Rokubacteria is a phylogenetic clade of as-yet-uncultivated prokaryotes, which are detected in diverse terrestrial habitats and are commonly addressed as members of the rare biosphere. This clade was originally described as a candidate phylum; however, based on the results of comparative genome analysis, was later defined as the order-level lineage, Rokubacteriales, within the phylum Methylomirabilota. The physiology and lifestyles of these bacteria are poorly understood. A dataset of 16S rRNA gene reads retrieved from four boreal raised bogs and six eutrophic fens was examined for the presence of the Rokubacteriales; the latter were detected exclusively in fens. Their relative abundance varied between 0.2 and 4% of all bacteria and was positively correlated with pH, total nitrogen content, and availability of Ca and Mg. To test an earlier published hypothesis regarding the presence of methanotrophic capabilities in Rokubacteria, peat samples were incubated with 10% methane for four weeks. No response to methane availability was detected for the Rokubacteriales, while clear a increase in relative abundance was observed for the conventional Methylococcales methanotrophs. The search for methane monooxygenase encoding genes in 60 currently available Rokubacteriales metagenomes yielded negative results, although copper-containing monooxygenases were encoded by some members of this order. This study suggests that peat-inhabiting Rokubacteriales are neutrophilic non-methanotrophic bacteria that colonize nitrogen-rich wetlands.

Large areas in Northern Russia are covered by extensive mires, which represent a complex mosaic of ombrotrophic raised bogs, minerotrophic and eutrophic fens, all in a close proximity to each other. In this paper, we compared microbial diversity patterns in the surface peat layers of the neighbouring raised bogs and eutrophic fens that are located within two geographically remote mire sites in Vologda region using 16S rRNA gene sequencing. Regardless of location, the microbial communities in raised bogs were highly similar to each other but were clearly distinct from those in eutrophic fens. Bogs were dominated by the Acidobacteria (30%–40% of total 16S rRNA gene reads), which belong to the orders Acidobacteriales and Bryobacterales. Other bog-specific bacteria included the Phycisphaera-like group WD2101 and the families Isosphaeraceae and Gemmataceae of the Planctomycetes, orders Opitutales and Pedosphaerales of the Verrucomicrobia and a particular group of alphaproteobacteria within the Rhizobiales. In contrast, fens hosted Anaerolineae-affiliated Chloroflexi, Vicinamibacteria- and Blastocatellia-affiliated Acidobacteria, Rokubacteria, uncultivated group OM190 of the Planctomycetes and several groups of betaproteobacteria. The Patescibacteria were detected in both types of wetlands but their relative abundance was higher in fens. A number of key parameters that define the distribution of particular bacterial groups in mires were identified.

Methanotrophic verrucomicrobia of the order Methylacidiphilales are known as extremely acidophilic, thermophilic or mesophilic bacteria that inhabit acidic geothermal ecosystems. The occurrence of verrucomicrobial methanotrophs in other types of acidic environments remains an open question. Notably, Methylacidiphilales-affiliated 16S rRNA gene sequences are commonly retrieved from acidic (pH 3.5–5.5) peatlands. In this study, we compared the patterns of verrucomicrobial diversity in four acidic raised bogs and six neutral fens located in European North Russia. Methylacidiphilales-like 16S rRNA gene reads displaying 83–86% similarity to 16S rRNA gene sequences of currently described verrucomicrobial methanotrophs were recovered exclusively from raised bogs. Laboratory incubation of peat samples with 10% methane for 3 weeks resulted in the pronounced increase of a relative abundance of alphaproteobacterial methanotrophs, while no response was detected for Methylacidiphilales-affiliated bacteria. Three metagenome-assembled genomes (MAGs) of peat-inhabiting Methylacidiphilales bacteria were reconstructed and examined for the presence of genes encoding methane monooxygenase enzymes and autotrophic carbon fixation pathways. None of these genomic determinants were detected in assembled MAGs. Metabolic reconstructions predicted a heterotrophic metabolism, with a potential to hydrolyze several plant-derived polysaccharides. As suggested by our analysis, peat-inhabiting representatives of the Methylacidiphilales are acidophilic aerobic heterotrophs, which comprise a sister family of the methanotrophic Methylacidiphilaceae.

