Fungi have an important role in nutrient cycling in most ecosystems on Earth, yet their ecology and functionality in deep continental subsurface remain unknown. Here, we report the first observations of active fungal colonization of mica schist in the deep continental biosphere and the ability of deep subsurface fungi to attach to rock surfaces under in situ conditions in groundwater at 500 and 967 m depth in Precambrian bedrock. We present an in situ subsurface biofilm trap, designed to reveal sessile microbial communities on rock surface in deep continental groundwater, using Outokumpu Deep Drill Hole, in eastern Finland, as a test site. The observed fungal phyla in Outokumpu subsurface were Basidiomycota, Ascomycota, and Mortierellomycota. In addition, significant proportion of the community represented unclassified Fungi. Sessile fungal communities on mica schist surfaces differed from the planktic fungal communities. The main bacterial phyla were Firmicutes, Proteobacteria, and Actinobacteriota. Biofilm formation on rock surfaces is a slow process and our results indicate that fungal and bacterial communities dominate the early surface attachment process, when pristine mineral surfaces are exposed to deep subsurface ecosystems. Various fungi showed statistically significant cross-kingdom correlation with both thiosulfate and sulfate reducing bacteria, e.g., SRB2 with fungi Debaryomyces hansenii.

The deep terrestrial biosphere hosts vast sessile rock surface communities and biofilms, but thus far, mostly planktic communities have been studied. We enriched deep subsurface microbial communities on mica schist in microcosms containing bedrock groundwater from the depth of 500 m from Outokumpu, Finland. The biofilms were visualized using scanning electron microscopy, revealing numerous different microbial cell morphologies and attachment strategies on the mica schist surface, e.g., bacteria with outer membrane vesicle-like structures, hair-like extracellular extensions, and long tubular cell structures expanding over hundreds of micrometers over mica schist surfaces. Bacterial communities were analyzed with amplicon sequencing showing that Pseudomonas, Desulfosporosinus, Hydrogenophaga, and Brevundimonas genera dominated communities after 8–40 months of incubation. A total of 21 metagenome assembled genomes from sessile rock surface metagenomes identified genes involved in biofilm formation, as well as a wide variety of metabolic traits indicating a high degree of environmental adaptivity to oligotrophic environment and potential for shifting between multiple energy or carbon sources. In addition, we detected ubiquitous organic carbon oxidation and capacity for arsenate and selenate reduction within our rocky MAGs. Our results agree with the previously suggested interaction between the deep subsurface microbial communities and the rock surfaces, and that this interaction could be crucial for sustaining life in the harsh anoxic and oligotrophic deep subsurface of crystalline bedrock environment.

In Finland, the repositories for low and intermediate-level radioactive waste (LLW and ILW) will be situated at three different geographical locations in about 60 to 100 m deep granite bedrock where the waste and waste containers can be subjected to anoxic groundwater containing microbes. The composition of groundwater varies in terms of chemistry and microbial activity in different locations. In this study, groundwater from the three repository areas was analyzed in respect to chemistry and microbial community. Corrosion tendency of three steel grades, carbon steel AISI/SAE 1005 and stainless steels AISI 304 and 316L, was studied in these groundwater environments using electrochemical methods. As a reference, measurements were also performed in simulated groundwater without microbes. The measurements show that corrosivity of the water and thus the steels’ performance differs depending on water origin. In addition, the groundwater differed remarkably in their chemical composition as well as abundance and diversity of microbial community between the sites. Consequently, the local environment has to be considered when evaluating the long-term evolution of disposal concepts.

In Finland, the repositories for low- and intermediate-level waste (LLW and ILW) are situated in three different geographical locations in deep granite bedrock, where waste containers can be subjected to anoxic groundwater containing microbes. The composition of groundwater varies in terms of chemistry and microbial activity in different locations. In this study, groundwater from three repository areas was analyzed with regard to chemistry and microbial community. The corrosion tendency of three steel grades, carbon steel AISI/SAE 1005 and stainless steels AISI 304 and 316L, was studied in these groundwater environments using electrochemical methods. As a reference, measurements were also performed in simulated groundwater that did not contain microbes. The measurements show that the corrosivity of the water, and thus, the steels’ performance, differed depending on its location of origin. Additionally, the groundwater differed remarkably in their chemical composition as well as the abundance and diversity of the microbial community within them. Consequently, the local environment has to be considered when evaluating the long-term safety of the disposal of nuclear waste.