Halobacterium yunchengense sp. nov., Natronomonas amylolytica sp. nov., Halorientalis halophila sp. nov., Halobellus salinisoli sp. nov., halophilic archaea isolated from a saline lake and inland saline soil.

Four halophilic archaeal strains YCN1T, YCN58T, LT38T, and LT62T were isolated from Yuncheng Salt Lake (Shanxi, China) and Tarim Basin (Xinjiang, China), respectively. Phylogenetic and phylogenomic analyses showed that these four strains tightly cluster with related species of Halobacterium, Natronomonas, Halorientalis, and Halobellus, respectively. The AAI, ANI, and dDDH values between these four strains and their related species of respective genera were lower than the proposed threshold values for species delineation. Strains YCN1T, YCN58T, LT38T, and LT62T could be differentiated from the current species of Halobacterium, Natronomonas, Halorientalis, and Halobellus, respectively, based on the comparison of diverse phenotypic characteristics. The polar lipid profiles of these four strains were closely similar to those of respective relatives within the genera Halobacterium, Natronomonas, Halorientalis, and Halobellus, respectively. The phenotypic, phylogenetic, and genome-based analyses indicated that strains YCN1T, YCN58T, LT38T, and LT62T represent respective novel species within the genera Halobacterium, Natronomonas, Halorentalis, and Halobellus, for which the names Halobacterium yunchengense sp. nov., Natronomonas amylolytica sp. nov., Halorientalis halophila sp. nov., and Halobellus salinisoli sp. nov. are proposed, respectively.

Borg extrachromosomal elements of methane-oxidizing archaea have conserved and expressed genetic repertoires.

Borgs are huge extrachromosomal elements (ECE) of anaerobic methane-consuming "Candidatus Methanoperedens" archaea. Here, we used nanopore sequencing to validate published complete genomes curated from short reads and to reconstruct new genomes. 13 complete and four near-complete linear genomes share 40 genes that define a largely syntenous genome backbone. We use these conserved genes to identify new Borgs from peatland soil and to delineate Borg phylogeny, revealing two major clades. Remarkably, Borg genes encoding nanowire-like electron-transferring cytochromes and cell surface proteins are more highly expressed than those of host Methanoperedens, indicating that Borgs augment the Methanoperedens activity in situ. We reconstructed the first complete 4.00 Mbp genome for a Methanoperedens that is inferred to be a Borg host and predicted its methylation motifs, which differ from pervasive TC and CC methylation motifs of the Borgs. Thus, methylation may enable Methanoperedens to distinguish their genomes from those of Borgs. Very high Borg to Methanoperedens ratios and structural predictions suggest that Borgs may be capable of encapsulation. The findings clearly define Borgs as a distinct class of ECE with shared genomic signatures, establish their diversification from a common ancestor with genetic inheritance, and raise the possibility of periodic existence outside of host cells.

A genome catalogue of mercury-methylating bacteria and archaea from sediments of a boreal river facing human disturbances.

Methyl mercury, a toxic compound, is produced by anaerobic microbes and magnifies in aquatic food webs, affecting the health of animals and humans. The exploration of mercury methylators based on genomes is still limited, especially in the context of river ecosystems. To address this knowledge gap, we developed a genome catalogue of potential mercury-methylating microorganisms. This was based on the presence of hgcAB from the sediments of a river affected by two run-of-river hydroelectric dams, logging activities and a wildfire. Through the use of genome-resolved metagenomics, we discovered a unique and diverse group of mercury methylators. These were dominated by members of the metabolically versatile Bacteroidota and were particularly rich in microbes that ferment butyrate. By comparing the diversity and abundance of mercury methylators between sites subjected to different disturbances, we found that ongoing disturbances, such as the input of organic matter related to logging activities, were particularly conducive to the establishment of a mercury-methylating niche. Finally, to gain a deeper understanding of the environmental factors that shape the diversity of mercury methylators, we compared the mercury-methylating genome catalogue with the broader microbial community. The results suggest that mercury methylators respond to environmental conditions in a manner similar to the overall microbial community. Therefore, it is crucial to interpret the diversity and abundance of mercury methylators within their specific ecological context.

Description of Asgardarchaeum abyssi gen. nov. spec. nov., a novel species within the class Asgardarchaeia and phylum Asgardarchaeota in accordance with the SeqCode.

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.

Minimal and hybrid hydrogenases are active from archaea.

