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Recent omics studies have provided invaluable insights into the metabolic potential, adaptation, and evolution of novel archaeal lineages from a variety of extreme environments. We utilized a genome-resolved metagenomic approach to recover eight medium- to high-quality metagenome-assembled genomes (MAGs) that likely represent a new order ("Candidatus Sysuiplasmatales") in the class Thermoplasmata from mine tailings and acid mine drainage (AMD) sediments sampled from two copper mines in South China. 16S rRNA gene-based analyses revealed a narrow habitat range for these uncultured archaea limited to AMD and hot spring-related environments. Metabolic reconstruction indicated a facultatively anaerobic heterotrophic lifestyle. This may allow the archaea to adapt to oxygen fluctuations and is thus in marked contrast to the majority of lineages in the domain Archaea, which typically show obligately anaerobic metabolisms. Notably, "Ca. Sysuiplasmatales" could conserve energy through degradation of fatty acids, amino acid metabolism, and oxidation of reduced inorganic sulfur compounds (RISCs), suggesting that they may contribute to acid generation in the extreme mine environments. Unlike the closely related orders Methanomassiliicoccales and "Candidatus Gimiplasmatales," "Ca. Sysuiplasmatales" lacks the capacity to perform methanogenesis and carbon fixation. Ancestral state reconstruction indicated that "Ca. Sysuiplasmatales," the closely related orders Methanomassiliicoccales and "Ca. Gimiplasmatales," and the orders SG8-5 and RBG-16-68-12 originated from a facultatively anaerobic ancestor capable of carbon fixation via the bacterial-type H4F Wood-Ljungdahl pathway (WLP). Their metabolic divergence might be attributed to different evolutionary paths. IMPORTANCE A wide array of archaea populate Earth's extreme environments; therefore, they may play important roles in mediating biogeochemical processes such as iron and sulfur cycling. However, our knowledge of archaeal biology and evolution is still limited, since the majority of the archaeal diversity is uncultured. For instance, most order-level lineages except Thermoplasmatales, Aciduliprofundales, and Methanomassiliicoccales within Thermoplasmata do not have cultured representatives. Here, we report the discovery and genomic characterization of a novel order, "Ca. Sysuiplasmatales," within Thermoplasmata in extremely acidic mine environments. "Ca. Sysuiplasmatales" are inferred to be facultatively anaerobic heterotrophs and likely contribute to acid generation through the oxidation of RISCs. The physiological divergence between "Ca. Sysuiplasmatales" and closely related Thermoplasmata lineages may be attributed to different evolutionary paths. These results expand our knowledge of archaea in the extreme mine ecosystem.

In previous publications, it was hypothesized that Micrarchaeota cells are covered by two individual membrane systems. This study proofs that at least the recently cultivated “Candidatus Micrarchaeum harzensis A_DKE” possesses an S-layer covering its cytoplasmic membrane. The potential S-layer protein was found to be among the proteins with the highest abundance in A_DKE and in silico characterization of its primary structure indicated homologies to other known S-layer proteins. Homologs of this protein were found in other Micrarchaeota genomes, which raises the question, whether the ability to form an S-layer is a common trait within this phylum. The S-layer protein seems to be glycosylated and the Micrarchaeum expresses genes for N-glycosylation under cultivation conditions, despite not being able to synthesize carbohydrates. Electron micrographs of freeze-etched samples of a previously described co-culture, containing Micrarchaeum A_DKE and a Thermoplasmatales member as its host organism, verified the hypothesis of an S-layer on the surface of A_DKE. Both organisms are clearly distinguishable by cell size, shape and surface structure.

Acidophilic archaea of the archaeal Richmond Mine acidophilic nanoorganisms (ARMAN) group from the uncultured candidate phylum “Candidatus Micrarchaeota” have small genomes and cell sizes and are known to be metabolically dependent and physically associated with their Thermoplasmatales hosts. However, phylogenetically diverse “Ca. Micrarchaeota” are widely distributed in various nonacidic environments, and it remains uncertain because of the lack of complete genomes whether they are also devoted to a partner-dependent lifestyle. Here, we obtained nine metagenome-assembled genomes of “Ca. Micrarchaeota” from the sediments of a meromictic freshwater lake, including a complete, closed 1.2 Mbp genome of “Ca. Micrarchaeota” Sv326, an archaeon phylogenetically distant from the ARMAN lineage. Genome analysis revealed that, contrary to ARMAN “Ca. Micrarchaeota,” the Sv326 archaeon has complete glycolytic pathways and ATP generation mechanisms in substrate phosphorylation reactions, the capacities to utilize some sugars and amino acids as substrates, and pathways for de novo nucleotide biosynthesis but lacked an aerobic respiratory chain. We suppose that Sv326 is a free-living scavenger rather than an obligate parasite/symbiont. Comparative analysis of “Ca. Micrarchaeota” genomes representing different order-level divisions indicated that evolution of the “Ca. Micrarchaeota” from a free-living “Candidatus Diapherotrites”-like ancestor involved losses of important metabolic pathways in different lineages and gains of specific functions in the course of adaptation to a partner-dependent lifestyle and specific environmental conditions. The ARMAN group represents the most pronounced case of genome reduction and gene loss, while the Sv326 lineage appeared to be rather close to the ancestral state of the “Ca. Micrarchaeota” in terms of metabolic potential. IMPORTANCE The recently described superphylum DPANN includes several phyla of uncultivated archaea with small cell sizes, reduced genomes, and limited metabolic capabilities. One of these phyla, “Ca. Micrarchaeota,” comprises an enigmatic group of archaea found in acid mine drainage environments, the archaeal Richmond Mine acidophilic nanoorganisms (ARMAN) group. Analysis of their reduced genomes revealed the absence of key metabolic pathways consistent with their partner-associated lifestyle, and physical associations of ARMAN cells with their hosts were documented. However, “Ca. Micrarchaeota” include several lineages besides the ARMAN group found in nonacidic environments, and none of them have been characterized. Here, we report a complete genome of “Ca. Micrarchaeota” from a non-ARMAN lineage. Analysis of this genome revealed the presence of metabolic capacities lost in ARMAN genomes that could enable a free-living lifestyle. These results expand our understanding of genetic diversity, lifestyle, and evolution of “Ca. Micrarchaeota.”

