Bioleaching of pyritic coal wastes: bioprospecting and efficiency of selected consortia.

Pyrite-bearing coal wastes are responsible of the formation of acid mine drainage (AMD), and their management to mitigate environmental impacts is a challenge to the coal mine industry in Europe and worldwide. The European CEReS project sought to develop a generic co-processing strategy to reuse and recycle coal wastes, based on removal of AMD generating potential through bioleaching. Chemolitoautotrophic iron- and sulfur-oxidizing microbial consortia were enriched from a Polish coal waste at 30 °C and 48 °C, but not 42 °C. Pyrite leaching yield, determined from bioleaching tests in 2-L stirred bioreactors, was best with the 48 °C endogenous consortium (80%), then the 42 °C exogenous BRGM-KCC consortium (71%), and finally the 30 °C endogenous consortium (50%). 16S rRNA gene-targeted metagenomics from five surface locations on the dump waste revealed a microbial community adapted to the site context, composed of iron- and/or sulfur-oxidizing genera thriving in low pH and metal rich environments and involved in AMD generation. All together, the results confirmed the predisposition of the pyritic coal waste to bioleaching and the potential of endogenous microorganisms for efficient bioleaching at 48 °C. The good leaching yields open the perspective to optimize further and scale-up the bioleaching process.

Anthropogenic remediation of heavy metals selects against natural microbial remediation.

In an era of unprecedented environmental change, there have been increasing ecological and global public health concerns associated with exposure to anthropogenic pollutants. While there is a pressing need to remediate polluted ecosystems, human intervention might unwittingly oppose selection for natural detoxification, which is primarily carried out by microbes. We test this possibility in the context of a ubiquitous chemical remediation strategy aimed at targeting metal pollution: the addition of lime-containing materials. Here, we show that raising pH by liming decreased the availability of toxic metals in acidic mine-degraded soils, but as a consequence selected against microbial taxa that naturally remediate soil through the production of metal-binding siderophores. Our results therefore highlight the crucial need to consider the eco-evolutionary consequences of human environmental strategies on microbial ecosystem services and other traits.

Multiple Osmotic Stress Responses in Acidihalobacter prosperus Result in Tolerance to Chloride Ions.

Extremely acidophilic microorganisms (pH optima for growth of ≤3) are utilized for the extraction of metals from sulfide minerals in the industrial biotechnology of "biomining." A long term goal for biomining has been development of microbial consortia able to withstand increased chloride concentrations for use in regions where freshwater is scarce. However, when challenged by elevated salt, acidophiles experience both osmotic stress and an acidification of the cytoplasm due to a collapse of the inside positive membrane potential, leading to an influx of protons. In this study, we tested the ability of the halotolerant acidophile Acidihalobacter prosperus to grow and catalyze sulfide mineral dissolution in elevated concentrations of salt and identified chloride tolerance mechanisms in Ac. prosperus as well as the chloride susceptible species, Acidithiobacillus ferrooxidans. Ac. prosperus had optimum iron oxidation at 20 g L-1 NaCl while At. ferrooxidans iron oxidation was inhibited in the presence of 6 g L-1 NaCl. The tolerance to chloride in Ac. prosperus was consistent with electron microscopy, determination of cell viability, and bioleaching capability. The Ac. prosperus proteomic response to elevated chloride concentrations included the production of osmotic stress regulators that potentially induced production of the compatible solute, ectoine uptake protein, and increased iron oxidation resulting in heightened electron flow to drive proton export by the F0F1 ATPase. In contrast, At. ferrooxidans responded to low levels of Cl- with a generalized stress response, decreased iron oxidation, and an increase in central carbon metabolism. One potential adaptation to high chloride in the Ac. prosperus Rus protein involved in ferrous iron oxidation was an increase in the negativity of the surface potential of Rus Form I (and Form II) that could help explain how it can be active under elevated chloride concentrations. These data have been used to create a model of chloride tolerance in the salt tolerant and susceptible species Ac. prosperus and At. ferrooxidans, respectively.

Biodiversity and geochemistry of an extremely acidic, low-temperature subterranean environment sustained by chemolithotrophy.

