Taiwan is situated in the subtropical region and its geographical location and topographical features contribute to a rich ecological diversity and scenic landscapes. We investigated the diversity of methanogens in different environments of Taiwan using a culture-dependent method. This report presents the characterization and taxonomy of six hydrogenotrophic methanogens obtained from cold seep sediments (strain FWC-SCC1T and FWC-SCC3T), marine sediments (strain CWC-02T and YWC-01T), estuarine sediments (strain Afa-1T), and a hot spring well (strain Wushi-C6T) in Taiwan. The proposed names of the six novel species are Methanoculleus frigidifontis (type strain FWC-SCC1T=BCRC AR10056T=NBRC 113993T), Methanoculleus oceani (CWC-02T=BCRC AR10055T=NBRC 113992T), Methanoculleus methanifontis (FWC-SCC3T=BCRC AR10057T=NBRC 113994T), Methanoculleus nereidis (YWC-01T=BCRC AR10060T=NBRC 114597T), Methanoculleus formosensis (Afa-1T=BCRC AR10054T=NBRC 113995T), and Methanoculleus caldifontis (Wushi-06T=BCRC AR10059T= NBRC 114596T).

Wetwood of living trees is a habitat of methanogenic archaea, but the ubiquity of methanogenic archaea in the trunk of various trees has not been revealed. The present study analysed methanogenic archaeal communities inside coniferous and broadleaved trees in a cold temperate mountain forest by culture-dependent or independent techniques. Heartwood and sapwood segments were obtained from the trunk of seven tree species, Cryptomeria japonica, Quercus crispula, Fraxinus mandshurica, Acer pictum, Aesculus turbinata, Magnolia obovata, and Populus tremula. Amplicon sequencing analysis of 16S rRNA genes showed that Methanobacteriaceae predominated the archaeal communities and Methanomassiliicoccaceae also inhabited some trees. Real-time PCR analysis detected methanogenic archaeal mcrA genes from all the tree species, with a maximum of 107 copies g-1 dry wood. Digital PCR analysis also detected mcrA genes derived from Methanobacterium spp. and Methanobrevibacter spp. from several samples, with a maximum of 105 and 104 copies g-1 dry wood. The enumeration by the most probable number method demonstrated the inhabitation of viable methanogenic archaea inside the trees; 106 cells g-1 dry wood was enumerated from a heartwood sample of C. japonica. Methanogenic archaea related to Methanobacterium beijingense were cultivated from a heartwood sample of Q. crispula and F. mandshurica. The present study demonstrated that the inside of various trees is a common habitat for methanogenic archaeal communities and a potential source of methane in forest ecosystems.

Surveying bacterial and archaeal microbial communities in host and environmental studies requires the collection and storage of samples. Many studies are conducted in distant locations challenging these prerequisites. The use of preserving buffers is an important alternative when lacking access to cryopreservation, however, its effectivity for samples with challenging chemistry or samples that provide opportunities for fast bacterial or archaeal growth upon exposure to an aerobic environment, like peat samples, requires methodological assessment. Here, in combination with an identified optimal DNA extraction kit for peat soil samples, we test the application of several commercial and a homemade preservation buffer and make recommendations on the method that can most effectively preserve a microbiome reflective of the original state. In treatments with a non-optimal buffer or in the absence, we observed notable community shifts beginning as early as three days post-preservation lowering diversity and community evenness, with growth-driven artifacts from a few specific phyla. However other buffers retain a very close composition relative to the original state, and we described several metrics to understand some variation across them. Due to the chemical effects of preservation buffers, it is critical to test their compatibility and reliability to preserve the original bacterial and archaeal community in different environments.

Ciliates are a diverse group of protists known for their ability to establish various partnerships and thrive in a wide variety of oxygen-depleted environments. Most anaerobic ciliates harbor methanogens, one of the few known archaea living intracellularly. These methanogens increase the metabolic efficiency of host fermentation via syntrophic use of host end-product in methanogenesis. Despite the ubiquity of these symbioses in anoxic habitats, patterns of symbiont specificity and fidelity are not well known. We surveyed two unrelated, commonly found groups of anaerobic ciliates, the Plagiopylea and Metopida, isolated from anoxic marine sediments. We sequenced host 18S rRNA and symbiont 16S rRNA marker genes as well as the symbiont internal transcribed spacer region from our cultured ciliates to identify hosts and their associated methanogenic symbionts. We found that marine ciliates from both of these co-occurring, divergent groups harbor closely related yet distinct intracellular archaea within the Methanocorpusculum genus. The symbionts appear to be stable at the host species level, but at higher taxonomic levels, there is evidence that symbiont replacements have occurred. Gaining insight into this unique association will deepen our understanding of the complex transmission modes of marine microbial symbionts, and the mutualistic microbial interactions occurring across domains of life.