Microbial hydrogen (H2) cycling underpins the diversity and functionality of diverse anoxic ecosystems. Among the three evolutionarily distinct hydrogenase superfamilies responsible, [FeFe] hydrogenases were thought to be restricted to bacteria and eukaryotes. Here, we show that anaerobic archaea encode diverse, active, and ancient lineages of [FeFe] hydrogenases through combining analysis of existing and new genomes with extensive biochemical experiments. [FeFe] hydrogenases are encoded by genomes of nine archaeal phyla and expressed by H2-producing Asgard archaeon cultures. We report an ultraminimal hydrogenase in DPANN archaea that binds the catalytic H-cluster and produces H2. Moreover, we identify and characterize remarkable hybrid complexes formed through the fusion of [FeFe] and [NiFe] hydrogenases in ten other archaeal orders. Phylogenetic analysis and structural modeling suggest a deep evolutionary history of hybrid hydrogenases. These findings reveal new metabolic adaptations of archaea, streamlined H2 catalysts for biotechnological development, and a surprisingly intertwined evolutionary history between the two major H2-metabolizing enzymes.

Asgard archaeal selenoproteome reveals a roadmap for the archaea-to-eukaryote transition of selenocysteine incorporation machinery.

Selenocysteine (Sec) is encoded by the UGA codon that normally functions as a stop signal and is specifically incorporated into selenoproteins via a unique recoding mechanism. The translational recoding of UGA as Sec is directed by an unusual RNA structure, the SECIS element. Although archaea and eukaryotes adopt similar Sec encoding machinery, the SECIS elements have no similarities to each other with regard to sequence and structure. We analyzed >400 Asgard archaeal genomes to examine the occurrence of both Sec encoding system and selenoproteins in this archaeal superphylum, the closest prokaryotic relatives of eukaryotes. A comprehensive map of Sec utilization trait has been generated, providing the most detailed understanding of the use of this nonstandard amino acid in Asgard archaea so far. By characterizing the selenoproteomes of all organisms, several selenoprotein-rich phyla and species were identified. Most Asgard archaeal selenoprotein genes possess eukaryotic SECIS-like structures with varying degrees of diversity. Moreover, euryarchaeal SECIS elements might originate from Asgard archaeal SECIS elements via lateral gene transfer, indicating a complex and dynamic scenario of the evolution of SECIS element within archaea. Finally, a roadmap for the transition of eukaryotic SECIS elements from archaea was proposed, and selenophosphate synthetase may serve as a potential intermediate for the generation of ancestral eukaryotic SECIS element. Our results offer new insights into a deeper understanding of the evolution of Sec insertion machinery.

EzBioCloud: a genome-driven database and platform for microbiome identification and discovery.

With the continued evolution of DNA sequencing technologies, the role of genome sequence data has become more integral in the classification and identification of Bacteria and Archaea. Six years after introducing EzBioCloud, an integrated platform representing the taxonomic hierarchy of Bacteria and Archaea through quality-controlled 16S rRNA gene and genome sequences, we present an updated version, that further refines and expands its capabilities. The current update recognizes the growing need for accurate taxonomic information as defining a species increasingly relies on genome sequence comparisons. We also incorporated an advanced strategy for addressing underrepresented or less studied lineages, bolstering the comprehensiveness and accuracy of our database. Our rigorous quality control protocols remain, where whole-genome assemblies from the NCBI Assembly Database undergo stringent screening to remove low-quality sequence data. These are then passed through our enhanced identification bioinformatics pipeline which initiates a 16S rRNA gene similarity search and then calculates the average nucleotide identity (ANI). For genome sequences lacking a 16S rRNA sequence and without a closely related genomic representative for ANI calculation, we apply a different ANI approach using bacterial core genes for improved taxonomic placement (core gene ANI, cgANI). Because of the increase in genome sequences available in NCBI and our newly introduced cgANI method, EzBioCloud now encompasses a total of 109 835 species, of which 21 964 have validly published names. 47 896 are candidate species identified either through 16S rRNA sequence similarity (phylotypes) or through whole genome ANI (genomospecies), and the remaining 39 975 were positioned in the taxonomic tree by cgANI (species clusters). Our EzBioCloud database is accessible at www.ezbiocloud.net/db.

A non-methanogenic archaeon within the order Methanocellales.

