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Family GH1 glycosyl hydrolases are ubiquitous in prokaryotes and eukaryotes and are utilized in numerous industrial applications, including bioconversion of lignocelluloses. In this study, hyperacidophilic archaeon Cuniculiplasma divulgatum (S5T=JCM 30642T) was explored as a source of novel carbohydrate-active enzymes. The genome of C. divulgatum encodes three GH1 enzyme candidates, from which CIB12 and CIB13 were heterologously expressed and characterized. Phylogenetic analysis of CIB12 and CIB13 clustered them with β-glucosidases from genuinely thermophilic archaea including Thermoplasma acidophilum, Picrophilus torridus, Sulfolobus solfataricus, Pyrococcus furiosus , and Thermococcus kodakarensis. Purified enzymes showed maximal activities at pH 4.5–6.0 (CIB12) and 4.5–5.5 (CIB13) with optimal temperatures at 50°C, suggesting a high-temperature origin of Cuniculiplasma spp. ancestors. Crystal structures of both enzymes revealed a classical (α/β)8 TIM-barrel fold with the active site located inside the barrel close to the C-termini of β-strands including the catalytic residues Glu204 and Glu388 (CIB12), and Glu204 and Glu385 (CIB13). Both enzymes preferred cellobiose over lactose as substrates and were classified as cellobiohydrolases. Cellobiose addition increased the biomass yield of Cuniculiplasma cultures growing on peptides by 50%, suggesting that the cellobiohydrolases expand the carbon substrate range and hence environmental fitness of Cuniculiplasma. [ABSTRACT FROM AUTHOR]

Simple Summary: The application of extremophilic, acidophilic microbial populations for the bio-oxidation of gold-bearing sulfide concentrates in industry is a source of great interest in the research of these populations. Understanding the effects of different factors on the activity of such acidophilic populations may allow for the performance of the bio-oxidation of sulfide concentrates to be regulated. In the present work, we studied the effects of different temperatures as well as carbon sources (carbon dioxide and molasses) on the rate of the bio-oxidation of a gold-bearing pyrite–arsenopyrite concentrate as well as on the composition of the microbial population performing this process. It was shown that an increase in the temperature from 40 to 50 °C led to a decrease in the intensity of bio-oxidation, while the application of additional carbon dioxide as a carbon source made it possible to prevent the inhibition of bio-oxidation due to a temperature increase. An analysis of the populations formed under different experimental conditions revealed that both temperature and carbon dioxide affected the composition of the microbial population which, in turn, may explain the effect on the bio-oxidation performance. Thus, the use of additional carbon dioxide may be proposed as the method to increase the efficiency of bio-oxidation and to prevent the negative effect of a temperature increase on the activity of bio-oxidation. In the present study, the effect of additional carbon sources (carbon dioxide and molasses) on the bio-oxidation of a pyrite–arsenopyrite concentrate at temperatures of 40–50 °C was studied, and novel data regarding the patterns of the bio-oxidation of gold-bearing sulfide concentrates and the composition of the microbial populations performing these processes were obtained. At 40 °C, additional carbon sources did not affect the bio-oxidation efficiency. At the same time, the application of additional carbon dioxide improved the bio-oxidation performance at temperatures of 45 and 50 °C and made it possible to avoid the inhibition of bio-oxidation due to an increase in the temperature. Therefore, the use of additional carbon dioxide may be proposed to prevent the negative effect of an increase in temperature on the bio-oxidation of sulfide concentrates. 16S rRNA gene profiling revealed archaea of the family Thermoplasmataceae (Acidiplasma, Ferroplasma, Cuniculiplasma, and A-plasma group) and bacteria of the genera Leptospirillum, with Sulfobacillus and Acidithiobacillus among the dominant groups in the community. Temperature influenced the composition of the communities to a greater extent than the additional sources of carbon and the mode of operation of the bioreactor. Elevating the temperature from 40 °C to 50 °C resulted in increases in the shares of Acidiplasma and Sulfobacillus and decreases in the relative abundances of Ferroplasma, Leptospirillum, and Acidithiobacillus, while Cuniculiplasma and A-plasma were more abundant at 45 °C. A metagenomic analysis of the studied population made it possible to characterize novel archaea belonging to an uncultivated, poorly-studied group of Thermoplasmatales which potentially plays an important role in the bio-oxidation process. Based on an analysis of the complete genome, we propose describing the novel species and novel genus as "Candidatus Carboxiplasma ferriphilum" gen. nov., spec. nov. [ABSTRACT FROM AUTHOR]

Background: Archaea of the order Thermoplasmatales are widely distributed in natural acidic areas and are amongst the most acidophilic prokaryotic organisms known so far. These organisms are difficult to culture, with currently only six genera validly published since the discovery of Thermoplasma acidophilum in 1970. Moreover, known great diversity of uncultured Thermoplasmatales represents microbial dark matter and underlines the necessity of efforts in cultivation and study of these archaea. Organisms from the order Thermoplasmatales affiliated with the so-called "alphabet-plasmas", and collectively dubbed "E-plasma", were the focus of this study. These archaea were found predominantly in the hyperacidic site PM4 of Parys Mountain, Wales, UK, making up to 58% of total metagenomic reads. However, these archaea escaped all cultivation attempts. Results: Their genome-based metabolism revealed its peptidolytic potential, in line with the physiology of the previously studied Thermoplasmatales isolates. Analyses of the genome and evolutionary history reconstruction have shown both the gain and loss of genes, that may have contributed to the success of the "E-plasma" in hyperacidic environment compared to their community neighbours. Notable genes among them are involved in the following molecular processes: signal transduction, stress response and glyoxylate shunt, as well as multiple copies of genes associated with various cellular functions; from energy production and conversion, replication, recombination, and repair, to cell wall/membrane/envelope biogenesis and archaella production. History events reconstruction shows that these genes, acquired by putative common ancestors, may determine the evolutionary and functional divergences of "E-plasma", which is much more developed than other representatives of the order Thermoplasmatales. In addition, the ancestral hereditary reconstruction strongly indicates the placement of Thermogymnomonas acidicola close to the root of the Thermoplasmatales. Conclusions: This study has analysed the metagenome-assembled genome of "E-plasma", which denotes the basis of their predominance in Parys Mountain environmental microbiome, their global ubiquity, and points into the right direction of further cultivation attempts. The results suggest distinct evolutionary trajectories of organisms comprising the order Thermoplasmatales, which is important for the understanding of their evolution and lifestyle. [ABSTRACT FROM AUTHOR]