Upland soils of tundra function as a constant sink for atmospheric CH₄ but the identity of methane oxidizers in these soils remains poorly understood. Methane uptake rates of −0.4 to −0.6 mg CH₄-C m⁻² day⁻¹ were determined by the static chamber method in a mildly acidic upland soil of the lichen-dominated forested tundra, North Siberia, Russia. The maximal CH₄ oxidation activity was localized in an organic surface soil layer underlying the lichen cover. Molecular identification of methanotrophic bacteria based on retrieval of the pmoA gene revealed Upland Soil Cluster Alpha (USCα) as the only detectable methanotroph group. Quantification of these pmoA gene fragments by means of specific qPCR assay detected ~10⁷pmoA gene copies g⁻¹ dry soil. The pmoA diversity was represented by seven closely related phylotypes; the most abundant phylotype displayed 97.5% identity to pmoA of Candidatus Methyloaffinis lahnbergensis. Further analysis of prokaryote diversity in this soil did not reveal 16S rRNA gene fragments from well-studied methanotrophs of the order Methylococcales and the family Methylocystaceae. The largest group of reads (~4% of all bacterial 16S rRNA gene fragments) that could potentially belong to methanotrophs was classified as uncultivated Beijerinckiaceae bacteria. These reads displayed 96–100 and 95–98% sequence similarity to 16S rRNA gene of Candidatus Methyloaffinis lahnbergensis and “Methylocapsa gorgona” MG08, respectively, and were represented by eight species-level operational taxonomic units (OTUs), two of which were highly abundant. These identification results characterize subarctic upland soils, which are exposed to atmospheric methane concentrations only, as a unique habitat colonized mostly by USCα methanotrophs.

Carotenoids are secondary metabolites that exhibit antioxidant properties and are characterized by a striking range of colorations from red to yellow. These natural pigments are synthesized by a wide range of eukaryotic and prokaryotic organisms. Among the latter, carotenoid-producing methanotrophic bacteria, which display fast growth on methane or natural gas, are of particular interest as potential producers of a feed protein enriched with carotenoids. Until recently, Methylomonas strain 16a and Methylomonas sp. ZR1 remained the only representatives of the genus for which detailed carotenoid profile was determined. In this study, we analyzed the genome sequences of five strains of Methylomonas species whose pigmentation varied from white and yellow to orange and red, and identified carotenoids produced by these bacteria. Carotenoids synthesized using four pigmented strains included C30 fraction, primarily composed of 4,4’-diaplycopene-4,4’-dioic acid and 4,4’-diaplycopenoic acid, as well as C40 fraction with the major compound represented by 1,1’-dihydroxy-3,4-didehydrolycopene. The genomes of studied Methylomonas strains varied in size between 4.59 and 5.45 Mb and contained 4201–4735 protein-coding genes. These genomes and 35 reference Methylomonas genomes available in the GenBank were examined for the presence of genes encoding carotenoid biosynthesis. Genomes of all pigmented Methylomonas strains harbored genes necessary for the synthesis of 4,4’-diaplycopene-4,4’-dioic acid. Non-pigmented “Methylomonas montana” MW1^T lacked the crtN gene required for carotenoid production. Nearly all strains possessed phytoene desaturases, which explained their ability to naturally synthesize lycopene. Thus, members of the genus Methylomonas can potentially be considered as producers of C30 and C40 carotenoids from methane.

The family Gemmataceae accomodates aerobic, chemoorganotrophic planctomycetes with large genome sizes, is mostly distributed in freshwater and terrestrial environments. However, these bacteria have recently also been found in locations relevant to human health. Since the antimicrobial resistance genes (AMR) from environmental resistome have the potential to be transferred to pathogens, it is essential to explore the resistant capabilities of environmental bacteria. In this study, the reconstruction of in silico resistome was performed for all nine available gemmata genomes. Furthermore, the genome of the newly isolated yet-undescribed strain G18 was sequenced and added to all analyses steps. Selected genomes were screened for the presence of mobile genetic elements. The flanking location of mobilizable genomic milieu around the AMR genes was of particular interest since such colocalization may appear to promote the horizontal gene transfer (HGT) events. Moreover the antibiotic susceptibility profile of six phylogenetically distinct strains of Gemmataceae planctomycetes was determined.