Serpentinization, a geochemical process found on modern and ancient Earth, provides an ultra-reducing environment that can support microbial methanogenesis and acetogenesis. Several groups of archaea, such as the order Methanocellales, are characterized by their ability to produce methane. Here, we generate metagenomic sequences from serpentinized springs in The Cedars, California, and construct a circularized metagenome-assembled genome of a Methanocellales archaeon, termed Met12, that lacks essential methanogenesis genes. The genome includes genes for an acetyl-CoA pathway, but lacks genes encoding methanogenesis enzymes such as methyl-coenzyme M reductase, heterodisulfide reductases and hydrogenases. In situ transcriptomic analyses reveal high expression of a multi-heme c-type cytochrome, and heterologous expression of this protein in a model bacterium demonstrates that it is capable of accepting electrons. Our results suggest that Met12, within the order Methanocellales, is not a methanogen but a CO2-reducing, electron-fueled acetogen without electron bifurcation.

How Do Thermophiles Organize Their Genomes?

All cells must maintain the structural and functional integrity of the genome under a wide range of environments. High temperatures pose a formidable challenge to cells by denaturing the DNA double helix, causing chemical damage to DNA, and increasing the random thermal motion of chromosomes. Thermophiles, predominantly classified as bacteria or archaea, exhibit an exceptional capacity to mitigate these detrimental effects and prosper under extreme thermal conditions, with some species tolerating temperatures higher than 100°C. Their genomes are mainly characterized by the presence of reverse gyrase, a unique topoisomerase that introduces positive supercoils into DNA. This enzyme has been suggested to maintain the genome integrity of thermophiles by limiting DNA melting and mediating DNA repair. Previous studies provided significant insights into the mechanisms by which NAPs, histones, SMC superfamily proteins, and polyamines affect the 3D genomes of thermophiles across different scales. Here, I discuss current knowledge of the genome organization in thermophiles and pertinent research questions for future investigations.

Fusion/fission protein family identification in Archaea.

The majority of newly discovered archaeal lineages remain without a cultivated representative, but scarce experimental data from the cultivated organisms show that they harbor distinct functional repertoires. To unveil the ecological as well as evolutionary impact of Archaea from metagenomics, new computational methods need to be developed, followed by in-depth analysis. Among them is the genome-wide protein fusion screening performed here. Natural fusions and fissions of genes not only contribute to microbial evolution but also complicate the correct identification and functional annotation of sequences. The products of these processes can be defined as fusion (or composite) proteins, the ones consisting of two or more domains originally encoded by different genes and split proteins, and the ones originating from the separation of a gene in two (fission). Fusion identifications are required for proper phylogenetic reconstructions and metabolic pathway completeness assessments, while mappings between fused and unfused proteins can fill some of the existing gaps in metabolic models. In the archaeal genome-wide screening, more than 1,900 fusion/fission protein clusters were identified, belonging to both newly sequenced and well-studied lineages. These protein families are mainly associated with different types of metabolism, genetic, and cellular processes. Moreover, 162 of the identified fusion/fission protein families are archaeal specific, having no identified fused homolog within the bacterial domain. Our approach was validated by the identification of experimentally characterized fusion/fission cases. However, around 25% of the identified fusion/fission families lack functional annotations for both composite and split states, showing the need for experimental characterization in Archaea.IMPORTANCEGenome-wide fusion screening has never been performed in Archaea on a broad taxonomic scale. The overlay of multiple computational techniques allows the detection of a fine-grained set of predicted fusion/fission families, instead of rough estimations based on conserved domain annotations only. The exhaustive mapping of fused proteins to bacterial organisms allows us to capture fusion/fission families that are specific to archaeal biology, as well as to identify links between bacterial and archaeal lineages based on cooccurrence of taxonomically restricted proteins and their sequence features. Furthermore, the identification of poorly characterized lineage-specific fusion proteins opens up possibilities for future experimental and computational investigations. This approach enhances our understanding of Archaea in general and provides potential candidates for in-depth studies in the future.

Investigating the nature of prokaryotic genomic island locations within a genome.

