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Workreport 2017-11



Metabolic Pathways of Deep Groundwater Microbiomes and Sulphide Formation at Olkiluoto


Blomberg, P., Itävaara, M., Marjamaa, K., Salavirta, H., Arvas, M., Miettinen, H., Vikman, M.



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This work is connected to Olkiluoto microbiological site characterization studies at the final disposal site of high radioactive wastes. The aim was to discover the mechanisms of sulphide-formation in deep subsurface groundwaters in Olkiluoto.

Geomicrobes have an important role in the transformation of minerals, in the decomposition and biosynthesis of organic compounds, and in the chemical changes of the groundwater composition. Microbial metabolic pathways and genes connected to these processes can therefore be used to monitor geobiological cycles. In order to study the sulphide-formation in two groundwater samples (OL-KR6/125−130 m with high sulphate concentration and no sulphides and OL-KR13/404.5−405.5 m with low sulphate concentration but high sulphide concentration), metagenomic and metatranscriptomic analyses were performed. Microbial diversity analyses were performed to tentatively relate the geomicrobiological processes and metabolic pathways with the organisms catalysing them.

The deep groundwater microbial biomass was filtered, DNA and mRNA extracted and sequenced. Proteins were predicted and metabolic pathways were studied based on the comparison of expressed genes (metatrancriptomes) to genes (metagenomes). The metabolism of the multispecies microbial communities, here called microbiomes, were very complex. All species in the microbiome were dependent on each other’s metabolism and on the prevailing physicochemical environmental conditions and available electron
acceptors and donors. Microbial metabolic pathways associated with geobiological cycles were studied. The analyses focused on the metabolism of nitrogen, sulphur, iron and carbon (methane and CO2).

Sulphate-reduction occurred by several routes. The metabolic pathways for both assimilatory and dissimilatory sulphate-reduction were active in both samples. Although several sulphate-reducing organisms were present in both samples, bacteria typically associated with the oxidation of sulphur compounds were most abundant. These bacteria may have been catalysing the oxidation of hydrogen sulphide to zero-valent sulphur, e.g. intracellular polysulphide, or catalysing the net disproportionation of zero-valent sulphur to sulphate and hydrogen sulphide.

Nitrogen and sulphur cycles connect to each other in several ways. E.g. nitrite as an electron acceptor can compete with sulphate. Furthermore, some of the enzymes in nitrogen metabolism were structurally and/or functionally similar to some of the enzymes in sulphur metabolism. Nitrogen fixation was the most active metabolic route for nitrogen utilization, transforming molecular nitrogen into ammonium. Although nitrification was not detected, both samples revealed active pathways for the consumption of nitrate and

Sulphur-reduction requires an electron donor such as hydrogen gas, methane, or organic carbon originating from dead biomass, all of which were deemed probable in these two samples.

The difference in sulphide accumulation into groundwater in the studied samples may be a result of the following observations. The archaea in OL-KR13/404.5−405.5 m were dominated by methanogens (methane-producing organisms), while the archaea in OLKR6/125−130 m were dominated by methanotrophs (methane-consuming organisms), although methanogens were also present. OL-KR6/125−130 m exclusively contained a large amount of direct methane-oxidizing organisms, thus indicating access to a yet unidentified electron acceptor. OL-KR13/404.5−405.5 m contained a larger fraction of anaerobic methane oxidizers utilizing reverse methanogenesis for the consumption of
methane. ANME-2 archaea and syntrophic bacteria were found abundantly in both samples, but more abundantly in OL-KR13/404.5−405.5 m. The OL-KR6/125−130 m groundwater contained a significant amount of soluble iron(II), which would have precipitated any free sulphide, thus removing it from the groundwater.

As a conclusion; Metabolic pathways connected to anaerobic methane oxidation, reverse methanogenesis in particular, were abundant in both samples, which may indicate a role for methane as a carbon source and an electron donor for the reduction of sulphate to sulphide in Olkiluoto groundwater. This finding is not lessened or contradicted by methane also being produced in these microbial communities.


Sulphate reduction, sulphide formation, deep groundwater, geomicrobiology, metabolic pathways, metagenomics, metatranscriptomics


WR 2017-11_web (pdf) (6.6 MB)


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