The geochemical dynamics and composition of microbial communities within a low-temperature (≈ 8.5°C), long-abandoned (> 90 years) underground pyrite mine (Cae Coch, located in north Wales) were investigated. Surface water percolating through fractures in the residual pyrite ore body that forms the roof of the mine becomes extremely acidic and iron-enriched due to microbially accelerated oxidative dissolution of the sulfide mineral. Water droplets on the mine roof were found to host a very limited diversity of exclusively autotrophic microorganisms, dominated by the recently described psychrotolerant iron/sulfur-oxidizing acidophile Acidithiobacillus ferrivorans, and smaller numbers of iron-oxidizing Leptospirillum ferrooxidans. In contrast, flowing water within the mine chamber was colonized with vast macroscopic microbial growths, in the form of acid streamers and microbial stalactites, where the dominant microorganisms were Betaproteobacteria (autotrophic iron oxidizers such as 'Ferrovum myxofaciens' and a bacterium related to Gallionella ferruginea). An isolated pool within the mine showed some similarity (although greater biodiversity) to the roof droplets, and was the only site where archaea were relatively abundant. Bacteria not previously associated with extremely acidic, metal-rich environments (a Sphingomonas sp. and Ralstonia pickettii) were found within the abandoned mine. Data supported the hypothesis that the Cae Coch ecosystem is underpinned by acidophilic, mostly autotrophic, bacteria that use ferrous iron present in the pyrite ore body as their source of energy, with a limited role for sulfur-based autotrophy. Results of this study highlight the importance of novel bacterial species (At. ferrivorans and acidophilic iron-oxidizing Betaproteobacteria) in mediating mineral oxidation and redox transformations of iron in acidic, low-temperature environments.

Structure, function, and evolution of the Thiomonas spp. genome.

Bacteria of the Thiomonas genus are ubiquitous in extreme environments, such as arsenic-rich acid mine drainage (AMD). The genome of one of these strains, Thiomonas sp. 3As, was sequenced, annotated, and examined, revealing specific adaptations allowing this bacterium to survive and grow in its highly toxic environment. In order to explore genomic diversity as well as genetic evolution in Thiomonas spp., a comparative genomic hybridization (CGH) approach was used on eight different strains of the Thiomonas genus, including five strains of the same species. Our results suggest that the Thiomonas genome has evolved through the gain or loss of genomic islands and that this evolution is influenced by the specific environmental conditions in which the strains live.

Carbon and arsenic metabolism in Thiomonas strains: differences revealed diverse adaptation processes.

BACKGROUND: Thiomonas strains are ubiquitous in arsenic-contaminated environments. Differences between Thiomonas strains in the way they have adapted and respond to arsenic have never been studied in detail. For this purpose, five Thiomonas strains, that are interesting in terms of arsenic metabolism were selected: T. arsenivorans, Thiomonas spp. WJ68 and 3As are able to oxidise As(III), while Thiomonas sp. Ynys1 and T. perometabolis are not. Moreover, T. arsenivorans and 3As present interesting physiological traits, in particular that these strains are able to use As(III) as an electron donor. RESULTS: The metabolism of carbon and arsenic was compared in the five Thiomonas strains belonging to two distinct phylogenetic groups. Greater physiological differences were found between these strains than might have been suggested by 16S rRNA/rpoA gene phylogeny, especially regarding arsenic metabolism. Physiologically, T. perometabolis and Ynys1 were unable to oxidise As(III) and were less arsenic-resistant than the other strains. Genetically, they appeared to lack the aox arsenic-oxidising genes and carried only a single ars arsenic resistance operon. Thiomonas arsenivorans belonged to a distinct phylogenetic group and increased its autotrophic metabolism when arsenic concentration increased. Differential proteomic analysis revealed that in T. arsenivorans, the rbc/cbb genes involved in the assimilation of inorganic carbon were induced in the presence of arsenic, whereas these genes were repressed in Thiomonas sp. 3As. CONCLUSION: Taken together, these results show that these closely related bacteria differ substantially in their response to arsenic, amongst other factors, and suggest different relationships between carbon assimilation and arsenic metabolism.

Dissimilatory ferrous iron oxidation at a low pH: a novel trait identified in the bacterial subclass Rubrobacteridae.

A novel iron-oxidizing acidophilic actinobacterium was isolated from spoil material at an abandoned copper mine. Phylogenetic analysis placed the isolate within the Rubrobacteridae subclass of the Actinobacteria. Its optimum temperature and pH for growth are 30-35 degrees C and pH 3.0, respectively. Although it could catalyze the dissimilatory oxidation of ferrous iron, growth yields declined progressively in media containing ferrous iron concentrations >100 microM. The isolate, Pa33, did not grow or oxidize iron in the absence of organic carbon, and appeared to be an obligate heterotroph. Specific rates of iron oxidation were much smaller than those determined for the autotrophic iron-oxidizing proteobacterium Acidithiobacillus ferrooxidans and the heterotrophic iron-oxidizing actinobacterium Ferrimicrobium acidiphilum. Iron oxidation by isolate Pa33 appears to be a defensive mechanism, in which iron oxidation converts a soluble species to which the bacterium is sensitive to an oxidized species (ferric iron) that is highly insoluble in the spoil from which it was isolated. This is the first report of acidophily or dissimilatory iron oxidation within the Rubrobacteridae subclass and one of very few within the Actinobacteria phylum as a whole.