An anaerobic, mesophilic, syntrophic, archaeon strain MK-D1T, was isolated as a pure co-culture with Methanogenium sp. strain MK-MG from deep-sea methane seep sediment. This organism is, to our knowledge, the first cultured representative of \'Asgard\' archaea, an archaeal group closely related to eukaryotes. Here, we describe the detailed physiology and phylogeny of MK-D1T and propose Promethearchaeum syntrophicum gen. nov., sp. nov. to accommodate this strain. Cells were non-motile, small cocci, approximately 300-750 nm in diameter and produced membrane vesicles, chains of blebs and membrane-based protrusions. MK-D1T grew at 4-30 °C with optimum growth at 20 °C. The strain grew chemoorganotrophically with amino acids, peptides and yeast extract with obligate dependence on syntrophy with H2-/formate-utilizing organisms. MK-D1T showed the fastest growth and highest maximum cell yield when grown with yeast extract as the substrate: approximately 3 months to full growth, reaching up to 6.7×106 16S rRNA gene copies ml-1. MK-D1T had a circular 4.32 Mb chromosome with a DNA G+C content of 31.1 mol%. The results of phylogenetic analyses of the 16S rRNA gene and conserved marker proteins indicated that the strain is affiliated with \'Asgard\' archaea and more specifically DHVC1/DSAG/MBG-B and \'Lokiarchaeota\'/\'Lokiarchaeia\'. On the basis of the results of 16S rRNA gene sequence analysis, the most closely related isolated relatives were Infirmifilum lucidum 3507LTT (76.09 %) and Methanothermobacter tenebrarum RMAST (77.45 %) and the closest relative in enrichment culture was Candidatus \'Lokiarchaeum ossiferum\' (95.39 %). The type strain of the type species is MK-D1T (JCM 39240T and JAMSTEC no. 115508). We propose the associated family, order, class, phylum, and kingdom as Promethearchaeaceae fam. nov., Promethearchaeales ord. nov., Promethearchaeia class. nov., Promethearchaeota phyl. nov., and Promethearchaeati regn. nov., respectively. These are in accordance with ICNP Rules 8 and 22 for nomenclature, Rule 30(3)(b) for validation and maintenance of the type strain, and Rule 31a for description as a member of an unambiguous syntrophic association.

Asgardarchaeota, commonly referred to as Asgard archaea, is a candidatus phylum-rank archaeal clade that includes the closest archaeal relatives of eukaryotes. Despite their prevalence in the scientific literature, the name Asgardarchaeota lacks nomenclatural validation. Here, we describe a novel high-quality metagenome-assembled genome (MAG), AB3033_2TS, proposed to serve as the nomenclatural type for the species Asgardarchaeum abyssiTS according to the rules of the SeqCode. Based on protein content and compositional features, we infer that A. abyssi AB3033_2TS is an acetogenic chemoheterotroph, possibly a facultative lithoautotroph, and is adapted to a thermophilic lifestyle. Utilizing genomes from Asgard archaea, TACK, and Euryarchaea, we perform phylogenomic reconstructions using the GTDB archaeal marker genes, the current reference set for taxonomic classification. Calibrating relative evolutionary divergence (RED) values for Asgardarchaeota using established Thermoproteota lineages in the GTDB r207 reference tree, we establish a robust classification and propose Asgardarchaeum as the type genus for the family Asgardarchaeaceae (fam. nov)., the order Asgardarchaeales (ord. nov.), the class Asgardarchaeia (class. nov.), and the phylum Asgardarchaeota (phyl. nov.). This effort aims to preserve taxonomic congruence in the scientific literature.

Haloferax and Halobellus are the representatives of the family Haloferacaceae and they are dominant in hypersaline ecosystems. Some Haloferax and Halobellus species exhibit a close evolutionary relationship. Genomic, phylogenetic (based on 16S rRNA gene sequence), and phylogenomic analysis were performed to evaluate the taxonomic positions of the genera Haloferax and Halobellus. Based on the results we propose to reclassify Halobellus ramosii as a later heterotypic synonym of Halobellus inordinatus; Haloferax lucentense and Haloferax alexandrinum as later heterotypic synonyms of Haloferax volcanii.

