Physiological potential and evolutionary trajectories of syntrophic sulfate-reducing bacterial partners of anaerobic methanotrophic archaea.

Sulfate-coupled anaerobic oxidation of methane (AOM) is performed by multicellular consortia of anaerobic methanotrophic archaea (ANME) in obligate syntrophic partnership with sulfate-reducing bacteria (SRB). Diverse ANME and SRB clades co-associate but the physiological basis for their adaptation and diversification is not well understood. In this work, we used comparative metagenomics and phylogenetics to investigate the metabolic adaptation among the 4 main syntrophic SRB clades (HotSeep-1, Seep-SRB2, Seep-SRB1a, and Seep-SRB1g) and identified features associated with their syntrophic lifestyle that distinguish them from their non-syntrophic evolutionary neighbors in the phylum Desulfobacterota. We show that the protein complexes involved in direct interspecies electron transfer (DIET) from ANME to the SRB outer membrane are conserved between the syntrophic lineages. In contrast, the proteins involved in electron transfer within the SRB inner membrane differ between clades, indicative of convergent evolution in the adaptation to a syntrophic lifestyle. Our analysis suggests that in most cases, this adaptation likely occurred after the acquisition of the DIET complexes in an ancestral clade and involve horizontal gene transfers within pathways for electron transfer (CbcBA) and biofilm formation (Pel). We also provide evidence for unique adaptations within syntrophic SRB clades, which vary depending on the archaeal partner. Among the most widespread syntrophic SRB, Seep-SRB1a, subclades that specifically partner ANME-2a are missing the cobalamin synthesis pathway, suggestive of nutritional dependency on its partner, while closely related Seep-SRB1a partners of ANME-2c lack nutritional auxotrophies. Our work provides insight into the features associated with DIET-based syntrophy and the adaptation of SRB towards it.

Giant virus diversity and host interactions through global metagenomics.

Our current knowledge about nucleocytoplasmic large DNA viruses (NCLDVs) is largely derived from viral isolates that are co-cultivated with protists and algae. Here we reconstructed 2,074 NCLDV genomes from sampling sites across the globe by building on the rapidly increasing amount of publicly available metagenome data. This led to an 11-fold increase in phylogenetic diversity and a parallel 10-fold expansion in functional diversity. Analysis of 58,023 major capsid proteins from large and giant viruses using metagenomic data revealed the global distribution patterns and cosmopolitan nature of these viruses. The discovered viral genomes encoded a wide range of proteins with putative roles in photosynthesis and diverse substrate transport processes, indicating that host reprogramming is probably a common strategy in the NCLDVs. Furthermore, inferences of horizontal gene transfer connected viral lineages to diverse eukaryotic hosts. We anticipate that the global diversity of NCLDVs that we describe here will establish giant viruses-which are associated with most major eukaryotic lineages-as important players in ecosystems across Earth's biomes.

IMG/PR: a database of plasmids from genomes and metagenomes with rich annotations and metadata.

Plasmids are mobile genetic elements found in many clades of Archaea and Bacteria. They drive horizontal gene transfer, impacting ecological and evolutionary processes within microbial communities, and hold substantial importance in human health and biotechnology. To support plasmid research and provide scientists with data of an unprecedented diversity of plasmid sequences, we introduce the IMG/PR database, a new resource encompassing 699 973 plasmid sequences derived from genomes, metagenomes and metatranscriptomes. IMG/PR is the first database to provide data of plasmid that were systematically identified from diverse microbiome samples. IMG/PR plasmids are associated with rich metadata that includes geographical and ecosystem information, host taxonomy, similarity to other plasmids, functional annotation, presence of genes involved in conjugation and antibiotic resistance. The database offers diverse methods for exploring its extensive plasmid collection, enabling users to navigate plasmids through metadata-centric queries, plasmid comparisons and BLAST searches. The web interface for IMG/PR is accessible at https://img.jgi.doe.gov/pr. Plasmid metadata and sequences can be downloaded from https://genome.jgi.doe.gov/portal/IMG_PR.

Comparative genomics reveals electron transfer and syntrophic mechanisms differentiating methanotrophic and methanogenic archaea.