Breakdown of plant biomass in rumen depends on interactions between bacteria, archaea, fungi, and protozoa; however, the majority of studies of the microbiome of ruminants, including the few studies of the rumen of camels, only studied one of these microbial groups. In this study, we applied total rRNA sequencing to identify active microbial communities in 22 solid and liquid rumen samples from 11 camels. These camels were reared at three stations that use different feeding systems: clover, hay and wheat straw (G1), fresh clover (G2), and wheat straw (G3). Bacteria dominated the libraries of sequence reads generated from all rumen samples, followed by protozoa, archaea, and fungi respectively. Firmicutes, Thermoplasmatales, Diplodinium, and Neocallimastix dominated bacterial, archaeal, protozoal and fungal communities, respectively in all samples. Libraries generated from camels reared at facility G2, where they were fed fresh clover, showed the highest alpha diversity. Principal co-ordinate analysis and linear discriminate analysis showed clusters associated with facility/feed and the relative abundance of microbes varied between liquid and solid fractions. This provides preliminary evidence that bacteria dominate the microbial communities of the camel rumen and these communities differ significantly between populations of domesticated camels.

Aquatic environments of volcanic origin provide an exceptional opportunity to study the adaptations of microbial communities to early planet life conditions such as high temperatures, high metal concentrations, and low pH. Here, we characterized the prokaryotic communities and physicochemical properties of seepage sites at the bottom of the Poas Volcano crater and the Agrio River, two geologically related extremely acidic environments located in the Central Volcanic mountain range of Costa Rica. Both locations hold a very low pH (pH 1.79-2.20) and have high sulfate and iron concentrations (Fe = 47-206 mg/L, SO42- = 1170-2460 mg/L measured as S), but significant differences in their temperature (90.0–95.0°C in the seepages at Poas Volcano versus 19.1–26.6 °C in Agrio River) and in the abundance of elemental sulfur. Based on the analysis of 16S rRNA gene sequences, we determined that Sulfobacillus spp., sulfur-oxidizing bacteria, represented more than half (58.4–78.4%) of the sequences in Poas Volcano seepage sites, while Agrio River was dominated by the iron- and sulfur-oxidizing Leptospirillum (7.4–55.5%) and members of the archeal order Thermoplasmatales (16.0-58.2%). Both environments share some chemical characteristics and part of their microbiota, however the temperature and the presence of reduced sulfur are likely the main distinguishing feature ultimately shaping their microbial communities. Our data suggest that in the Poas Volcano-Agrio River system there is a common metabolism but with specialization of species that adapt to the physicochemical conditions of each environment. Universidad de Costa Rica/[809-B6-524]/UCR/Costa Rica Ministerio Federal de Educación e Investigación/[FI-255B-17]/BMBF/Alemania Ministerio de Ciencia, Tecnología y Telecomunicaciones/[FI-255B-17]/MICITT/Costa Rica European Research Council/[ERC250350-IPBSL]/ERC/Unión Europea UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigaciones en Productos Naturales (CIPRONA) UCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de Química UCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela Centroamericana de Geología UCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de Biología UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Electroquímica y Energía Química (CELEQ)

Signal peptides help newly synthesized proteins reach the cell membrane or be secreted. As part of a biological process key to immune response and surveillance in humans, and associated with diseases, for example, Alzheimer, remnant signal peptides and other transmembrane segments are proteolyzed by the intramembrane aspartyl protease (IAP) enzyme family. Here, we identified IAP orthologs throughout the tree of life. In addition to eukaryotes, IAPs are encoded in metabolically diverse archaea from a wide range of environments. We found three distinct clades of archaeal IAPs: (a) Euryarchaeota (eg, halophilic Halobacteriales, methanogenic Methanosarcinales and Methanomicrobiales, marine Poseidoniales, acidophilic Thermoplasmatales, hyperthermophilic Archaeoglobus spp.), (b) DPANN, and (c) Bathyarchaeota, Crenarchaeota, and Asgard. IAPs were also present in bacterial genomes from uncultivated members of Candidate Phylum Radiation, perhaps due to horizontal gene transfer from DPANN archaeal lineages. Sequence analysis of the catalytic motif YD…GXGD (where X is any amino acid) in IAPs from archaea and bacteria reveals WD in Lokiarchaeota and many residue types in the X position. Gene neighborhood analysis in halophilic archaea shows IAP genes near corrinoid transporters (btuCDF genes). In marine Euryarchaeota, a putative BtuF-like domain is found in N-terminus of the IAP gene, suggesting a role for these IAPs in metal ion cofactor or other nutrient scavenging. Interestingly, eukaryotic IAP family members appear to have evolved either from Euryarchaeota or from Asgard archaea. Taken together, our phylogenetic and bioinformatics analysis should prompt experiments to probe the biological roles of IAPs in prokaryotic secretomes.

© 2020 by the authors., The archaeon Cuniculiplasma divulgatum is ubiquitous in acidic environments with low-to-moderate temperatures. However, molecular mechanisms underlying its ability to thrive at lower temperatures remain unexplored. Using mass spectrometry (MS)-based proteomics, we analysed the effect of short-term (3 h) exposure to cold. The C. divulgatum genome encodes 2016 protein-coding genes, from which 819 proteins were identified in the cells grown under optimal conditions. In line with the peptidolytic lifestyle of C. divulgatum, its intracellular proteome revealed the abundance of proteases, ABC transporters and cytochrome C oxidase. From 747 quantifiable polypeptides, the levels of 582 proteins showed no change after the cold shock, whereas 104 proteins were upregulated suggesting that they might be contributing to cold adaptation. The highest increase in expression appeared in low-abundance (0.001–0.005 fmol%) proteins for polypeptides’ hydrolysis (metal-dependent hydrolase), oxidation of amino acids (FAD-dependent oxidoreductase), pyrimidine biosynthesis (aspartate carbamoyltransferase regulatory chain proteins), citrate cycle (2-oxoacid ferredoxin oxidoreductase) and ATP production (V type ATP synthase). Importantly, the cold shock induced a substantial increase (6% and 9%) in expression of the most-abundant proteins, thermosome beta subunit and glutamate dehydrogenase. This study has outlined potential mechanisms of environmental fitness of Cuniculiplasma spp. allowing them to colonise acidic settings at low/moderate temperatures., We acknowledge the support of the Centre for Environmental Biotechnology Project (CEB) part-funded by the European Regional Development Fund (ERDF) through the Welsh Government. R.B. and B.P. acknowledge also the support of the Supercomputing Wales project, which is part-funded by the European Regional Development Fund (ERDF) via the Welsh Government.

Recently, the order Thermoplasmatales was expanded through the cultivation and description of species Cuniculiplasma divulgatum and corresponding family Cuniculiplasmataceae. Initially isolated from acidic streamers, signatures of these archaea were ubiquitously found in various low-pH settings. Eight genomes with various levels of completeness are currently available, all of which exhibit very high sequence identities and genomic conservation. Co-existence of Cuniculiplasmataceae with archaeal Richmond Mine acidophilic nanoorganisms (‘ARMAN’)-related archaea representing an intriguing group within the “microbial dark matter” suggests their common fundamental environmental strategy and metabolic networking. The specific case of “Candidatus Mancarchaeum acidiphilum” Mia14 phylogenetically affiliated with “Ca. Micrarchaeota” from the superphylum “Ca. Diapherotrites” along with the presence of other representatives of ‘DPANN’ with significantly reduced genomes points at a high probability of close interactions between the latter and various Thermoplasmatales abundant in situ. This review critically assesses our knowledge on specific functional role and potential of the members of Cuniculiplasmataceae abundant in acidophilic microbiomes through the analysis of distribution, physiological and genomic patterns, and their interactions with ‘ARMAN’-related archaea. Electronic supplementary material The online version of this article (10.1007/s00792-018-1071-2) contains supplementary material, which is available to authorized users.

