Metabolic potential and survival strategies in MAGs
In general, the anvi’o pipeline using co-assembly showed the best binning results for our eighteen metagenomes, generating a total of 158 MAGs. We included in our analyses only 1 MAG produced through the idba-ud assembler and MaxBin binning since this MAG belonged to a taxon (Calditrichia) which was not achieved through the anvi’o pipeline (Supplementary Figure 3, Supplementary Table 4). From the 159 MAGs, 12 were assigned as Archaea through GTDB-Tk and GhostKoala, belonging to Nitrososphaerales (Candidatus Nitrosocaldus according with GhostKoala taxonomy) (2), Nitrosoarchaeum (1),Nitrosotenius (1), Nitrospumilus (1), Desulfurococcales (Aeropyrum according with GhostKoala taxonomy) (1), Acidilobaceae (1), Pyrodictiaceae (2) and Woesearchaeia (Nanoarchaeota) (3). The bacterial MAGs were classified through GTDB-Tk and GhostKoala as the following phyla: Acidobacteriota (1), Aquificota (2), Bacteroidota (92), Calditrichota (5), Campylobacterota (1), Chloroflexota (3), Cyanobacteriota (1), Firmicutes (1), Nitrospirota (2), Patescibacteria (4) and Proteobacteria (35) (Supplementary Figure 4, Supplementary Table 4). A total of 13 MAGs were considered as high quality and 82 as medium quality drafts.
The MAGs were selected for functional annotation by their quality and based on groups related to extremophiles and associated to sulfur and nitrogen metabolisms. These 11 selected MAGs were assigned as DI_MAG_00003 (Sulfurimonas ), DI_MAG_00004 (Hydrogenothermaceae/Persephonella ), DI_MAG_00006 (Promineofilaceae/Candidatus Promineofilum ), DI_MAG_00010 (Caldilineaceae/Caldilinea ), DI_MAG_00011 (Thermonemataceae), DI_MAG_00019 (Chitinophagaceae), DI_MAG_00020 (Pyrodictiaceae/Pyrodictium ), DI_MAG_00021 (Dojkabacteria), DI_MAG_00022 (Woesearchaeia/archaeon GW2011_AR20), DI_MAG_00049 (Nitrososphaerales/Candidatus Nitrosocaldus ) and DI_MAG_FBB2_12 (Calditrichia) (Table 1).
We identified in the high-quality DI_MAG_00004 (Hydrogenothermaceae/Persephonella , ~ 97% completeness) genes for nitrate reduction (narGHI andnirA ), denitrification (narGHI ), nitrification (narGH ), sulfate reduction (sat , cysH , sir ), sulfur and thiosulfate oxidation (soxAXBYZ ), and incomplete pathways for carbon fixation (Figure 6a). This MAG had several genes associated with stress response, especially oxidative stress (e.g. superoxide reductase and dismutase, rubrerythrin and rubredoxin) and thermal response (e.g. groES , hsp20 and hspR ), as different DNA repair mechanisms, including photolyase repair (Figure 6b). In general, genes involved with the nitrogen cycle were identified in almost all selected MAGs, except for MAGs DI_MAG_00020, DI_MAG_00021, and DI_MAG_00022. Sulfate reduction genes were also detected in different selected MAGs, except for MAGs DI_MAG_00020, DI_MAG_00021, DI_MAG_00022 and DI_MAG_FBB2_12. All MAGs had incomplete pathways for carbon fixation, except for DI_MAG_00004 and DI_MAG_00021 (Figure 6a).
Different cold-shock genes were detected among MAGs; DI_MAG_00006 was the one which presented more csp genes. We did not find anycsp genes in archaeal MAGs (DI_MAG_00020, DI_MAG_00022, and DI_MAG_00049). However, we observed genes in all selected MAGs that were related to different heat-shock responses, includinggroEL/groES genes in DI_MAG_00004, DI_MAG_00020, and DI_MAG_00021. Thermosome (thsA ) and reverse gyrase genes were identified in all the selected MAGs assigned as Archaea (DI_MAG_00020, DI_MAG_00022, and DI_MAG_00049). Although all MAGs showed the potential presence of oxidative stress response (except DI_MAG_00049), rubrerythrin and rubredoxin genes were only observed in DI_MAG_00004 and DI_MAG_00003. Different DNA repair mechanisms were identified in selected MAGs, such as several recombination genes (rec genes), DNA mismatch repair (mut genes), nucleotide excision repair (uvr genes), double-strand break repair (her A, only in archaeal-selected MAGs) and photolyase repair (only in DI_MAG_00004) (Figure 6b).