Horizontal gene transfer (HGT) is a powerful evolutionary force that considerably shapes the structure of prokaryotic genomes and is associated with genomic islands (GIs). A GI is a DNA segment composed of transferred genes that can be found within a prokaryotic genome, obtained through HGT. Much research has focused on detecting GIs in genomes, but here we pursue a new course, which is identifying possible preferred locations of GIs in the prokaryotic genome. Here, we identify the locations of the GIs within prokaryotic genomes to examine patterns in those locations. Prokaryotic GIs were analyzed according to the genome structure that they are located in, whether it be a circular or a linear genome. The analytical investigations employed are: (1) studying the GI locations in relation to the origin of replication (oriC); (2) exploring the distances between GIs; and (3) determining the distribution of GIs across the genomes. For each of the investigations, the analysis was performed on all of the GIs in the data set. Moreover, to void bias caused by the distribution of the genomes represented, the GIs in one genome from each species and the GIs of the most frequent species are also analyzed. Overall, the results showed that there are preferred sites for the GIs in the genome. In the linear genomes, these sites are usually located in the oriC region and terminus region, while in the circular genomes, they are located solely in the terminus region. These results also showed that the distance distribution between the GIs is almost exponential, which proves that GIs have preferred sites within genomes. The oriC and termniuns are preferred sites for the GIs and a possible natural explanation for this could be connected to the content of the oriC region. Moreover, the content of the GIs in terms of its protein families was studied and the results demonstrated that the majority of frequent protein families are close to identical in each section.

Analysis of nearly 3000 archaeal genomes from terrestrial geothermal springs sheds light on interconnected biogeochemical processes.

Terrestrial geothermal springs are physicochemically diverse and host abundant populations of Archaea. However, the diversity, functionality, and geological influences of these Archaea are not well understood. Here we explore the genomic diversity of Archaea in 152 metagenomes from 48 geothermal springs in Tengchong, China, collected from 2016 to 2021. Our dataset is comprised of 2949 archaeal metagenome-assembled genomes spanning 12 phyla and 392 newly identified species, which increases the known species diversity of Archaea by ~48.6%. The structures and potential functions of the archaeal communities are strongly influenced by temperature and pH, with high-temperature acidic and alkaline springs favoring archaeal abundance over Bacteria. Genome-resolved metagenomics and metatranscriptomics provide insights into the potential ecological niches of these Archaea and their potential roles in carbon, sulfur, nitrogen, and hydrogen metabolism. Furthermore, our findings illustrate the interplay of competition and cooperation among Archaea in biogeochemical cycles, possibly arising from overlapping functional niches and metabolic handoffs. Taken together, our study expands the genomic diversity of Archaea inhabiting geothermal springs and provides a foundation for more incisive study of biogeochemical processes mediated by Archaea in geothermal ecosystems.

AnnoView enables large-scale analysis, comparison, and visualization of microbial gene neighborhoods.

The analysis and comparison of gene neighborhoods is a powerful approach for exploring microbial genome structure, function, and evolution. Although numerous tools exist for genome visualization and comparison, genome exploration across large genomic databases or user-generated datasets remains a challenge. Here, we introduce AnnoView, a web server designed for interactive exploration of gene neighborhoods across the bacterial and archaeal tree of life. Our server offers users the ability to identify, compare, and visualize gene neighborhoods of interest from 30 238 bacterial genomes and 1672 archaeal genomes, through integration with the comprehensive Genome Taxonomy Database and AnnoTree databases. Identified gene neighborhoods can be visualized using pre-computed functional annotations from different sources such as KEGG, Pfam and TIGRFAM, or clustered based on similarity. Alternatively, users can upload and explore their own custom genomic datasets in GBK, GFF or CSV format, or use AnnoView as a genome browser for relatively small genomes (e.g. viruses and plasmids). Ultimately, we anticipate that AnnoView will catalyze biological discovery by enabling user-friendly search, comparison, and visualization of genomic data. AnnoView is available at http://annoview.uwaterloo.ca.

Genomic comparison of deep-sea hydrothermal genera related to Aeropyrum, Thermodiscus and Caldisphaera, and proposed emended description of the family Acidilobaceae.

