Advanced Materials Research Vols. 20-21

Abstract: The genus Sulfobacillus includes moderately thermophilic and thermotolerant acidophilic bacteria, which prefer mixotrophic growth conditions. These organisms are S. thermosulfidooxidans 1269Т, S. sibiricus N1Т and S. thermotolerans Kr1Т. The simultaneous presence of organic (for example yeast extract) and inorganic energy sources is essential for optimal growth. Carbon sources are both CO2 and organic compounds. During their mixotrophic growth, the basic pathways of carbohydrate catabolism of sulfobacilli are those of glycolysis, oxidative pentose phosphate and Entner–Doudorov (excepting strain N1). Assimilation of CO2 is carried out both auto- and heterotrophically. Final stages of oxidation of organic substances serve for cell biosynthesis. Bacteria are able to switch to organo- and autotrophic metabolism. At that the enzyme activities of pentose phosphate pathway are not detected. The cycle of tricarboxylic acids is disrupted at the level of 2-oxoglutarate dehydrogenase. The glyoxylate bypass is absent. The maximal protein and ATP values and high intensity of respiration in S. thermosulfidooxidans, S. sibiricus and S. thermotolerans cultures were determined under mixotrophic conditions. Studied strains due to their flexible carbon and energetic metabolism together with other microorganisms of communities (leptospirilli and archaea) participate in sulfide minerals processing at 40-450C without organic compounds supplement.

Abstract: The acidophilic heterotrophic bacteria Acidiphilium spp. were considered as a ministrant of Leptospirillum spp., Acidithiobacillus spp. and other autotrophic bacteria in acid mine drainage (AMD). 25 Acidiphilium-like bacterial strains were isolated from eight different mines in China and their physiology was characterized. Their marmatite-bioleaching ability has been investigated and compared with that of a mixed culture of Acidiphilium sp. and Acidithiobacillus ferrooxidans. The results indicated that some Acidiphilium-like strains exhibited a high marmatite-bioleaching ability, which was even higher than that of Acidithiobacillus ferrooxidans. The leaching efficiency was not higher in mixed than in pure culture. The phylogenetic relationship of the 25 Acidiphilium-like strains was analyzed by 16S rRNA sequencing, GyrB sequencing and rep-PCR genomic fingerprinting. The 25 strains and other eight species of Acidiphilium spp. were clustered into three groups.

Abstract: We investigated the oxidation of elemental sulfur in resting cells of Acidithiobacillus ferrooxidans in order to optimize the substrate for sulfur oxidation and to explore oxidation kinetics. We showed that although pH and temperature had no significant effect on the apparent Michaelis constant, cell concentration increased it.

Abstract: Cemented layers predominantly consisting of gels/poorly crystalline mineral phases have been formed as a consequence of mineral weathering in sulfidic tailings near Freiberg, Saxony, Germany. These layers function as natural attenuation barrier for toxic compounds and reduce oxidation and erosion processes of tailings surfaces. Quantitative molecular biological and cultivation methods were applied to investigate the role of microorganisms for mineral weathering and cemented layer formation. High resolution depth profiles of numbers of microorganisms showed maximal cell numbers in the oxidation zone where cemented layers had been formed. Highest total cell numbers of >109 cells g-1 dry weight (dw) were detected by SybrGreen direct counting. Using quantitative real-time PCR (Q-PCR) between 107 and 109 Bacteria g-1 dw and up to 108 Archaea g-1 dw were determined. As well high numbers of cultivable and living Bacteria could be detected by MPN (most probable number) for Fe(II)- and S-oxidizers and CARD-FISH (catalyzed reporter deposition - fluorescence in situ hybridization). Overall, the high numbers of microorganisms determined with various quantification techniques argue for a significant role of microorganisms in cemented layer formation due to microbial mineral weathering. It is hypothesized that EPS (extracellular polymeric substances) mediate the formation of secondary mineral phases.

Abstract: A culture independent molecular methodology was used to investigate the bacterial and archaeal microbial dynamics of leachate collected from a 60°C chalcopyrite bioleaching column inoculated with a known microbial consortium. A 16S rRNA gene clone library was constructed for both the bacterial and archaeal populations in the leachate from the column. PCR-RFLP analysis of these clone libraries indicate species dominance and generally low species diversity.