The anaerobic oxidation of methane coupled to sulfate reduction is a microbially mediated process requiring a syntrophic partnership between anaerobic methanotrophic (ANME) archaea and sulfate-reducing bacteria (SRB). Based on genome taxonomy, ANME lineages are polyphyletic within the phylum Halobacterota, none of which have been isolated in pure culture. Here, we reconstruct 28 ANME genomes from environmental metagenomes and flow sorted syntrophic consortia. Together with a reanalysis of previously published datasets, these genomes enable a comparative analysis of all marine ANME clades. We review the genomic features that separate ANME from their methanogenic relatives and identify what differentiates ANME clades. Large multiheme cytochromes and bioenergetic complexes predicted to be involved in novel electron bifurcation reactions are well distributed and conserved in the ANME archaea, while significant variations in the anabolic C1 pathways exists between clades. Our analysis raises the possibility that methylotrophic methanogenesis may have evolved from a methanotrophic ancestor.

The IMG/M data management and analysis system v.7: content updates and new features.

The Integrated Microbial Genomes & Microbiomes system (IMG/M: https://img.jgi.doe.gov/m/) at the Department of Energy (DOE) Joint Genome Institute (JGI) continues to provide support for users to perform comparative analysis of isolate and single cell genomes, metagenomes, and metatranscriptomes. In addition to datasets produced by the JGI, IMG v.7 also includes datasets imported from public sources such as NCBI Genbank, SRA, and the DOE National Microbiome Data Collaborative (NMDC), or submitted by external users. In the past couple years, we have continued our effort to help the user community by improving the annotation pipeline, upgrading the contents with new reference database versions, and adding new analysis functionalities such as advanced scaffold search, Average Nucleotide Identity (ANI) for high-quality metagenome bins, new cassette search, improved gene neighborhood display, and improvements to metatranscriptome data display and analysis. We also extended the collaboration and integration efforts with other DOE-funded projects such as NMDC and DOE Biology Knowledgebase (KBase).

IMG/VR v4: an expanded database of uncultivated virus genomes within a framework of extensive functional, taxonomic, and ecological metadata.

Viruses are widely recognized as critical members of all microbiomes. Metagenomics enables large-scale exploration of the global virosphere, progressively revealing the extensive genomic diversity of viruses on Earth and highlighting the myriad of ways by which viruses impact biological processes. IMG/VR provides access to the largest collection of viral sequences obtained from (meta)genomes, along with functional annotation and rich metadata. A web interface enables users to efficiently browse and search viruses based on genome features and/or sequence similarity. Here, we present the fourth version of IMG/VR, composed of >15 million virus genomes and genome fragments, a ≈6-fold increase in size compared to the previous version. These clustered into 8.7 million viral operational taxonomic units, including 231 408 with at least one high-quality representative. Viral sequences in IMG/VR are now systematically identified from genomes, metagenomes, and metatranscriptomes using a new detection approach (geNomad), and IMG standard annotation are complemented with genome quality estimation using CheckV, taxonomic classification reflecting the latest taxonomic standards, and microbial host taxonomy prediction. IMG/VR v4 is available at https://img.jgi.doe.gov/vr, and the underlying data are available to download at https://genome.jgi.doe.gov/portal/IMG_VR.

Horizontal gene transfer of the Mer operon is associated with large effects on the transcriptome and increased tolerance to mercury in nitrogen-fixing bacteria.