Deep-sea hydrothermal vents host archaeal and bacterial thermophilic communities, including taxonomically and functionally diverse Thermoproteota. Despite their prevalence in high-temperature submarine communities, Thermoproteota are chronically under-represented in genomic databases and issues have emerged regarding their nomenclature, particularly within the Aeropyrum-Thermodiscus-Caldisphaera. To resolve some of these problems, we identified 47 metagenome-assembled genomes (MAGs) within this clade, from 20 previously published deep-sea hydrothermal vent and submarine volcano metagenomes, and 24 MAGs from public databases. Using phylogenomic analysis, Genome Taxonomy Database Toolkit (GTDB-Tk) taxonomic assessment, 16S rRNA gene phylogeny, average amino acid identity (AAI) and functional gene patterns, we re-evaluated of the taxonomy of the Aeropyrum-Thermodiscus-Caldisphaera. At least nine genus-level clades were identified with two or more MAGs. In accordance with SeqCode requirements and recommendations, we propose names for three novel genera, viz. Tiamatella incendiivivens, Hestiella acidicharens and Calypsonella navitae. A fourth genus was also identified related to Thermodiscus maritimus, for which no available sequenced genome exists. We propose the novel species Thermodiscus eudorianus to describe our high-quality Thermodiscus MAG, which represents the type genome for the genus. All three novel genera and T. eudorianus are likely anaerobic heterotrophs, capable of fermenting protein-rich carbon sources, while some Tiamatella, Calypsonella and T. eudorianus may also reduce polysulfides, thiosulfate, sulfur and/or selenite, and the likely acidophile, Hestiella, may reduce nitrate and/or perchlorate. Based on phylogenomic evidence, we also propose the family Acidilobaceae be amended to include Caldisphaera, Aeropyrum, Thermodiscus and Stetteria and the novel genera described here.

Metabolic potential of Nitrososphaera-associated clades.

Soil ammonia-oxidizing archaea (AOA) play a crucial role in converting ammonia to nitrite, thereby mobilizing reactive nitrogen species into their soluble form, with a significant impact on nitrogen losses from terrestrial soils. Yet, our knowledge regarding their diversity and functions remains limited. In this study, we reconstructed 97 high-quality AOA metagenome-assembled genomes (MAGs) from 180 soil samples collected in Central Germany during 2014-2019 summers. These MAGs were affiliated with the order Nitrososphaerales and clustered into four family-level clades (NS-α/γ/δ/ε). Among these MAGs, 75 belonged to the most abundant but least understood δ-clade. Within the δ-clade, the amoA genes in three MAGs from neutral soils showed a 99.5% similarity to the fosmid clone 54d9, which has served as representative of the δ-clade for the past two decades since even today no cultivated representatives are available. Seventy-two MAGs constituted a distinct δ sub-clade, and their abundance and expression activity were more than twice that of other MAGs in slightly acidic soils. Unlike the less abundant clades (α, γ, and ε), the δ-MAGs possessed multiple highly expressed intracellular and extracellular carbohydrate-active enzymes responsible for carbohydrate binding (CBM32) and degradation (GH5), along with highly expressed genes involved in ammonia oxidation. Together, these results suggest metabolic versatility of uncultured soil AOA and a potential mixotrophic or chemolithoheterotrophic lifestyle among 54d9-like AOA.

Phenotypic and genomic characterization of Methanothermobacter wolfeii strain BSEL, a CO2-capturing archaeon with minimal nutrient requirements.

A new variant of Methanothermobacter wolfeii was isolated from an anaerobic digester using enrichment cultivation in anaerobic conditions. The new isolate was taxonomically identified via 16S rRNA gene sequencing and tagged as M. wolfeii BSEL. The whole genome of the new variant was sequenced and de novo assembled. Genomic variations between the BSEL strain and the type strain were discovered, suggesting evolutionary adaptations of the BSEL strain that conferred advantages while growing under a low concentration of nutrients. M. wolfeii BSEL displayed the highest specific growth rate ever reported for the wolfeii species (0.27 ± 0.03 h-1) using carbon dioxide (CO2) as unique carbon source and hydrogen (H2) as electron donor. M. wolfeii BSEL grew at this rate in an environment with ammonium (NH4+) as sole nitrogen source. The minerals content required to cultivate the BSEL strain was relatively low and resembled the ionic background of tap water without mineral supplements. Optimum growth rate for the new isolate was observed at 64°C and pH 8.3. In this work, it was shown that wastewater from a wastewater treatment facility can be used as a low-cost alternative medium to cultivate M. wolfeii BSEL. Continuous gas fermentation fed with a synthetic biogas mimic along with H2 in a bubble column bioreactor using M. wolfeii BSEL as biocatalyst resulted in a CO2 conversion efficiency of 97% and a final methane (CH4) titer of 98.5%v, demonstrating the ability of the new strain for upgrading biogas to renewable natural gas.IMPORTANCEAs a methanogenic archaeon, Methanothermobacter wolfeii uses CO2 as electron acceptor, producing CH4 as final product. The metabolism of M. wolfeii can be harnessed to capture CO2 from industrial emissions, besides producing a drop-in renewable biofuel to substitute fossil natural gas. If used as biocatalyst in new-generation CO2 sequestration processes, M. wolfeii has the potential to accelerate the decarbonization of the energy generation sector, which is the biggest contributor of CO2 emissions worldwide. Nonetheless, the development of CO2 sequestration archaeal-based biotechnology is still limited by an uncertainty in the requirements to cultivate methanogenic archaea and the unknown longevity of archaeal cultures. In this study, we report the adaptation, isolation, and phenotypic characterization of a novel variant of M. wolfeii, which is capable of maximum growth with minimal nutrients input. Our findings demonstrate the potential of this variant for the production of renewable natural gas, paving the way for the development of more efficient and sustainable CO2 sequestration processes.