Some bacteria and archaea have evolved metabolic strategies that enable them to live in environments contaminated by toxic metals. In fact, many bacteria and archaea take advantage of the redox sensitivity of these very same metals to gain energy via anaerobic respiration. Here, metagenomic techniques were developed and applied along with conventional physiological and ecological methods to elucidate multiple modes of adaptation of bacteria and archaea in metal-contaminated acid mine drainage and aquifer environments contaminated by mine tailings. These approaches provided insight into how these organisms survive and thrive in these environments and how they differentiate themselves from each other.Many of the microbial species in acid mine drainage and mine tailings-contaminated aquifer environments are difficult to culture in the laboratory. Thus, a focus of the research was metabolic analysis of these organisms via analysis of genes and genomes recovered from microbial communities and isolates. Many of the genes are novel, and likely required for specific environmental adaptation, but they are difficult or impossible to functionally characterize based on conventional homology methods. A new method was developed to deal with the challenge of identifying poorly annotated or hypothetical genes of importance in adaptation to metal-contaminated environments. This probabilistic approach is based on conserved gene order between the genomes of interest with distant relatives. The annotation method was used in conjunction with traditional comparative genomics to differentiate a group of co-occurring archaea based on their genetic metabolic potential. These microorganisms grow in biofilm communities in an acid mine drainage system within the Richmond Mine, near Redding in Northern California, USA. Microbial biofilms growing at the air-solution interface were sampled from solutions with pH values of < 1.2, temperatures of up to 48 °C, and mM concentrations of zinc, copper, arsenic, and sub-molar concentrations of dissolved iron. We used a metagenomic approach in which DNA was extracted from biofilm samples, sequenced, and analyzed in order to evaluate differences in the metabolic potential of five closely related Thermoplasmatales archaea and one distant relative. A subset of these organisms appears to be capable of iron oxidation, whereas others appear to live primarily heterotrophically. Another subset is potentially capable of CO oxidation. There are also major differences in motility within the group.A metal-contaminated aquifer adjacent to the Colorado River in Colorado, USA, was studied to investigate microorganisms adapted to high vanadium concentrations. A vanadium-reducing Betaproteobacterium of the genus Simplicispira was isolated (strain BDI). This organism's genome encodes a large number of toxic metal resistance, chemotaxis, motility, and conjugation-related genes that likely allow it to detoxify, avoid contaminants and rapidly adapt in a changing environment. Physiological characterization in the laboratory shows that it is a facultative anaerobic nitrate-reducer capable of reduction of up to 3 mM vanadate.In order to determine the effect of vanadium contamination on the aquifer community structure, soluble vanadium was added to an in-well, flow-through sediment column. Reduction of dissolved vanadate was documented, along with an increase in the number of cells capable of vanadium reduction, and an increase in the relative abundance of strain BDI. An increase in the relative abundance of three families of known vanadium reducing bacteria (Commomonadaceae, Geobacteraceae, and Pseudomonadaceae) was also noted. This experiment confirmed the environmental importance of BDI, and microbial vanadium reduction in response to acetate addition. Following short-term acetate addition to the aquifer, vanadium remained immobile for at least two years. Because the organisms stimulated by amendment were resident in the aquifer and removal of vanadium from solution persists, the acetate addition approach has significant potential for bioremediation of vanadium.In summary, this research used culture-based and culture-independent techniques to elucidate microbial metabolisms that allow organisms to colonize metal-contaminated environments. Vanadium reduction was linked to specific subsurface bacteria, one of which was isolated and characterized. The findings have significance in the fields of genomics, metagenomics, microbial ecology and biogeochemistry, and have potential application for bioremediation.

Small acidophilic archaea belonging to Micrarchaeota and Parvarchaeota phyla are known to physically interact with some Thermoplasmatales members in nature. However, due to a lack of cultivation and limited genomes on hand, their biodiversity, metabolisms, and physiologies remain largely unresolved. Here, we obtained 39 genomes from acid mine drainage (AMD) and hot spring environments around the world. 16S rRNA gene based analyses revealed that Parvarchaeota were only detected in AMD and hot spring habitats, while Micrarchaeota were also detected in others including soil, peat, hypersaline mat, and freshwater, suggesting a considerable higher diversity and broader than expected habitat distribution for this phylum. Despite their small genomes (0.64-1.08 Mb), these archaea may contribute to carbon and nitrogen cycling by degrading multiple saccharides and proteins, and produce ATP via aerobic respiration and fermentation. Additionally, we identified several syntenic genes with homology to those involved in iron oxidation in six Parvarchaeota genomes, suggesting their potential role in iron cycling. However, both phyla lack biosynthetic pathways for amino acids and nucleotides, suggesting that they likely scavenge these biomolecules from the environment and/or other community members. Moreover, low-oxygen enrichments in laboratory confirmed our speculation that both phyla are microaerobic/anaerobic, based on several specific genes identified in them. Furthermore, phylogenetic analyses provide insights into the close evolutionary history of energy related functionalities between both phyla with Thermoplasmatales. These results expand our understanding of these elusive archaea by revealing their involvement in carbon, nitrogen, and iron cycling, and suggest their potential interactions with Thermoplasmatales on genomic scale.

The crystal structure of the Ta1207 protein fromThermoplasma acidophilumis reported. Ta1207 was identified in a screen for high-molecular-weight protein complexes inT. acidophilum. In solution, Ta1207 forms homopentamers of 188 kDa. The crystal structure of recombinant Ta1207 solved by Se-MAD at 2.4 Å resolution revealed a complex with fivefold symmetry. In the crystal lattice, calcium ions induce the formation of a nanocage from two pentamers. The Ta1207 protomers comprise two domains with the same novel α/β topology. A deep pocket with a binding site for a negatively charged group suggests that Ta1207 functions as an intracellular receptor for an unknown ligand. Homologues of Ta1207 occur only in Thermoplasmatales and its function might be related to the extreme lifestyle of these archaea. The thermostable Ta1207 complex might provide a useful fivefold-symmetric scaffold for future nanotechnological applications.