Four halophilic archaeal strains, BCD28T, BND7T, PSR21T, and PSRA2T, were isolated from coastal and inland saline soil, respectively. The 16S rRNA and rpoB\' gene sequence similarities among these four strains and current species of Halomarina were 95.9-96.6% and 86.9-90.3%, respectively. Phylogenetic and phylogenomic analyses revealed that these four strains tightly cluster with the current species of the genus Halomarina. The AAI, ANI, and dDDH values among these four strains and current species of Halomarina were 65.3-68.4%, 75.8-77.7%, and 20.3-22.0%, respectively, clearly below the threshold values for species demarcation. Strains BCD28T, BND7T, PSR21T, and PSRA2T could be differentiated from the current species of Halomarina based on the comparison of diverse phenotypic characteristics. The major polar lipids of these four strains were phosphatidylglycerol (PG), phosphatidylglycerol phosphate methyl ester (PGP-Me), and four to five glycolipids. Phosphatidylglycerol sulfate (PGS) was only detected in strain BND7T. The phenotypic, phylogenetic, and genome-based analyses suggested that strains BCD28T (= CGMCC 1.18776T = JCM 34908T), BND7T (= CGMCC 1.18778T = JCM 34910T), PSR21T (= CGMCC 1.17027T = JCM 34147T), and PSRA2T (= CGMCC 1.17214T = JCM 34148T) represent four novel species of the genus Halomarina, for which the names Halomarina litorea sp. nov., Halomarina pelagica sp. nov., Halomarina halobia sp. nov., and Halomarina ordinaria sp. nov. are proposed.

Use of curldlan, an insoluble β-1,3-glucan, as an enrichment substrate under aerobic conditions resulted in the selection from hypersaline soda lakes of a single natronarchaeon, strain AArc-curdl1. This organism is an obligately aerobic saccharolytic, possessing a poorly explored (in Archaea) potential to utilize beta-1-3 glucans, being only a second example of a haloarchaeon with this ability known in pure culture. The main phenotypic property of the isolate is the ability to grow with insoluble β-1,3-backboned glucans, i.e. curdlan and pachyman. Furthermore, the strain utilized starch family α-glucans, beta-fructan inulin and a limited spectrum of sugars. The major ether-bound membrane polar phospholipids included PGP-Me and PG. The glyco- and sulfolipids were absent. The major respiratory menaquinone is MK-8:8. According to phylogenomic analysis, AArc-curdl1 represents a separate species in the recently described genus Natronosalvus within the family Natrialbaceae. The closest related species is Natronosalvus amylolyticus (ANI, AAI and DDH values of 90.2, 91.6 and 44 %, respectively). On the basis of its unique physiological properties and phylogenomic distance, strain AArc-curdl1T is classified as a novel species Natronosalvus hydrolyticus sp. nov. (=JCM 34865 = UQM 41566).

BACKGROUND: Saline lakes are home to various archaea that play special and crucial roles in the global biogeochemical cycle. The Qinghai-Tibet Plateau hosts a large number of lakes with diverse salinity ranging from 0.1 to over 400 g/L, harboring complex and diverse archaea. To the best of our knowledge, the formation mechanisms and potential ecological roles of archaea in Qinghai-Tibetan Plateau saline lakes remain largely unknown.
RESULTS: Using High-throughput Illumina sequencing, we uncovered the vastly distinct archaea communities between two typical saline lakes with significant salinity differences on the Qinghai Tibet Plateau (Qinghai saline lake and Chaka hypersaline lake) and suggested archaea played different important roles in methanogenesis-related and nitrate reduction-related functions of these two lakes, respectively. Rather than the individual effect of salinity, the composite effect of salinity with diverse environmental parameters (e.g., temperature, chlorophyll a, total nitrogen, and total phosphorus) dominated the explanation of the variations in archaeal community structure in different habitats. Based on the network analysis, we further found the correlations between dominant archaeal OTUs were tight but significantly different between the two habitats, implying that archaeal interactions may also largely determine the shape of archaeal communities.
CONCLUSIONS: The present study improved our understanding of the structure and function of archaea in different saline lakes on the Qinghai-Tibet Plateau and provided a new perspective on the mechanisms underlying shaping their communities.