Using the pan-genomic framework for the discovery of genomic islands in the haloarchaeon Halorubrum ezzemoulense.

In this study, we use pan-genomics to characterize the genomic variability of the widely dispersed halophilic archaeal species Halorubrum ezzemoulense (Hez). We include a multi-regional sampling of newly sequenced, high-quality draft genomes. The pan-genome graph of the species reveals 50 genomic islands that represent rare accessory genetic capabilities available to members. Most notably, we observe rearrangements that have led to the insertion/recombination/replacement of mutually exclusive genomic islands in equivalent genome positions ("homeocassettes"). These conflicting islands encode for similar functions, but homologs from islands located between the same core genes exhibit high divergence on the amino acid level, while the neighboring core genes are nearly identical. Both islands of a homeocassette often coexist in the same geographic location, suggesting that either island may be beyond the reach of selective sweeps and that these loci of divergence between Hez members are maintained and persist long term. This implies that subsections of the population have different niche preferences and rare metabolic capabilities. After an evaluation of the gene content in the homeocassettes, we speculate that these islands may play a role in the speciation, niche adaptability, and group selection dynamics in Hez. Though homeocassettes are first described in this study, similar replacements and divergence of genes on genomic islands have been previously reported in other Haloarchaea and distantly related Archaea, suggesting that homeocassettes may be a feature in a wide range of organisms outside of Hez.IMPORTANCEThis study catalogs the rare genes discovered in strains of the species Halorubrum ezzemoulense (Hez), an obligate halophilic archaeon, through the perspective of its pan-genome. These rare genes are often found to be arranged on islands that confer metabolic and transport functions and contain genes that have eluded previous studies. The discovery of divergent, but homologous islands occupying equivalent genome positions ("homeocassettes") in different genomes, reveals significant new information on genome evolution in Hez. Homeocassette pairs encode for similar functions, but their dissimilarity and distribution imply high rates of recombination, different specializations, and niche preferences in Hez. The coexistence of both islands of a homeocassette pair in multiple environments demonstrates that both islands are beyond the reach of selective sweeps and that these genome content differences between strains persist long term. The switch between islands through recombination under different environmental conditions may lead to a greater range of niche adaptability in Hez.

A global survey of prokaryotic genomes reveals the eco-evolutionary pressures driving horizontal gene transfer.

Horizontal gene transfer, the exchange of genetic material through means other than reproduction, is a fundamental force in prokaryotic genome evolution. Genomic persistence of horizontally transferred genes has been shown to be influenced by both ecological and evolutionary factors. However, there is limited availability of ecological information about species other than the habitats from which they were isolated, which has prevented a deeper exploration of ecological contributions to horizontal gene transfer. Here we focus on transfers detected through comparison of individual gene trees to the species tree, assessing the distribution of gene-exchanging prokaryotes across over a million environmental sequencing samples. By analysing detected horizontal gene transfer events, we show distinct functional profiles for recent versus old events. Although most genes transferred are part of the accessory genome, genes transferred earlier in evolution tend to be more ubiquitous within present-day species. We find that co-occurring, interacting and high-abundance species tend to exchange more genes. Finally, we show that host-associated specialist species are most likely to exchange genes with other host-associated specialist species, whereas species found across different habitats have similar gene exchange rates irrespective of their preferred habitat. Our study covers an unprecedented scale of integrated horizontal gene transfer and environmental information, highlighting broad eco-evolutionary trends.

Halomontanus rarus gen. nov., sp. nov., a novel halophilic archaeon of the family Natrialbaceae from salt lakes on the Qinghai-Xizang Plateau.