Acid sulfate soil (ASS) is sensitive to redox fluctuations induced by climate change and human activities. Oxidation of sulfur and sulfide in ASS leads to the release of acid and consequently metals, posing severe hazards to coastal environment, while microbial contribution and response to oxidation is poorly understood. Here we used metagenomic sequencing to delineate the shift in microbial community structures and functional genes in ASS upon exposure to aerobic conditions. Aerobic incubation resulted in significant shifts in microbial communities in both topsoil and parent material. Archaea decreased significantly in the parent material after aerobic incubation. The relative abundance of sulfur cycling genes in the parent material layer was significantly higher than those in the topsoil, and multiple sulfide oxidation genes increased after aerobic oxidation. Metagenomic assembly enabled construction of eight key draft genomes from ASS. Three of them (GS3, GS6 and GT3) are novel strains of Thermoplasmatales, Acidothermales (Acidothermus) and Acidimicrobiales, respectively. Functional gene annotation of these population genomes revealed a dominance of sulfur cycling genes and acid tolerant genes. These findings highlight the microbial response to environmental change and identify the ecological adaptation and survival strategies of microbial communities in acid sulfate soils.

Deep-sea hydrothermal vents are amongst the most extreme environments on Earth and represent interesting targets for marine bioprospecting and biodiscovery. The microbial communities in hydrothermal vents are often dominated by chemolithoautotrophs utilizing simple chemical compounds, though the full extent of their heterotrophic abilities is still being explored. In the bioprocessing industry, where degradation of complex organic materials often is a major challenge, new microbial solutions are heavily needed. To meet these needs, we have developed novel in situ incubators and tested if deployment of recalcitrant materials from fish farming and wood-pulping industries introduced changes in the microbial community structure in hot marine hydrothermal sediments. The incubation chambers were deployed in sediments at the Bruse vent site located within the Jan Mayen vent field for 1 year, after which the microbial populations in the chambers were profiled by 16S rRNA Ion Torrent amplicon sequencing. A total of 921 operational taxonomic units (OTUs) were assigned into 74 different phyla where differences in community structure were observed depending on the incubated material, chamber depth below the sea floor and/or temperature. A high fraction of putative heterotrophic microbial lineages related to cultivated members within the Thermotogales were observed. However, considerable fractions of previously uncultivated and novel Thermotogales and Bacteroidetes were also identified. Moreover, several novel lineages (e.g., members within the DPANN superphylum, unidentified archaeal lineages, unclassified Thermoplasmatales and Candidatus division BRC-1 bacterium) of as-yet uncultivated thermophilic archaea and bacteria were identified. Overall, our data illustrate that amendment of hydrothermal vent communities by in situ incubation of biomass induces shifts in community structure toward increased fractions of heterotrophic microorganisms. The technologies utilized here could aid in subsequent metagenomics-based enzyme discovery for diverse industries. publishedVersion

We describe the geochemistry and microbial diversity of a pristine environment that resembles an acid rock drainage (ARD) but it is actually the result of hydrothermal and volcanic influences. We designate this environment, and other comparable sites, as volcanic influenced acid rock drainage (VARD) systems. The metal content and sulfuric acid in this ecosystem stem from the volcanic milieu and not from the product of pyrite oxidation. Based on the analysis of 16S rRNA gene amplicons, we report the microbial community structure in the pristine San Cayetano Costa Rican VARD environment (pH = 2.94–3.06, sulfate ~ 0.87–1.19 g L−1, iron ~ 35–61 mg L−1 (waters), and ~ 8–293 g kg−1 (sediments)). San Cayetano was found to be dominated by microorganisms involved in the geochemical cycling of iron, sulfur, and nitrogen; however, the identity and abundance of the species changed with the oxygen content (0.40–6.06 mg L−1) along the river course. The hypoxic source of San Cayetano is dominated by a putative anaerobic sulfate-reducing Deltaproteobacterium. Sulfur-oxidizing bacteria such as Acidithiobacillus or Sulfobacillus are found in smaller proportions with respect to typical ARD. In the oxic downstream, we identified aerobic iron-oxidizers (Leptospirillum, Acidithrix, Ferrovum) and heterotrophic bacteria (Burkholderiaceae bacterium, Trichococcus, Acidocella). Thermoplasmatales archaea closely related to environmental phylotypes found in other ARD niches were also observed throughout the entire ecosystem. Overall, our study shows the differences and similarities in the diversity and distribution of the microbial communities between an ARD and a VARD system at the source and along the oxygen gradient that establishes on the course of the river., This work was supported by The Vice-rectory of Research of Universidad de Costa Rica (project number VI 809-B6-524), the Costa Rican Ministry of Science, Technology and Telecommunication (MICITT) and Federal Ministry of Education and Research (BMBF) (project VolcanZyme contract No FI-255B-17), and the ERC grant IPBSL (ERC250350-IPBSL). MM acknowledges government funding from the Transitorio I of the National Law 8488 for Emergencies and Risk Prevention in Costa Rica. F.P-S. is supported by grant IJC2018-035180-I from the Spanish Ministry of Science and Innovation.

Numerous archaeal lineages are known to inhabit marine subsurface sediments, although their distributions, metabolic capacities, and interspecies interactions are still not well understood. Abundant and diverse archaea were recently reported in Costa Rica (CR) margin subseafloor sediments recovered during IODP Expedition 334. Here, we recover metagenome-assembled genomes (MAGs) of archaea from the CR margin and compare them to their relatives from shallower settings. We describe 31 MAGs of six different archaeal lineages (Lokiarchaeota, Thorarchaeota, Heimdallarchaeota, Bathyarcheota, Thermoplasmatales, and Hadesarchaea) and thoroughly analyze representative MAGs from the phyla Lokiarchaeota and Bathyarchaeota. Our analysis suggests the potential capability of Lokiarchaeota members to anaerobically degrade aliphatic and aromatic hydrocarbons. We show it is genetically possible and energetically feasible for Lokiarchaeota to degrade benzoate if they associate with organisms using nitrate, nitrite, and sulfite as electron acceptors, which suggests a possibility of syntrophic relationships between Lokiarchaeota and nitrite and sulfite reducing bacteria. The novel Bathyarchaeota lineage possesses an incomplete methanogenesis pathway lacking the methyl coenzyme M reductase complex and encodes a noncanonical acetogenic pathway potentially coupling methylotrophy to acetogenesis via the methyl branch of Wood–Ljungdahl pathway. These metabolic characteristics suggest the potential of this Bathyarchaeota lineage to be a transition between methanogenic and acetogenic Bathyarchaeota lineages. This work expands our knowledge about the metabolic functional repertoire of marine benthic archaea.