BACKGROUND: Mercury (Hg) is highly toxic and has the potential to cause severe health problems for humans and foraging animals when transported into edible plant parts. Soil rhizobia that form symbiosis with legumes may possess mechanisms to prevent heavy metal translocation from roots to shoots in plants by exporting metals from nodules or compartmentalizing metal ions inside nodules. Horizontal gene transfer has potential to confer immediate de novo adaptations to stress. We used comparative genomics of high quality de novo assemblies to identify structural differences in the genomes of nitrogen-fixing rhizobia that were isolated from a mercury (Hg) mine site that show high variation in their tolerance to Hg. RESULTS: Our analyses identified multiple structurally conserved merA homologs in the genomes of Sinorhizobium medicae and Rhizobium leguminosarum but only the strains that possessed a Mer operon exhibited 10-fold increased tolerance to Hg. RNAseq analysis revealed nearly all genes in the Mer operon were significantly up-regulated in response to Hg stress in free-living conditions and in nodules. In both free-living and nodule environments, we found the Hg-tolerant strains with a Mer operon exhibited the fewest number of differentially expressed genes (DEGs) in the genome, indicating a rapid and efficient detoxification of Hg from the cells that reduced general stress responses to the Hg-treatment. Expression changes in S. medicae while in bacteroids showed that both rhizobia strain and host-plant tolerance affected the number of DEGs. Aside from Mer operon genes, nif genes which are involved in nitrogenase activity in S. medicae showed significant up-regulation in the most Hg-tolerant strain while inside the most Hg-accumulating host-plant. Transfer of a plasmid containing the Mer operon from the most tolerant strain to low-tolerant strains resulted in an immediate increase in Hg tolerance, indicating that the Mer operon is able to confer hyper tolerance to Hg. CONCLUSIONS: Mer operons have not been previously reported in nitrogen-fixing rhizobia. This study demonstrates a pivotal role of the Mer operon in effective mercury detoxification and hypertolerance in nitrogen-fixing rhizobia. This finding has major implications not only for soil bioremediation, but also host plants growing in mercury contaminated soils.

Antiquaquibacter oligotrophicus gen. nov., sp. nov., a novel oligotrophic bacterium from groundwater.

In this study, a Gram-stain-positive, non-motile, oxidase- and catalase-negative, rod-shaped, bacterial strain (SG_E_30_P1T) that formed light yellow colonies was isolated from a groundwater sample of Sztaravoda spring, Hungary. Based on 16S rRNA phylogenetic and phylogenomic analyses, the strain was found to form a distinct linage within the family Microbacteriaceae. Its closest relatives in terms of near full-length 16S rRNA gene sequences are Salinibacterium hongtaonis MH299814 (97.72 % sequence similarity) and Leifsonia psychrotolerans GQ406810 (97.57 %). The novel strain grows optimally at 20-28 °C, at neutral pH and in the presence of NaCl (1-2 w/v%). Strain SG_E_30_P1T contains MK-7 and B-type peptidoglycan with diaminobutyrate as the diagnostic amino acid. The major cellular fatty acids are anteiso-C15 : 0, iso-C16 : 0 and iso-C14 : 0, and the polar lipid profile is composed of diphosphatidylglycerol and phosphatidylglycerol, as well as an unidentified aminoglycolipid, aminophospholipid and some unidentified phospholipids. The assembled draft genome is a contig with a total length of 2 897 968 bp and a DNA G+C content of 65.5 mol%. Amino acid identity values with it closest relatives with sequenced genomes of <62.54 %, as well as other genome distance results, indicate that this bacterium represents a novel genus within the family Microbacteriaceae. We suggest that SG_E_30_P1T (=DSM 111415T=NCAIM B.02656T) represents the type strain of a novel genus and species for which the name Antiquaquibacter oligotrophicus gen. nov., sp. nov. is proposed.

IMG/VR v3: an integrated ecological and evolutionary framework for interrogating genomes of uncultivated viruses.

Viruses are integral components of all ecosystems and microbiomes on Earth. Through pervasive infections of their cellular hosts, viruses can reshape microbial community structure and drive global nutrient cycling. Over the past decade, viral sequences identified from genomes and metagenomes have provided an unprecedented view of viral genome diversity in nature. Since 2016, the IMG/VR database has provided access to the largest collection of viral sequences obtained from (meta)genomes. Here, we present the third version of IMG/VR, composed of 18 373 cultivated and 2 314 329 uncultivated viral genomes (UViGs), nearly tripling the total number of sequences compared to the previous version. These clustered into 935 362 viral Operational Taxonomic Units (vOTUs), including 188 930 with two or more members. UViGs in IMG/VR are now reported as single viral contigs, integrated proviruses or genome bins, and are annotated with a new standardized pipeline including genome quality estimation using CheckV, taxonomic classification reflecting the latest ICTV update, and expanded host taxonomy prediction. The new IMG/VR interface enables users to efficiently browse, search, and select UViGs based on genome features and/or sequence similarity. IMG/VR v3 is available at https://img.jgi.doe.gov/vr, and the underlying data are available to download at https://genome.jgi.doe.gov/portal/IMG_VR.

