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Background: The current view suggests that in low-temperature acidic environments, archaea are significantly less abundant than bacteria. Thus, this study of the microbiome of Parys Mountain (Anglesey, UK) sheds light on the generality of this current assumption. Parys Mountain is a historically important copper mine and its acid mine drainage (AMD) water streams are characterised by constant moderate temperatures (8-18 °C), extremely low pH (1.7) and high concentrations of soluble iron and other metal cations., Results: Metagenomic and SSU rRNA amplicon sequencing of DNA from Parys Mountain revealed a significant proportion of archaea affiliated with Euryarchaeota, which accounted for ca. 67% of the community. Within this phylum, potentially new clades of Thermoplasmata were overrepresented (58%), with the most predominant group being "E-plasma", alongside low-abundant Cuniculiplasmataceae, 'Ca. Micrarchaeota' and 'Terrestrial Miscellaneous Euryarchaeal Group' (TMEG) archaea, which were phylogenetically close to Methanomassilicoccales and clustered with counterparts from acidic/moderately acidic settings. In the sediment, archaea and Thermoplasmata contributed the highest numbers in V3-V4 amplicon reads, in contrast with the water body community, where Proteobacteria, Nitrospirae, Acidobacteria and Actinobacteria outnumbered archaea. Cultivation efforts revealed the abundance of archaeal sequences closely related to Cuniculiplasma divulgatum in an enrichment culture established from the filterable fraction of the water sample. Enrichment cultures with unfiltered samples showed the presence of Ferrimicrobium acidiphilum, C. divulgatum, 'Ca. Mancarchaeum acidiphilum Mia14', 'Ca. Micrarchaeota'-related and diverse minor (< 2%) bacterial metagenomic reads., Conclusion: Contrary to expectation, our study showed a high abundance of archaea in this extremely acidic mine-impacted environment. Further, archaeal populations were dominated by one particular group, suggesting that they are functionally important. The prevalence of archaea over bacteria in these microbiomes and their spatial distribution patterns represents a novel and important advance in our understanding of acidophile ecology. We also demonstrated a procedure for the specific enrichment of cell wall-deficient members of the archaeal component of this community, although the large fraction of archaeal taxa remained unculturable. Lastly, we identified a separate clustering of globally occurring acidophilic members of TMEG that collectively belong to a distinct order within Thermoplasmata with yet unclear functional roles in the ecosystem.

The genus Carboxydocella forms a deeply branching family in the class Clostridia and is currently represented by three physiologically diverse species of thermophilic prokaryotes. The type strain of the type species, Carboxydocella thermautotrophica 41 T , is an obligate chemolithoautotroph growing exclusively by hydrogenogenic CO oxidation. Another strain, isolated from a hot spring at Uzon caldera, Kamchatka in the course of this work, is capable of coupling carboxydotrophy and dissimilatory reduction of Fe(III) from oxic and phyllosilicate minerals. The processes of carboxydotrophy and Fe(III) reduction appeared to be interdependent in this strain. The genomes of both isolates were sequenced, assembled into single chromosome sequences (for strain 41 T a plasmid sequence was also assembled) and analyzed. Genome analysis revealed that each of the two strains possessed six genes encoding diverse Ni,Fe-containing CO dehydrogenases (maximum reported in complete prokaryotic genomes), indicating crucial role of carbon monoxide in C. thermautotrophica metabolism. Both strains possessed a set of 30 multiheme c -type cytochromes, but only the newly isolated Fe-reducing strain 019 had one extra gene of a 17-heme cytochrome, which is proposed to represent a novel determinant of dissimilatory iron reduction in prokaryotes. Mössbauer studies revealed that strain 019 induced reductive transformation of the abundant ferric/ferrous-mica mineral glauconite to siderite during carboxydotrophic growth. Reconstruction of the C. thermautotrophica strains energy metabolism is the first comprehensive genome analysis of a representative of the deep phylogenetic branch Clostridia Incertae Sedis, family V. Our data provide insights into energy metabolism of C. thermautotrophica with an emphasis on its ecological implications.