Numerous archaeal lineages are known to inhabit marine subsurface sediments, although their distributions, metabolic capacities and interspecies interactions are still not well understood. Abundant and diverse archaea were recently reported in Costa Rica (CR) margin subseafloor sediments recovered during IODP Expedition 334. Here, we recover metagenome-assembled genomes (MAGs) of archaea from the CR-margin and compare them to their relatives from shallower settings. We describe 31 MAGs of 6 different archaeal lineages (Lokiarchaeota, Thorarchaeota, Heimdallarchaeota, Bathyarcheota, Thermoplasmatales and Hadesarchaea) and thoroughly analyze representative MAGs from the phyla Lokiarchaeota and Bathyarchaeota. Our analysis suggests the potential capabilities of Lokiarchaeota members to anaerobically degrade aliphatic and aromatic hydrocarbons. We show it is genetically possible and energetically feasible for Lokiarchaeota to degrade benzoate if they associate with organisms using nitrate, nitrite and sulfite as electron acceptors, which suggests a possibility of syntrophic relationships between Lokiarchaeota and nitrite and sulfite reducers. The novel Bathyarchaeota lineage possesses an incomplete methanogenesis pathway lacking the methyl co-enzyme M reductase complex and encodes a non-canonical acetogenic pathway potentially coupling methylotrophy to acetogenesis via the methyl branch of Wood-Ljundahl pathway. These novel metabolic characteristics suggest the potential of this Bathyarchaeota lineage to be a transition between methanogenic and acetogenic Bathyarchaeota lineages. This work substantially expands our knowledge about the metabolic function repertoire of marine benthic archaea.

The impact of hydrothermal activity on wider ocean geochemistry and microbial ecology remains a topic of much interest. To explore whether hydrothermal microbial signatures are exported to surrounding marine sediments or if such organisms serve as an important source of sedimentary organic matter, we determined the distributions of glycerol dialkyl glycerol tetraether (GDGT) membrane lipids in surficial normal marine sediments, metalliferous sediments and low-temperature hydrothermal deposits at newly discovered hydrothermal fields and adjacent areas at the Southwest Indian Ridge (SWIR). The GDGTs in those samples varied significantly, evidently representing a variable influence of the hydrothermal activity. GDGT compositions of surficial background sediments in SWIR were similar to those commonly observed in marine sediments, dominated by GDGTs associated with marine planktonic archaea and especially GDGT−0 and crenarchaeol. In contrast, the GDGTs of metalliferous sediments strongly impacted by hydrothermal activity and low-temperature hydrothermal deposits were markedly different, characterized by high relative abundances of isoprenoid GDGTs (iGDGTs) bearing multiple rings (yielding a higher ring index), low relative abundances of crenarchaeol, and the presence of glycerol monoalkyl glycerol tetraether lipids (GMGTs; so called ‘H-tetraethers’) that were absent in the normal marine sediments. Sources for these hydrothermal-specific tetraether lipids likely include methanogens and anaerobic methanotrophic archaea (GDGT−0 and GDGT−1–3, respectively), Thermoprotei and Thermoplasmatales (elevated GDGT−3 and 4), and other thermophilic archaea including Methanobacteriales (GMGTs). Deposits influenced by low-temperature hydrothermal activity also contained higher abundances of branched GDGTs (brGDGTs) typically attributed to soil bacteria. The more distal metalliferous sediments influenced by the neutrally buoyant plume did not contain putative hydrothermal GDGTs, having the same GDGT distribution as the background sediments. This suggests that the neutrally buoyant plume has a limited potential to directly influence the organic matter inputs to surrounding sediments, due to a rapidly waning chemosynthetic microbial contribution relative to normal marine contributions as the plume dispersed and was diluted.

Cueva de la Mora (CM) is an acidic, meromictic pit lake in the Iberian Pyrite Belt characterized by extremely high metal(loid) concentrations and strong gradients in oxygen, metal, and nutrient concentrations. We hypothesized that geochemical variations with depth would result in differences in community composition and in metal resistance strategies among active microbial populations. We also hypothesized that metal resistance gene (MRG) expression would correlate with toxicity levels for dissolved metal species in the lake. Water samples were collected in the upper oxic layer, chemocline, and deep anoxic layer of the lake for shotgun metagenomic and metatranscriptomic sequencing. Metagenomic analyses revealed dramatic differences in the composition of the microbial communities with depth, consistent with changing geochemistry. Based on relative abundance of taxa identified in each metagenome, Eukaryotes (predominantly Coccomyxa) dominated the upper layer, while Archaea (predominantly Thermoplasmatales) dominated the deep layer, and a combination of Bacteria and Eukaryotes were abundant at the chemocline. We compared metal resistance across communities using a curated list of protein-coding MRGs with KEGG Orthology identifiers (KOs) and found that there were broad differences in the metal resistance strategies (e.g., intracellular metal accumulation) expressed by Eukaryotes, Bacteria, and Archaea. Although normalized abundances of MRG and MRG expression were generally higher in the deep layer, expression of metal-specific genes was not strongly related to variations in specific metal concentrations, especially for Cu and As. We also compared MRG potential and expression in metagenome assembled genomes (MAGs) from the deep layer, where metal concentrations are highest. Consistent with previous work showing differences in metal resistance mechanisms even at the strain level, MRG expression patterns varied strongly among MAG populations from the same depth. Some MAG populations expressed very few MRG known to date, suggesting that novel metal resistance strategies remain to be discovered in uncultivated acidophiles.

[Background] The current view suggests that in low-temperature acidic environments, archaea are significantly less abundant than bacteria. Thus, this study of the microbiome of Parys Mountain (Anglesey, UK) sheds light on the generality of this current assumption. Parys Mountain is a historically important copper mine and its acid mine drainage (AMD) water streams are characterised by constant moderate temperatures (8–18 °C), extremely low pH (1.7) and high concentrations of soluble iron and other metal cations., [Results] Metagenomic and SSU rRNA amplicon sequencing of DNA from Parys Mountain revealed a significant proportion of archaea affiliated with Euryarchaeota, which accounted for ca. 67% of the community. Within this phylum, potentially new clades of Thermoplasmata were overrepresented (58%), with the most predominant group being “E-plasma”, alongside low-abundant Cuniculiplasmataceae, ‘Ca. Micrarchaeota’ and ‘Terrestrial Miscellaneous Euryarchaeal Group’ (TMEG) archaea, which were phylogenetically close to Methanomassilicoccales and clustered with counterparts from acidic/moderately acidic settings. In the sediment, archaea and Thermoplasmata contributed the highest numbers in V3-V4 amplicon reads, in contrast with the water body community, where Proteobacteria, Nitrospirae, Acidobacteria and Actinobacteria outnumbered archaea. Cultivation efforts revealed the abundance of archaeal sequences closely related to Cuniculiplasma divulgatum in an enrichment culture established from the filterable fraction of the water sample. Enrichment cultures with unfiltered samples showed the presence of Ferrimicrobium acidiphilum, C. divulgatum, ‘Ca. Mancarchaeum acidiphilum Mia14’, ‘Ca. Micrarchaeota’-related and diverse minor (, [Conclusion] Contrary to expectation, our study showed a high abundance of archaea in this extremely acidic mine-impacted environment. Further, archaeal populations were dominated by one particular group, suggesting that they are functionally important. The prevalence of archaea over bacteria in these microbiomes and their spatial distribution patterns represents a novel and important advance in our understanding of acidophile ecology. We also demonstrated a procedure for the specific enrichment of cell wall-deficient members of the archaeal component of this community, although the large fraction of archaeal taxa remained unculturable. Lastly, we identified a separate clustering of globally occurring acidophilic members of TMEG that collectively belong to a distinct order within Thermoplasmata with yet unclear functional roles in the ecosystem., The work of OVG, PNG and SVT was supported by the Royal Society UK-Russia Exchange Grant #IE 160224 (RFBR 16-54-10072 КО project). OVG and PNG acknowledge ERA Net IB2 Project MetaCat through UK Biotechnology and Biological Sciences Research Council (BBSRC) Grant BB/M029085/1 and the support of the Centre of Environmental Biotechnology Project funded by the European Regional Development Fund (ERDF) through the Welsh Government. RB and PNG acknowledge the support of the Supercomputing Wales project, which is part-funded by the European Regional Development Fund (ERDF) via the Welsh Government. The work of AAK and IVK was supported by the Russian Science Foundation grant # 18-44-04024. MF acknowledges grants PCIN-2014-107 (within ERA NET IB2 grant ERA-IB-14-030—MetaCat), PCIN-2017-078 (within the Marine Biotechnology ERA-NET (ERA-MBT) funded under the European Commission’s Seventh Framework Programme, 2013-2017, Grant agreement 604814), BIO2014-54494-R and BIO2017-85522-R from the Spanish Ministry of Economy and Competitiveness.

