Methanogenic archaea are major players in the global carbon cycle and in the biotechnology of anaerobic digestion. The phylum Euryarchaeota includes diverse groups of methanogens that are interspersed with non-methanogenic lineages. So far, methanogens inhabiting hypersaline environments have been identified only within the order Methanosarcinales. We report the discovery of a deep phylogenetic lineage of extremophilic methanogens in hypersaline lakes and present analysis of two nearly complete genomes from this group. Within the phylum Euryarchaeota, these isolates form a separate, class-level lineage ‘Methanonatronarchaeia’ that is most closely related to the class Halobacteria. Similar to the Halobacteria, ‘Methanonatronarchaeia’ are extremely halophilic and do not accumulate organic osmoprotectants. The high intracellular concentration of potassium implies that ‘Methanonatronarchaeia’ employ the ‘salt-in’ osmoprotection strategy. These methanogens are heterotrophic methyl-reducers that use C₁-methylated compounds as electron acceptors and formate or hydrogen as electron donors. The genomes contain an incomplete and apparently inactivated set of genes encoding the upper branch of methyl group oxidation to CO₂ as well as membrane-bound heterodisulfide reductase and cytochromes. These features differentiate ‘Methanonatronarchaeia’ from all known methyl-reducing methanogens. The discovery of extremely halophilic, methyl-reducing methanogens related to haloarchaea provides insights into the origin of methanogenesis and shows that the strategies employed by methanogens to thrive in salt-saturating conditions are not limited to the classical methylotrophic pathway.

Hypersaline soda lakes are characterized by extreme high soluble carbonate alkalinity. Despite the high pH and salt content, highly diverse microbial communities are known to be present in soda lake brines but the microbiome of soda lake sediments received much less attention of microbiologists. Here, we performed metagenomic sequencing on soda lake sediments to give the first extensive overview of the taxonomic diversity found in these complex, extreme environments and to gain novel physiological insights into the most abundant, uncultured prokaryote lineages. We sequenced five metagenomes obtained from four surface sediments of Siberian soda lakes with a pH 10 and a salt content between 70 and 400 g L−1. The recovered 16S rRNA gene sequences were mostly from Bacteria, even in the salt-saturated lakes. Most OTUs were assigned to uncultured families. We reconstructed 871 metagenome-assembled genomes (MAGs) spanning more than 45 phyla and discovered the first extremophilic members of the Candidate Phyla Radiation (CPR). Five new species of CPR were among the most dominant community members. Novel dominant lineages were found within previously well-characterized functional groups involved in carbon, sulfur, and nitrogen cycling. Moreover, key enzymes of the Wood-Ljungdahl pathway were encoded within at least four bacterial phyla never previously associated with this ancient anaerobic pathway for carbon fixation and dissimilation, including the Actinobacteria. Our first sequencing effort of hypersaline soda lake sediment metagenomes led to two important advances. First, we showed the existence and obtained the first genomes of haloalkaliphilic members of the CPR and several hundred other novel prokaryote lineages. The soda lake CPR is a functionally diverse group, but the most abundant organisms in this study are likely fermenters with a possible role in primary carbon degradation. Second, we found evidence for the presence of the Wood-Ljungdahl pathway in many more taxonomic groups than those encompassing known homo-acetogens, sulfate-reducers, and methanogens. Since only few environmental metagenomics studies have targeted sediment microbial communities and never to this extent, we expect that our findings are relevant not only for the understanding of haloalkaline environments but can also be used to set targets for future studies on marine and freshwater sediments.

A novel nitrite-oxidizing bacterium (NOB), strain Lb^T, was isolated from a nitrifying bioreactor with a high loading of ammonium bicarbonate in a mineral medium with nitrite as the energy source. The cells were oval (lancet-shaped) rods with pointed edges, non-motile, Gram-positive (by staining and from the cell wall structure) and non-spore-forming. Strain Lb^Twas an obligately aerobic, chemolitoautotrophic NOB, utilizing nitrite or formate as the energy source and CO₂as the carbon source. Ammonium served as the only source of assimilated nitrogen. Growth with nitrite was optimal at pH 6.8–7.5 and at 40 °C (maximum 46 °C). The membrane lipids consisted of C₂₀alkyl 1,2-diols with the dominant fatty acids being 10MeC₁₈and C_{18 : 1ω9}. The peptidoglycan lackedmeso-DAP but contained ornithine and lysine. The dominant lipoquinone was MK-8. Phylogenetic analyses of the 16s rRNA gene sequence placed strain Lb^Tinto the class Thermomicrobia of the phylum Chloroflexi with Sphaerobacter thermophilus as the closest relative. On the basis of physiological and phylogenetic data, it is proposed that strain Lb^Trepresents a novel species of a new genus, with the suggested nameNitrolancea hollandicagen. nov., sp. nov. The type strain of the type species is Lb^T( = DSM 23161^T = UNIQEM U798^T).

Significance: We report on cultivation and characterization of an association between Candidatus Nanohalobium constans and its host, the chitinotrophic haloarchaeon Halomicrobium LC1Hm, obtained from a crystallizer pond of marine solar salterns. High-quality nanohaloarchael genome sequence in conjunction with electron- and fluorescence microscopy, growth analysis, and proteomic and metabolomic data revealed mutually beneficial interactions between two archaea, and allowed dissection of the mechanisms for these interactions. Owing to their ubiquity in hypersaline environments, Nanohaloarchaeota may play a role in carbon turnover and ecosystem functioning, yet insights into the nature of this have been lacking. Here, we provide evidence that nanohaloarchaea can expand the range of available substrates for the haloarchaeon, suggesting that the ectosymbiont increases the metabolic capacity of the host.

Methanogenic enrichments from hypersaline lakes at moderate thermophilic conditions have resulted in the cultivation of an unknown deep lineage of euryarchaeota related to the class Halobacteria . Eleven soda lake isolates and three salt lake enrichment cultures were methyl-reducing methanogens that utilize C1 methylated compounds as electron acceptors and H2 or formate as electron donors, but they were unable to grow on either substrates alone or to form methane from acetate. They are extreme halophiles, growing optimally at 4 M total Na+ and the first representatives of methanogens employing the ‘salt-in’ osmoprotective mechanism. The salt lake subgroup is neutrophilic, whereas the soda lake isolates are obligate alkaliphiles, with an optimum around pH 9.5. Both grow optimally at 50 °C. The genetic diversity inside the two subgroups is very low, indicating that the soda and salt lake clusters consist of a single genetic species each. The phylogenetic distance between the two subgroups is in the range of distant genera, whereas the distance to other euryarchaea is below 83 % identity of the 16S rRNA gene. These isolates and enriched methanogens, together with closely related environmental clones from hypersaline habitats (the SA1 group), form a novel class-level clade in the phylum Euryarchaeota. On the basis of distinct phenotypic and genetic properties, the soda lake isolates are classified into a new genus and species, Methanonatronarchaeum thermophilum, with the type strain AMET1T (DSM 28684T=NBRC 110805T=UNIQEM U982T), and the salt lake methanogens into a candidate genus and species ‘Candidatus Methanohalarchaeum thermophilum’. These organisms are proposed to form novel family, order and class Methanonatronarchaeaceae fam. nov., Methanonatronarchaeales ord. nov. and Methanonatronarchaeia classis nov., within the phylum Euryarchaeota .