The IMG/M data management and analysis system v.6.0: new tools and advanced capabilities.

The Integrated Microbial Genomes & Microbiomes system (IMG/M: https://img.jgi.doe.gov/m/) contains annotated isolate genome and metagenome datasets sequenced at the DOE's Joint Genome Institute (JGI), submitted by external users, or imported from public sources such as NCBI. IMG v 6.0 includes advanced search functions and a new tool for statistical analysis of mixed sets of genomes and metagenome bins. The new IMG web user interface also has a new Help page with additional documentation and webinar tutorials to help users better understand how to use various IMG functions and tools for their research. New datasets have been processed with the prokaryotic annotation pipeline v.5, which includes extended protein family assignments.

Community Structure and Microbial Associations in Sediment-Free Methanotrophic Enrichment Cultures from a Marine Methane Seep.

Syntrophic consortia of anaerobic methanotrophic archaea (ANME) and sulfate-reducing bacteria (SRB) consume large amounts of methane and serve as the foundational microorganisms in marine methane seeps. Despite their importance in the carbon cycle, research on the physiology of ANME-SRB consortia has been hampered by the slow growth and complex physicochemical environment the consortia inhabit. Here, we report successful sediment-free enrichment of ANME-SRB consortia from deep-sea methane seep sediments in the Santa Monica Basin, California. Anoxic Percoll density gradients and size-selective filtration were used to separate ANME-SRB consortia from sediment particles and single cells to accelerate the cultivation process. Over a 3-year period, a subset of the sediment-associated ANME and SRB lineages, predominantly comprised of ANME-2a/2b ("Candidatus Methanocomedenaceae") and their syntrophic bacterial partners, SEEP-SRB1/2, adapted and grew under defined laboratory conditions. Metagenome-assembled genomes from several enrichments revealed that ANME-2a, SEEP-SRB1, and Methanococcoides in different enrichments from the same inoculum represented distinct species, whereas other coenriched microorganisms were closely related at the species level. This suggests that ANME, SRB, and Methanococcoides are more genetically diverse than other members in methane seeps. Flow cytometry sorting and sequencing of cell aggregates revealed that Methanococcoides, Anaerolineales, and SEEP-SRB1 were overrepresented in multiple ANME-2a cell aggregates relative to the bulk metagenomes, suggesting they were physically associated and possibly interacting. Overall, this study represents a successful case of selective cultivation of anaerobic slow-growing microorganisms from sediments based on their physical characteristics, introducing new opportunities for detailed genomic, physiological, biochemical, and ecological analyses. IMPORTANCE Biological anaerobic oxidation of methane (AOM) coupled with sulfate reduction represents a large methane sink in global ocean sediments. Methane consumption is carried out by syntrophic archaeal-bacterial consortia and fuels a unique ecosystem, yet the interactions in these slow-growing syntrophic consortia and with other associated community members remain poorly understood. The significance of this study is the establishment of sediment-free enrichment cultures of anaerobic methanotrophic archaea and sulfate-reducing bacteria performing AOM with sulfate using selective cultivation approaches based on size, density, and metabolism. By reconstructing microbial genomes and analyzing community composition of the enrichment cultures and cell aggregates, we shed light on the diversity of microorganisms physically associated with AOM consortia beyond the core syntrophic partners. These enrichment cultures offer simplified model systems to extend our understanding of the diversity of microbial interactions within marine methane seeps.

Faunimonas pinastri gen. nov., sp. nov., an endophyte from a pine tree of the family Pleomorphomonadaceae, class Alphaproteobacteria.

Bacterial strain A52C2T was isolated from the endophytic microbial community of a Pinus pinaster tree trunk and characterized. Strain A52C2T stained Gram-negative and formed rod-shaped cells that grew optimally at 30 °C and at pH 6.0-7.0. The G+C content of the DNA was 65.1 mol %. The respiratory quinone was ubiquinone 10, and the major fatty acids were cyclo-C19:0 ω8c and C18:0, representing 70.1 % of the total fatty acids. Phylogenetic analyses based on the 16S rRNA gene sequences placed strain A52C2T in a distinct lineage within the order Hyphomicrobiales, family Pleomorphomonadaceae. The 16S rRNA gene sequence similarities of A52C2T to that of Mongoliimonas terrestris and Oharaeibacter diazotrophicus were 93.15 and 93.2 %, respectively. The draft genome sequence of strain A52C2T comprises 4 196 045 bases with a 195-fold mapped coverage of the genome. The assembled genome consists of 43 contigs of more than 1 000 bp (N50 contig size was 209 720 bp). The genome encodes 4033 putative coding sequences. The phylogenetic, phenotypic and chemotaxonomic data showed that strain A52C2T (=UCCCB 130T=CECT 8949T=LMG 29042T) represents the type of a novel species and genus, for which we propose the name Faunimonas pinastri gen. nov., sp. nov.