Here we report the chemical and microbial characterization of the surface water of a CO2-rich hydrothermal vent known in Costa Rica as Borbollones, located at Tenorio Volcano National Park. The Borbollones showed a temperature surrounding 60 °C, a pH of 2.4 and the gas released has a composition of ~ 97% CO2, ~ 0.07% H2S, ~ 2.3% N2 and ~ 0.12% CH4. Other chemical species such as sulfate and iron were found at high levels with respect to typical fresh water bodies. Analysis by 16S rRNA gene metabarcoding revealed that in Borbollones predominates an archaeon from the order Thermoplasmatales and one bacterium from the genus Sulfurimonas. Other sulfur- (genera Thiomonas, Acidithiobacillus, Sulfuriferula, and Sulfuricurvum) and iron-oxidizing bacteria (genera Sideroxydans, Gallionella, and Ferrovum) were identified. Our results show that CO2-influenced surface water of Borbollones contains microorganisms that are usually found in acid rock drainage environments or sulfur-rich hydrothermal vents. To our knowledge, this is the first microbiological characterization of a CO2-dominated hydrothermal spring from Central America and expands our understanding of those extreme ecosystems., The Vice-rectory of Research of Universidad de Costa Rica (809-B6-524), CENIBiot and by the ERC grant IPBSL (ERC250350-IPBSL) supported this research. Data Intensive Academic Grid, which is supported by the USA National Science Foundation (0959894) provided computational resources. F.P-S. is supported by the Spanish Economy and Competitiveness Ministry (MINECO) grant CTM2016-80095-C2-1-R. MMC acknowledges government funding from the Transitorio I of the National Law 8488 for Emergencies and Risk Prevention in Costa Rica. JMdM gratefully acknowledges support from the Deep Carbon Observatory Biology Meets Subduction project.

Background Sustainable management of voluminous and hazardous oily sludge produced by petroleum refineries remains a challenging problem worldwide. Characterization of microbial communities of petroleum contaminated sites has been considered as the essential prerequisite for implementation of suitable bioremediation strategies. Three petroleum refinery sludge samples from North Eastern India were analyzed using next-generation sequencing technology to explore the diversity and functional potential of inhabitant microorganisms and scope for their on-site bioremediation. Results All sludge samples were hydrocarbon rich, anaerobic and reduced with sulfate as major anion and several heavy metals. High throughput sequencing of V3-16S rRNA genes from sludge metagenomes revealed dominance of strictly anaerobic, fermentative, thermophilic, sulfate-reducing bacteria affiliated to Coprothermobacter, Fervidobacterium, Treponema, Syntrophus, Thermodesulfovibrio, Anaerolinea, Syntrophobacter, Anaerostipes, Anaerobaculum, etc., which have been well known for hydrocarbon degradation. Relatively higher proportions of archaea were detected by qPCR. Archaeal 16S rRNA gene sequences showed presence of methanogenic Methanobacterium, Methanosaeta, Thermoplasmatales, etc. Detection of known hydrocarbon utilizing aerobic/facultative anaerobic (Mycobacterium, Pseudomonas, Longilinea, Geobacter, etc.), nitrate reducing (Gordonia, Novosphigobium, etc.) and nitrogen fixing (Azovibrio, Rhodobacter, etc.) bacteria suggested niche specific guilds with aerobic, facultative anaerobic and strict anaerobic populations. Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt) predicted putative genetic repertoire of sludge microbiomes and their potential for hydrocarbon degradation; lipid-, nitrogen-, sulfur- and methane- metabolism. Methyl coenzyme M reductase A (mcrA) and dissimilatory sulfite reductase beta-subunit (dsrB) genes phylogeny confirmed methanogenic and sulfate-reducing activities within sludge environment endowed by hydrogenotrophic methanogens and sulfate-reducing Deltaproteobacteria and Firmicutes members. Conclusion Refinery sludge microbiomes were comprised of hydrocarbon degrading, fermentative, sulfate-reducing, syntrophic, nitrogen fixing and methanogenic microorganisms, which were in accordance with the prevailing physicochemical nature of the samples. Analysis of functional biomarker genes ascertained the activities of methanogenic and sulfate-reducing organisms within sludge environment. Overall data provided better insights on microbial diversity and activity in oil contaminated environment, which could be exploited suitably for in situ bioremediation of refinery sludge. Electronic supplementary material The online version of this article (10.1186/s12866-018-1275-8) contains supplementary material, which is available to authorized users.

© 2017 Finstad, Probst, Thomas, Andersen, Demergasso, Echeverría, Amundson and Banfield. Although once thought to be devoid of biology, recent studies have identified salt deposits as oases for life in the hyperarid Atacama Desert. To examine spatial patterns of microbial species and key nutrient sources, we genomically characterized 26 salt crusts from three sites along a fog gradient. The communities are dominated by a large variety of Halobacteriales and Bacteroidetes, plus a few algal and Cyanobacterial species. CRISPR locus analysis suggests the distribution of a single Cyanobacterial population among all sites. This is in stark contrast to the extremely high sample specificity of most other community members. Only present at the highest moisture site is a genomically characterized Thermoplasmatales archaeon (Marine Group II) and six Nanohaloarchaea, one of which is represented by a complete genome. Parcubacteria (OD1) and Saccharibacteria (TM7), not previously reported from hypersaline environments, were found at low abundances. We found no indication of a N2 fixation pathway in the communities, suggesting acquisition of bioavailable nitrogen from atmospherically derived nitrate. Samples cluster by site based on bacterial and archaeal abundance patterns and photosynthetic capacity decreases with increasing distance from the ocean. We conclude that moisture level, controlled by coastal fog intensity, is the strongest driver of community membership.

