New knowledge about methanogenic and methanotrophic Archaea diversity and activity in bottom sediments is important for understanding of processes involved to the transportation of carbon and biogenic compounds from terrestrial ecosystems to aquatic ones, and their biogeochemical transformation. The bottom sediments of lakes and rivers are the hotspots of methane production, but composition and activity of methanogenic and methanotrophic microorganisms remains poorly described for the bottom sediments of the great Arctic rivers. In our study we collected the bottom sediments accumulating along the left and right banks of the Yenisei River in 18 sites located between 56.0ºN and 67.4ºN. All sediment samples were immediately frozen (-18oC) and kept frozen during transportation to laboratory. In parallel with sediment sampling we collected samples of dissolved greenhouse gases by head space method and further measured CO2 and CH4 concentrations and δ13С-CH4 and δ13С-CO2 (Picarro 2201-i, USA). Metagenomic studies were performed in the Core Centrum ‘Genomic Technologies, Proteomics and Cell Biology’ (All-Russia Research Institute for Agricultural Microbiology). DNA was isolated using the MN NucleoSpin Kit (MN, Germany). A Precellus 24 homogenizer (Bertin, USA) was used as a destructive mechanical action. The quality of DNA isolation was checked electrophoretically (1% agarose gel, Bio-Rad, USA) and by PCR (Bio-Rad T100 Thermal Cycler). DNA sequencing was performed via Illumina MiSEQ sequencing system (USA), with primers F515 (GTGCCAGCMGCCGCGTAA) and R806 (GGACTACVSGGGTATCTAAT) for V4 variable region of the 16s rDNA gene, according manufacturer`s manual. Sequence processing was performed in R, using followed libraries: phyloseq, dada2, ShorReads, Biostrings and ggplot2. In total 29 prokaryotes belonging to Achaea were identified in sediment samples including 15 metanogenic archaeons, 1 anaerobic methanotroph (Candidatus Methanoperedens), 5 prokaryotes of Thermoplasmata, 5 prokaryotes of phylum Thaumarchaeota, 1 prokaryote of Bathyarchaeia, 1 prokaryote of Woesearchaeia, and 1 prokaryote of Altiarchaeia. Methanogenic community structure of bottom sediments of the Yenisei River dominated by archaeons belonging to Methanosarcina, Methanosaeta and Methanoregula. The OTU abundance of these archaeons was larger in sediments collected between 56ºN and 61ºN. Along this channel segment the values of δ13С-CH4 in the dissolved methane has increased from -54 to -43‰ VPDB that indicated methylotrophic and acetoclastic methanogenesis. In the segment between 61ºN and 64ºN the OTU abundance of methanogenic Archaea decreased dramatically (5-190 times) which was accompanied by the sharp depletion of δ13С-CH4 up to -60 – -80 ‰ VPDB indicating the shift to hydrogenotrophic metabolic pathway of methane production. Also in this river area we observed increasing OTU abundance of anaerobic methanotrophs belonging to Candidatus Methanoperedens. Further North (64-67ºN) we observed enrichment of δ13С-CH4 and increasing in methanogenic community archaeons belonging to Methanosarcina and Methanoregula. Acknowledgments: this study was supported by the grant from the RFBR, the project № 18-05-60203_Arctica, and 19-05-50107.
