supports the notion that indigenous bacterial strains are highly effectual in metal dissolution but provides the basic vital conditions to upscale the bioleaching technique for metals dissolution.     

Gene expression modulation by chalcopyrite and bornite in Acidithiobacillus ferrooxidans

Acidithiobacillus ferrooxidans is a mesophilic, acidophilic, chemolithoautotrophic bacterium that obtains energy from the oxidation of ferrous iron (Fe²⁺), elemental sulfur and reduced sulfur compounds. The industrial interest in A. ferrooxidans resides in its capacity to oxidize insoluble metal sulfides into soluble metal sulfates, thus allowing the recovery of the desired metals from low-grade sulfide ores. In the present work, RNA arbitrarily primed PCR (RAP-PCR) was performed to identify cDNAs differentially expressed in A. ferrooxidans cells grown in the presence of Fe²⁺ and cells maintained for 24 h in the presence of the copper sulfides bornite and chalcopyrite. Eighteen cDNAs corresponding to genes with known function were identified, and their relative expression was further characterized by real-time quantitative PCR. Bornite had a mild effect on the expression of the 18 genes analyzed. None of these genes was down-regulated and among the few genes up-regulated, it is worth mentioning lepA and def-2 that are involved in protein synthesis. Chalcopyrite presented the most significant changes. Five genes related to protein processing were down-regulated, and another 5 genes related to the transport system were up-regulated. The up- and down-regulation of these genes in the presence of bornite and chalcopyrite could be due to alterations in the ideal pH, presence of copper ions in solution and nutrient limitation. The results suggest that gene expression modulation might be important for the A. ferrooxidans early response to copper sulfides.     

Rate of Pyrite Bioleaching by Thiobacillus ferrooxidans: Results of an Interlaboratory Comparison

Ten laboratories participated in an interlaboratory comparison of determination of bioleaching rates of a pyrite reference material. A standardized procedure and a single strain of Thiobacillus ferrooxidans were used in this study. The mean rate of bioleaching of the pyrite reference material was 12.4 mg of Fe per liter per h, with a coefficient of variation (percent relative standard deviation) of 32% as determined by eight laboratories. These results show the precision among laboratories of the determination of rates of pyrite bioleaching when a standard test procedure and reference material are used.     

Sulfur Chemistry in Bacterial Leaching of Pyrite

In the case of pyrite bioleaching by Leptospirillum ferrooxidans, an organism without sulfur-oxidizing capacity, besides the production of tetra- and pentathionate, a considerable accumulation of elemental sulfur occurred. A similar result was obtained for chemical oxidation assays with acidic, sterile iron(III) ion-containing solutions. In the case of Thiobacillus ferrooxidans, only slight amounts of elemental sulfur were detectable because of the organism's capacity to oxidize sulfur compounds. In the course of oxidative, chemical pyrite degradation under alkaline conditions, the accumulation of tetrathionate, trithionate, and thiosulfate occurred. The data indicate that thiosulfate, trithionate, tetrathionate, and disulfane-monosulfonic acid are key intermediate sulfur compounds in oxidative pyrite degradation. A novel (cyclic) leaching mechanism is proposed which basically is indirect.     

Wear Resistance and Non-Magnetic Layer Formation on 316L Implant Material with Plasma Nitriding

In this study, the applicability of plasma nitriding treatment in the production of non-magnetic and corrosion resistant layer on 316L stainless steel implant material was investigated. 316L stainless steel substrates were plasma nitrided at temperatures of 350 ℃, 375 ℃, 400 ℃, 425 ℃ and 450 ℃ for 2 h in a gas mixture of 50% N2-50% H2, respectively. It was determined that the treatment temperature is the most important factor on the properties of the corrosion resistant layer of 316L stainless steel. The results show that s-phase formed at the temperatures under 400 ℃, and at the temperatures above 400 ℃, instead of s-phase, CrN and y＇-Fe4N phases were observed in the modified layer. The electrical resistivity and surface roughness of the modified layer increase with treatment temperature. Under 400 ℃ the corrosion resistance increased with the temperature, above 400 ℃ it decreased with the increase in treatment temperature. It was analyzed that the electrical resistivity and the soft （ideal） ferro- magnetic properties of 316L stainless steel increased with treatment temperature during nitriding treatment. Also, plasma ni- triding at low temperatures provided magnetic behavior close to the ideal untreated 316L stainless steel.