&ldquo, Candidatus Micrarchaeota&rdquo, are widely distributed in acidic environments, however, their cultivability and our understanding of their interactions with potential hosts are very limited. Their habitats were so far attributed with acidic sites, soils, peats, freshwater systems, and hypersaline mats. Using cultivation and culture-independent approaches (16S rRNA gene clonal libraries, high-throughput amplicon sequencing of V3-V4 region of 16S rRNA genes), we surveyed the occurrence of these archaea in geothermal areas on Kamchatka Peninsula and Kunashir Island and assessed their taxonomic diversity in relation with another type of low-pH environment, acid mine drainage stream (Wales, UK). We detected &ldquo, Ca. Micrarchaeota&rdquo, in thermophilic heterotrophic enrichment cultures of Kunashir and Kamchatka that appeared as two different phylotypes, namely &ldquo, Ca. Mancarchaeum acidiphilum&rdquo, and ARMAN-2-related, alongside their potential hosts, Cuniculiplasma spp. and other Thermoplasmatales archaea without defined taxonomic position. These clusters of &ldquo, together with three other groups were also present in mesophilic acid mine drainage community. Present work expands our knowledge on the diversity of &ldquo, in thermophilic and mesophilic acidic environments, suggests cultivability patterns of acidophilic archaea and establishes potential links between low-abundance species of thermophilic &ldquo, and certain Thermoplasmatales, such as Cuniculiplasma spp. in situ.

A suite of molecular methods targeting 16S rRNA genes (i.e., DGGE, clone and high-throughput [HTP] amplicon library sequencing) was used to profile the microbial communities in deep Fennoscandian crystalline bedrock fracture fluids. Variation among bacterial 16S rRNA genes was examined with two commonly used primer pairs: P1/P2 and U968f/U1401r. DGGE using U968f/ U1401r mostly detected β-, γ-proteobacteria and Firmicutes, while P1/P2 primers additionally detected other proteobacterial clades and candidate divisions. However, in combination with clone libraries the U968f/U1401r primers detected a higher bacterial diversity than DGGE alone. HTP amplicon sequencing with P1/P2 revealed an abundance of the DGGE bacterial groups as well as many other bacterial taxa likely representing minor components of these communities. Archaeal diversity was investigated via DGGE or HTP amplicon sequencingusing primers A344F/ 519RP. The majority of archaea detected with HTP amplicon sequencing belonged to uncultured Thermoplasmatales and Pendant 33/DHVE3, 4, 6 groups. DGGE of the same samples detected mostly SAGMEG and Methanosarcinales archaea, but almost none of those were revealed by HTP amplicon sequencing. Overall, our results show that the inferred diversity and composition of microbial communities in deep fracture fluids is highly dependent on analytical technique and that the method should be carefully selected with this in mind.

We have studied the distribution of the archaeal communities in Rambla Salada (Murcia, Spain) over three different seasons and observed the influence upon them of the environmental variables, salinity, pH, oxygen and temperature. Samples were collected from three representative sites in order to gain an insight into the archaeal population of the rambla as a whole. Denaturing gradient gel electrophoresis patterns and diversity indexes indicate that the diversity of the archaeal community in Rambla Salada changed mainly according to the season. We found no significant differences between the types of sample studied: watery sediments and soils. The upwelling zone showed most diversity in its archaeal community. The overall archaeal community was composed mainly of Halobacteriales and Thermoplasmatales, accounting for 72.6 and 12.1 % of the total, respectively. Haloarcula was the most abundant genus, being present at all three sites during all three seasons. Some few Crenarchaeota were always found, mainly at low-salinity levels. Ordination canonical correspondence analysis demonstrated that salinity affected the structure of the community significantly, whilst pH, oxygen and temperature did so to a lesser extent. Most Halobacteriales correlated positively with salinity and pH, whilst Thermoplasmatales correlated negatively with both salinity and pH and positively with temperature and oxygen. The archaeal community with the highest diversity was sampled during June 2006, the season with the highest salt concentration. Catalyzed reporter deposition-fluorescence in situ hybridization showed that the percentage of archaea in Rambla Salada compared to the total number of microorganisms (as measured by DAPI) ranged from 11.1 to 16.7 %. Our research group had isolated the most abundant taxon, Haloarcula, previously in Rambla Salada using classical culture techniques, but on this occasion, using culture-independent methods, we were also able to identify some phylotypes, Halorubrum, Methanolobus, Natronomonas, Halomicrobium, Halobacterium, Halosimplex, uncultured Thermoplasmatales and uncultured Crenarchaeota, that had remained undetected during our earlier studies in this habitat.

A new clade of archaea has recently been proposed to constitute the seventh methanogenic order, the Methanomassiliicoccales , which is related to the Thermoplasmatales and the uncultivated archaeal clades deep-sea hydrothermal vent Euryarchaeota group 2 and marine group II Euryarchaeota but only distantly related to other methanogens. In this study, we investigated the membrane lipid composition of Methanomassiliicoccus luminyensis , the sole cultured representative of this seventh order. The lipid inventory of M. luminyensis comprises a unique assemblage of novel lipids as well as lipids otherwise typical for thermophilic, methanogenic, or halophilic archaea. For instance, glycerol sesterpanyl-phytanyl diether core lipids found mainly in halophilic archaea were detected, and so were compounds bearing either heptose or methoxylated glycosidic head groups, neither of which have been reported so far for other archaea. The absence of quinones or methanophenazines is consistent with a biochemistry of methanogenesis different from that of the methanophenazine-containing methylotrophic methanogens. The most distinctive characteristic of the membrane lipid composition of M. luminyensis , however, is the presence of tetraether lipids in which one glycerol backbone is replaced by either butane- or pentanetriol, i.e., lipids recently discovered in marine sediments. Butanetriol dibiphytanyl glycerol tetraether (BDGT) constitutes the most abundant core lipid type (>50% relative abundance) in M. luminyensis . We have thus identified a source for these unusual orphan lipids. The complementary analysis of diverse marine sediment samples showed that BDGTs are widespread in anoxic layers, suggesting an environmental significance of Methanomassiliicoccales and/or related BDGT producers beyond gastrointestinal tracts. IMPORTANCE Cellular membranes of members of all three domains of life, Archaea , Bacteria , and Eukarya , are largely formed by lipids in which glycerol serves as backbone for the hydrophobic alkyl chains. Recently, however, archaeal tetraether lipids with either butanetriol or pentanetriol as a backbone were identified in marine sediments and attributed to uncultured sediment-dwelling archaea. Here we show that the butanetriol-based dibiphytanyl tetraethers constitute the major lipids in Methanomassiliicoccus luminyensis , currently the only isolate of the novel seventh order of methanogens. Given the absence of these lipids in a large set of archaeal isolates, these compounds may be diagnostic for the Methanomassiliicoccales and/or closely related archaea.