Aquatic environments of volcanic origin provide an exceptional opportunity to study the adaptations of microorganisms to early planet life conditions. 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 Costa Rica. Both locations hold a low pH (1.79–2.20) and have high sulfate and iron concentrations (Fe = 47–206 mg/L, SO42− = 1170–2460 mg/L), but significant differences in their temperature (90.0–95.0 ºC in the seepages at Poas Volcano, 19.1–26.6 ºC in Agrio River) and in the elemental sulfur content. Based on the analysis of 16S rRNA gene sequences, we determined that Sulfobacillus spp. represented more than half of the sequences in Poas Volcano seepage sites, while Agrio River was dominated by Leptospirillum and members of the archaeal order Thermoplasmatales. Both environments share some chemical characteristics and part of their microbiota, however, the temperature and the reduced sulfur are likely the main distinguishing features, 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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Background: Metal sulfide mineral dissolution during bioleaching and acid mine drainage (AMD) formation creates an environment that is inhospitable to most life. Despite dominance by a small number of bacteria, AMD microbial biofilm communities contain a notable variety of coexisting and closely related Euryarchaea, most of which have defied cultivation efforts. For this reason, we used metagenomics to analyze variation in gene content that may contribute to niche differentiation among co-occurring AMD archaea. Our analyses targeted members of the Thermoplasmatales and related archaea. These results greatly expand genomic information available for this archaeal order. Results: We reconstructed near-complete genomes for uncultivated, relatively low abundance organisms A-, E-, and Gplasma, members of Thermoplasmatales order, and for a novel organism, Iplasma. Genomic analyses of these organisms, as well as Ferroplasma type I and II, reveal that all are facultative aerobic heterotrophs with the ability to use many of the same carbon substrates, including methanol. Most of the genomes share genes for toxic metal resistance and surface-layer production. Only Aplasma and Eplasma have a full suite of flagellar genes whereas all but the Ferroplasma spp. have genes for pili production. Cryogenic-electron microscopy (cryo-EM) and tomography (cryo-ET) strengthen these metagenomics-based ultrastructural predictions. Notably, only Aplasma, Gplasma and the Ferroplasma spp. have predicted iron oxidation genes and Eplasma and Iplasma lack most genes for cobalamin, valine, (iso)leucine and histidine synthesis. Conclusion: The Thermoplasmatales AMD archaea share a large number of metabolic capabilities. All of the uncultivated organisms studied here (A-, E-, G-, and Iplasma) are metabolically very similar to characterized Ferroplasma spp., differentiating themselves mainly in their genetic capabilities for biosynthesis, motility, and possibly iron oxidation. These results indicate that subtle, but important genomic differences, coupled with unknown differences in gene expression, distinguish these organisms enough to allow for co-existence. Overall this study reveals shared features of organisms from the Thermoplasmatales lineage and provides new insights into the functioning of AMD communities. [ABSTRACT FROM AUTHOR]

Abstract: Based on the molecular analysis of human subgingival plaque samples from 30 periodontitis patients a novel lineage of Archaea within the phylogenetic radiation of Thermoplasmatales was identified in 10% of cases. Co-occurrence of unique 16S rRNA gene and mcrA gene sequences suggests that this lineage corresponds to a hitherto unknown group of methanogens. [Copyright &y& Elsevier]

In view of the continuous increase in the number of described species of nonmethanogenic hyperthermophilic and related thermophilic archaea thriving in geothermally heated habitats (i.e., members of the orders Thermoproteales, Desulfurococcales, Sulfolobales, Thermoplasmatales, Thermococcales and Archaeoglobales), their current taxonomic status and taxonomic properties are reviewed. The introduction of 16S rRNA phylogeny has brought a more reliable taxonomic framework for these archaea, although, their phenotypic properties still remain as salient and descriptive features. Moreover, certain cellular components are potentially applicable to their taxonomic classification. [Copyright &y& Elsevier]

"Candidatus Micrarchaeota" 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 "Ca. Micrarchaeota" in thermophilic heterotrophic enrichment cultures of Kunashir and Kamchatka that appeared as two different phylotypes, namely "Ca. Mancarchaeum acidiphilum"-, and ARMAN-2-related, alongside their potential hosts, Cuniculiplasma spp. and other Thermoplasmatales archaea without defined taxonomic position. These clusters of "Ca. Micrarchaeota" together with three other groups were also present in mesophilic acid mine drainage community. Present work expands our knowledge on the diversity of "Ca. Micrarchaeota" in thermophilic and mesophilic acidic environments, suggests cultivability patterns of acidophilic archaea and establishes potential links between low-abundance species of thermophilic "Ca. Micrarchaeota" and certain Thermoplasmatales, such as Cuniculiplasma spp. in situ. [ABSTRACT FROM AUTHOR]