Two halophilic archaeal strains TS33T and KZCA124 were isolated from two distant salt lakes on the Qinghai-Xizang Plateau, respectively. Culture-independent analysis indicated that these two strains were original inhabitants but low abundant taxa in respective salt lakes. Strains TS33T and KZCA124 were able to grow at 20-60 °C (optimum were 42 and 35 °C, respectively), with 0.9-4.8 M NaCl (optimum were 3.0 and 2.6 M, respectively), with 0-0.7 M MgCl2 (optimum, 0.3 M) and at pH 5.0-9.5 (optimum were pH 7.5 and pH 7, respectively). The 16S rRNA and rpoB' gene similarities between these two strains were 99.7% and 99.4%, and these two similarities among strains TS33T, KZCA124, and existing species of the family Natrialbaceae were 90.6-95.5% and 84.4-89.3%, respectively. Phylogenetic and phylogenomic analyses indicated that strains TS33T and KZCA124 formed an independent branch separated from neighboring genera, Saliphagus, Natronosalvus, and Natronobiforma. The averagenucleotideidentity (ANI), digital DNA-DNAhybridization (dDDH), and average amino acid identity (AAI) values between strains TS33T and KZCA124 were 96.4%, 73.1%, and 96.7%, respectively, higher than the thresholds for species demarcation. The overall genome-related indexes between these two strains and existing species of family Natrialbaceae were 73-77%, 21-25%, and 63-70%, respectively, significantly lower than the species boundary thresholds. Strains TS33T and KZCA124 may represent a novel species of a new genus within the family Natrialbaceae judged by the cutoff value of AAI (≤76%) proposed to differentiate genera within the family Natrialbaceae. The major polar lipids of strains TS33T and KZCA124 were phosphatidic acid, phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester, sulfated mannosyl glucosyl diether, and sulfated galactosyl mannosyl glucosyl diether. These two strains could be distinguished from the related genera according to differential phenotypic characteristics. These phenotypic, phylogenetic, and genomic analyses revealed that strains TS33T (=KCTC 4310T = MCCC 4K00132T) and KZCA124 (=CGMCC 1.17432 = JCM 34316) represent a novel species of a new genus of the family Natrialbaceae and were named Halomontanus rarus gen. nov., sp. nov.

Cell surface architecture of the cultivated DPANN archaeon Nanobdella aerobiophila.

The DPANN archaeal clade includes obligately ectosymbiotic species. Their cell surfaces potentially play an important role in the symbiotic interaction between the ectosymbionts and their hosts. However, little is known about the mechanism of ectosymbiosis. Here, we show cell surface structures of the cultivated DPANN archaeon Nanobdella aerobiophila strain MJ1T and its host Metallosphaera sedula strain MJ1HA, using a variety of electron microscopy techniques, i.e., negative-staining transmission electron microscopy, quick-freeze deep-etch TEM, and 3D electron tomography. The thickness, unit size, and lattice symmetry of the S-layer of strain MJ1T were different from those of the host archaeon strain MJ1HA. Genomic and transcriptomic analyses highlighted the most highly expressed MJ1T gene for a putative S-layer protein with multiple glycosylation sites and immunoglobulin-like folds, which has no sequence homology to known S-layer proteins. In addition, genes for putative pectin lyase- or lectin-like extracellular proteins, which are potentially involved in symbiotic interaction, were found in the MJ1T genome based on in silico 3D protein structure prediction. Live cell imaging at the optimum growth temperature of 65°C indicated that cell complexes of strains MJ1T and MJ1HA were motile, but sole MJ1T cells were not. Taken together, we propose a model of the symbiotic interaction and cell cycle of Nanobdella aerobiophila.IMPORTANCEDPANN archaea are widely distributed in a variety of natural and artificial environments and may play a considerable role in the microbial ecosystem. All of the cultivated DPANN archaea so far need host organisms for their growth, i.e., obligately ectosymbiotic. However, the mechanism of the ectosymbiosis by DPANN archaea is largely unknown. To this end, we performed a comprehensive analysis of the cultivated DPANN archaeon, Nanobdella aerobiophila, using electron microscopy, live cell imaging, transcriptomics, and genomics, including 3D protein structure prediction. Based on the results, we propose a reasonable model of the symbiotic interaction and cell cycle of Nanobdella aerobiophila, which will enhance our understanding of the enigmatic physiology and ecological significance of DPANN archaea.