The Copahue Thermal Complex is located in the northwest of the province of Neuquen, Argentina in the Norquin department. It lies near the Chilean border, 376 km from the capital of the province at 2,010 meters above sea level. The Copahue complex is located in a geothermally active area in a volcanic caldera, surrounded by multiple fumaroles, potholes, vents, springs and a series of mineromedicinal water lagoons. Copahue is a system of spring acid waters, hyperthermal and largely of high mineralization. The so-called "mother sulfur lagoon" consists of hyperthermal sulfated water, with a dry residue 110°C, 1075 mg /l; pH: 5.6 and chemical analysis shows the following mineral concentrations (mg / l): chlorine 2.7; fluoride 0.3; sulfate 586.3; sodium 29.3; potassium 19.1; lithium 0.02; calcium 44.6; magnesium 11.3; total iron 2.2; ammonium 54.8. The mother sulfur lagoon produces mud that is composed of: smectite (39%), kaolinite (6%), silica (6%), pyrite (2%), alunite (2%), iron oxides (4%), and a strong presence of sulfur at 39%. The contact water maintains an acid pH of the order of 2.5 to 3, and contains dissolved substances on the order of 700 to 1000 mg / l, while the temperature of the water and the mud of this lagoon is one of the highest among the natural manifestations of Copahue. Regarding the mud, it can be added that it is an excellent adsorbent, has an ion exchange capacity of the order of 10 meq / g, and a cooling kinetics that makes it an adequate material to achieve thermal effects associated with its application. This lagoon is the main producer of mud used in the cabins. In terms of medical applications, sulfur is a particularly important component, which as pure mineral has disinfectant, keratolytic and antiseptic properties, and can also be transformed to give rise to other minerals that contain sulfur (in forms reduced and oxidized) such as alunite and pyrite. At the same time, both kaolinite and smectite act as excellent adsorbents, dermal protectors and give softness to peloids, while improving their thermal properties. The microbiological richness that it possesses is also an outstanding feature of this system. The prokaryotic biodiversity of the Sulfur Lagoon (LS T = 54° C and pH: 3) was studied using two complementary techniques: a) phylogenetic analysis of the 16S rRNA genes of bacteria and archaea; and b) in situ hybridization of specific fluorescent probes (FIHS) for a quantitative estimation of microbial biodiversity. The results obtained showed that in this thermal well 90% of the microorganisms are archaea, 6% are bacteria and 4% could not be detected by FIHS. Within the bacteria, species of Pseudomonas, Achromobacter, Acidiphilum, and Mezorhizobium were identified and among the archaea the dominant genera were Thermoplasmatales and Sulfolobus. Additionally, a species of the native genus Acidianus of Copahue was detected, which was named A. copahuensis and which has highly varied metabolic characteristics: it is acidophilic, thermophilic, facultative anaerobic, sulfur and iron oxidant. Finally, in LS, other thermophilic anaerobic archaea were also detected, such as Vulcanisaeta and Thermocladium. Copahue is a very interesting source of biodiversity both for its contribution to global biology and its biotechnological potential. The mineral richness of naturally matured mud and microbiota allows the treatment of a large number of dermatological, osteoarticular and rheumatological pathologies. Regarding the presentation of cases in dermatology, the following are highlighted: mud therapy combined with phototherapy in the treatment of psoriasis, and in the treatment of Rosacea and venous ulcers using cyanophytic algae bioglea. Finally, it is important to underscore the existence of a large number of cases documented each season that validate the effectiveness of treatment of different pathologies, for example the combination of mud with magnetotherapy and ultrasound in a patient with radiodermitic ulcers and pain treatment combining mud therapy and galvanic therapy.

Highly acidic (pH 0–1) biofilms, known as ‘snottites', form on the walls and ceilings of hydrogen sulfide-rich caves. We investigated the population structure, physiology and biogeochemistry of these biofilms using metagenomics, rRNA methods and lipid geochemistry. Snottites from the Frasassi cave system (Italy) are dominated (>70% of cells) by Acidithiobacillus thiooxidans, with smaller populations including an archaeon in the uncultivated ‘G-plasma' clade of Thermoplasmatales (>15%) and a bacterium in the Acidimicrobiaceae family (>5%). Based on metagenomic evidence, the Acidithiobacillus population is autotrophic (ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO), carboxysomes) and oxidizes sulfur by the sulfide–quinone reductase and sox pathways. No reads matching nitrogen fixation genes were detected in the metagenome, whereas multiple matches to nitrogen assimilation functions are present, consistent with geochemical evidence, that fixed nitrogen is available in the snottite environment to support autotrophic growth. Evidence for adaptations to extreme acidity include Acidithiobacillus sequences for cation transporters and hopanoid synthesis, and direct measurements of hopanoid membrane lipids. Based on combined metagenomic, molecular and geochemical evidence, we suggest that Acidithiobacillus is the snottite architect and main primary producer, and that snottite morphology and distributions in the cave environment are directly related to the supply of C, N and energy substrates from the cave atmosphere.

Molecular diversity of methanogens in the rumen of Korean black goats was investigated with 16S rRNA gene clone libraries using methanogen-specific primers. The libraries were composed of rumen fluid-associated methanogens (FAM) and rumen particle-associated methanogens (PAM) from rumen-fistulated Korean black goats. Among the 141 clones of the FAM library, the sequences were mostly related to two phyla, the Methanobacteriaceae family (77.3%) and the Thermoplasmatales family (22.7%); and among the 68 clones of the PAM library, sequences were also mainly clustered in the two phyla, the Thermoplasmatales family (63.24%) and the Methanobacteriaceae family (35.29%). Most of the sequenced clones in the two libraries were closely related to uncultured methanogenic archaeon. Quantitative real-time PCR revealed that PAM (8.97 log 10) had significantly higher (P < 0.01) density of methanogens by the methanogenic 16S rRNA gene copies than FAM (7.57 log 10). The two clone libraries also showed difference in Shannon index (FAM library 1.70 and PAM library 1.59) and Chao 1 estimator (FAM library 18 and PAM library 17 operational taxonomic units). Apparent differences found in the microbial community from the two 16S rRNA gene libraries could be a result of such factors as the chemical and physical nature of the target material surface, types or component of diets, the interaction between the methanogens and other microbes, and age of the experimental goats.