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Microbial communities of iron-rich water in the Pyhäsalmi mine, Finland, were investigated with high-throughput amplicon sequencing and qPCR targeting bacteria, archaea, and fungi. In addition, the abundance of Leptospirillum and Acidithiobacillus was assessed with genus-specific qPCR assays, and enrichment cultures targeting aerobic ferrous iron oxidizers and ferric iron reducers were established. The acidic (pH 1.4–2.3) mine water collected from 240 m, 500 m, and 600 m depth from within the mine had a high microbial diversity consisting of 63-114 bacterial, 10-13 archaeal, and 104-117 fungal genera. The most abundant microorganisms in the mine water were typical acid mine drainage (AMD) taxa, such as acidophilic, iron-oxidizing Leptospirillum, Acidiphilum, Acidithiobacillus, Ferrovum, and Thermoplasma. The fungi belonged mostly to the phylum Ascomycetes, although a great part of the fungal sequences remained unclassified. The number of archaeal 16S rRNA genes in the mine water was between 0.3 and 1.2 × 107 copies mL−1 in the samples from 500 m and 600 m, but only 3.9 × 103 at 240 m and archaea were in general not enriched in cultures. The number of fungal 5.8S rRNA genes was high only in the mine water from 500 m and 600 m, where 0.2–3.4 × 104 spore equivalents mL−1 were detected. A high number of Leptospirillum 16S rRNA genes, 0.6–1.6 × 1010 copies mL−1, were detected at 500 m and 600 m depth and in cultures containing ferrous iron, showing the importance of iron oxidizers in this environment. The abundance of bacteria in general was between 103 and 106 16S rRNA gene copies mL−1. Our results showed a high microbial diversity in the acid- and iron-impacted waters of the Pyhäsalmi mine, where Leptospirillum bacteria were especially prominent. These iron oxidizers are also the main nitrogen-fixing microorganisms in this ecosystem. [ABSTRACT FROM AUTHOR]

The long-term safety of final disposal of spent nuclear fuel in the deep geosphere is dependent on stability of biogeochemical conditions at the disposal site. Microbial processes, such as sulphate reduction and methanogenesis, may have profound effects on site biogeochemistry. In this study, sulphate-reducing bacteria and methane-producing archaea were investigated at depths ranging from 68 to 545 m in crystalline rock fractures at an intended spent nuclear fuel disposal site in Olkiluoto, Finland. Denaturing gradient gel electrophoresis detected diverse sulphate-reducing bacterial communities in all samples. Although the number of dsrB gene copies was below 103 copies ml−1 in all analyzed samples according to real-time quantitative PCR, their abundance was highest in samples that had the highest sulphate concentrations. Several distinct mcrA gene fragments were also recovered from most of the analyzed samples by cloning, although the number of methanogens was lower than that of sulphate-reducing bacteria when measured by mcrA-targeted quantitative PCR. The detected gene fragments were most closely related to sequences obtained from aquatic and deep subsurface environments. Results imply that sulphate reduction, methanogenesis, and anaerobic methane oxidation may all take place in the Olkiluoto deep geobiosphere. [ABSTRACT FROM AUTHOR]

Abstract: Members of the euryarchaeotal genera Methanolobus and Halobacterium as well as group 1.1c Crenarchaeota were enriched from ectomycorrhizal samples and cultured under anaerobic conditions. 16S rRNA gene sequences of Methanolobus were obtained in a H2 + CO2 atmosphere and autofluorescent putatively methanogenic microbial cells were detected by epifluorescence microscopy of the anaerobic methane-producing enrichment cultures. Halobacterium and group 1.1c Crenarchaeota grew anaerobically when either H2 or CH4 was added to the atmosphere. Group 1.1c Crenarchaeota were also enriched under aerobic conditions on mineral media, but only when methane or methanol was added as carbon sources. The 16S rRNA gene sequences of 1.1c Crenarchaeota grown under both anaerobic and aerobic conditions were highly similar. Our study demonstrates the growth of group 1.1c Crenarchaeota and Halobacteria derived from non-extreme soil environment in non-saline enrichments under anaerobic conditions. The results suggest that 1.1c Crenarchaeota may play a role in the cycling of C-1 substrates in the boreal forest soil ecosystem. [Copyright &y& Elsevier]

Archaea colonising defined compartments of Scots pine Suillus bovinus or Paxillus involutus mycorrhizospheres developed in forest humus-containing microcosms were investigated by nested polymerase chain reaction (PCR), cloning, restriction fragment length polymorphism (RFLP) and sequencing. Archaea representing six RFLP groups were detected in the system. Sequence analysis of clones representing the different RFLP types confirmed the presence of novel Finnish forest soil Crenarchaeota. Archaeal sequences were identified from mycorrhizas of both P. involutus and S. bovinus, at the margins of the external mycelium and in uncolonised humus but not from non-mycorrhizal short roots. Fungal and compartment-specific crenarchaeal occupation of mycorrhizospheres is discussed in relation to bacterial community distribution in similar systems. [Copyright &y& Elsevier]

It has only recently been discovered that naturally prevailing microorganisms have a notable role in flotation in addition to chemical process parameters and overall water quality. This study's aim was to assess the prevailing microbial communities in relation to process chemistry in a zinc and copper mineral flotation plant. Due to the limitations of cultivation-based microbial methods that detect only a fraction of the total microbial diversity, DNA-based methods were utilised. However, it was discovered that the DNA extraction methods need to be improved for these environments with high mineral particle content. Microbial communities and metabolism were studied with quantitative PCR and amplicon sequencing of bacterial, archaeal and fungal marker genes and shotgun sequencing. Bacteria dominated the microbial communities, but in addition, both archaea and fungi were present. The predominant bacterial metabolism included versatile sulfur compound oxidation. Putative Thiovirga sp. dominated in the zinc plant and the water circuit samples, whereas Thiobacillus spp. dominated the copper plant. Halothiobacillus spp. were also an apparent part of the community in all samples. Nitrogen metabolism was more related to assimilatory than dissimilatory nitrate and nitrite oxidation/reduction reactions. Abundance of heavy metal resistance genes emphasized the adaptation and competitive edge of the core microbiome in these extreme conditions compared to microorganisms freshly entering the process. [ABSTRACT FROM AUTHOR]

Crystalline bedrock has been chosen for deep geologic long-term storage of used nuclear fuel in Finland. The risks generated by the deep subsurface microbial communities in these disposal sites need to be well characterised in advance to ensure safety. Deep subsurface microbial communities in a steady state are unlikely to contribute to known risk factors, such as corrosion or gas production. However, the construction of the geological final-disposal facility, bedrock disturbances, and hydraulic gradients cause changes that affect the microbial steady-state. To study the induced metabolism of deep microbial communities in changing environmental conditions, the activating effect of different electron donors and acceptors were measured with redox sensing fluorescent dyes (5-Cyano-2,3-ditolyl tetrazolium chloride, CTC and RedoxSensor™ Green, RSG). Fluids originating from two different fracture zones of the Finnish disposal site in Olkiluoto were studied. These fracture fluids were very dissimilar both chemically and in terms of bacterial and archaeal diversity. However, the microbial communities of both fracture fluids were activated, especially with acetate, which indicates the important role of acetate as a preferred electron donor for Olkiluoto deep subsurface communities. [ABSTRACT FROM AUTHOR]