Abstract: The sediment microbial communities of a disused coal mine lake, Lake Kepwari (pH~4.5-5) were studied to understand how the natural microbial processes in an oligotrophic acidic mine lake system influence the iron and sulphur cycles. Most probable number (MPN) viable counts were used to enumerate the benthic bacteria at different depths. MPN results revealed an abundance of bacteria that were capable of growing in sulphate reducing medium with numbers in the range of 1 × 107 – 1 × 108 cells.g-1 of wet sediment. In contrast, MPN results showed much lower numbers of bacteria that were capable of growing in ferric reducing medium with 1 × 102 – 2 × 103 cells.g-1 of wet sediment detected. Serial decimal dilution cultures were used to isolate pure strains of benthic bacteria. Strains HP1, HP2 and HP3 were isolated from benthic lake sediments at 18 m, 0 m and 10 m water depths respectively. 16S rRNA gene sequence analysis of strain HP1 showed that the strain belonged to the genus Enterobacter, strain HP2 belonged to the Order Rhizobiales and strain HP3 belonged to the sub-order Micrococcineae. Denaturing gradient gel electrophoresis (DGGE) analysis of 16S rRNA gene fragments was used to profile the diversity of the benthic microbial communities at different depths. DGGE profiling of benthic sediments revealed that sediments contained mostly members of the Proteobacteria, Actinobacteria and Firmicutes phyla.

Abstract: This paper describes about microbial diversity in an iron oxidation tank of an AMD treatment plant established at an abandoned sulphur mine in Japan. Since the mining operation was stopped, this mine has produced strong acidic mine drainage, pH 1.8, including a high concentration of ferrous iron, 301mg/L, and the flow rate was about 4m3/min. In 2006, a pilot scale microbial iron oxidizing system was installed to remove total iron more easily from the AMD by oxidizing ferrous iron to ferric iron. From the start of this pilot operation, microbial diversity in the iron oxidation tank was investigated using a PCR-DGGE method for about two months. In the PCR, V3 region of 16S rRNA gene for Bacteria was amplified. The profiles of DGGE showed that there were three dominant species in the iron oxidation tank through the experimental period. Number of bands on DGGE profiles decreased with dates of sampling so that the microbial population became less diverse because of the iron oxidizing operation. There was a wide variety of bacterial species of even though conditions were strongly acidic.

Abstract: Facultative Fe(III)-reducing bacterium Acidiphilium SJH was incubated in media with ferric iron under various conditions with respect to oxygen availability for the growing cells. The bacteria oxidized organic substratum to carbon dioxide using oxygen and ferric iron as terminal electron acceptors. Ferric iron reduction was observed in all incubation modes. The distribution of reducing equivalents from the oxidation of organic carbon for the reduction of both O2 and Fe(III) was evaluated from CO2 production rate and O2 consumption rate. In fully aerobic conditions approximately 10 % of CO2 produced was coupled with reduction of Fe(III) as terminal electron acceptor. Under aerobic conditions, the ratio of CO2 produced to O2 consumed remained unaffected in a broad concentration range of dissolved oxygen. In the course of oxygen limitation (microaerobic conditions) the molar CO2 to O2 ratio increased from approx. 1 to 2 and even much more with respect to oxygen transfer rate during incubation. On the other hand no bacterial growth and extremely slow iron reduction was observed in obligatory anaerobic conditions in a reactor purged with either pure or CO2-enriched nitrogen.

Abstract: Acidophilic iron-oxidizing microorganisms are important for both, the environment and for biotechnological applications. Biogeneration of ferric from ferrous iron was studied using an iron-oxidizing consortium developed during polymetallic concentrate bioleaching. A promising iron oxidizing consortium was developed by adaptation and selection, which resulted in bacterial iron oxidation activity under the extreme conditions of 250 g/L ferrous sulphate as initial substrate and 500 g/L ferric sulphate. The development of iron oxidizers improved the iron oxidation rate from 0.019 to as high as 0.6 g/L/h in the shake flask studies with 25 % initial ferrous sulphate in the medium. The consortium showed dominance of Leptospirillum ferrooxidans and Acidithiobacillus ferrooxidans. A fixed film bioreactor was further developed to improve rates of iron oxidation. The developed fixed film bioreactor operated successfully for 200 batches without external addition of inoculum with the highest iron oxidation rate of 1.89 g/L/h. The biological ferric iron generation process provides continuous leaching agent in the form of ferric sulphate, which would be a promising eco-friendly biotechnological process for the indirect extraction of precious metals from sulphidic mineral concentrates.

Abstract: We present here a method for the electrochemical enrichment of chromium-reducing bacteria from environmental samples containing various microorganisms. An electrochemical cultivation system was constructed; this consisted of a Pt anode and a carbon cathode separated by an ion-exchange membrane. For an electrochemical cultivation, an environmental sample with chromium-reducing bacteria was inoculated into the anode well where a medium containing 0.1 mM Cr(VI) ions was poured. On the other hand, Cr(VI)-free medium was poured into the cathode well. Chromium-reducing bacteria grew selectively after 10 days of incubation and application of 1.0 V (vs. Ag/AgCl) of anode potential that reproduces Cr(VI) ions. This result suggested that Cr(VI) ions generated on the anode promoted the growth of chromium-reducing bacteria as their electron acceptor and due to Cr(VI) toxicity, inhibited the growth of other microorganisms.