SAR11 bacteria linked to ocean anoxia and nitrogen loss.

Bacteria of the SAR11 clade constitute up to one half of all microbial cells in the oxygen-rich surface ocean. SAR11 bacteria are also abundant in oxygen minimum zones (OMZs), where oxygen falls below detection and anaerobic microbes have vital roles in converting bioavailable nitrogen to N2 gas. Anaerobic metabolism has not yet been observed in SAR11, and it remains unknown how these bacteria contribute to OMZ biogeochemical cycling. Here, genomic analysis of single cells from the world's largest OMZ revealed previously uncharacterized SAR11 lineages with adaptations for life without oxygen, including genes for respiratory nitrate reductases (Nar). SAR11 nar genes were experimentally verified to encode proteins catalysing the nitrite-producing first step of denitrification and constituted ~40% of OMZ nar transcripts, with transcription peaking in the anoxic zone of maximum nitrate reduction activity. These results link SAR11 to pathways of ocean nitrogen loss, redefining the ecological niche of Earth's most abundant organismal group.

Hydrogenotrophic methanogenesis in archaeal phylum Verstraetearchaeota reveals the shared ancestry of all methanogens.

Methanogenic archaea are major contributors to the global carbon cycle and were long thought to belong exclusively to the euryarchaeal phylum. Discovery of the methanogenesis gene cluster methyl-coenzyme M reductase (Mcr) in the Bathyarchaeota, and thereafter the Verstraetearchaeota, led to a paradigm shift, pushing back the evolutionary origin of methanogenesis to predate that of the Euryarchaeota. The methylotrophic methanogenesis found in the non-Euryarchaota distinguished itself from the predominantly hydrogenotrophic methanogens found in euryarchaeal orders as the former do not couple methanogenesis to carbon fixation through the reductive acetyl-CoA [Wood-Ljungdahl pathway (WLP)], which was interpreted as evidence for independent evolution of the two methanogenesis pathways. Here, we report the discovery of a complete and divergent hydrogenotrophic methanogenesis pathway in a thermophilic order of the Verstraetearchaeota, which we have named Candidatus Methanohydrogenales, as well as the presence of the WLP in the crenarchaeal order Desulfurococcales. Our findings support the ancient origin of hydrogenotrophic methanogenesis, suggest that methylotrophic methanogenesis might be a later adaptation of specific orders, and provide insight into how the transition from hydrogenotrophic to methylotrophic methanogenesis might have occurred.

Hymenobacter caeli sp. nov., an airborne bacterium isolated from King George Island, Antarctica.

A rod-shaped and Gram-stain-negative bacterial strain 9AT, was isolated from an air sample collected at King George Island, maritime Antarctica. Phylogenetic analysis based on 16S rRNA gene sequence reveals that strain 9AT belongs to the genus Hymenobacter and shows the highest similarity to Hymenobacter coccineus CCM 8649T (96.8 %). The DNA G+C content based on the draft genome sequence is 64.9 mol%. Strain 9AT is strictly aerobic, psychrophilic, catalase-positive, oxidase-positive and non-motile. Growth is observed at 0-20 °C (optimum 10 °C), pH 6.0-8.0 (optimum pH 7.0), and in the absence of NaCl. The predominant menaquinone of strain 9AT is MK-7 and the major fatty acids comprise Summed Feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c; 25.2 %), iso-C15 : 0 (23.2 %), C16 : 1 ω5c (11.6 %), Summed Feature 4 (anteiso-C17 : 1 B/iso-C17 : 1 I) (9.6 %) and anteiso-C15 : 0 (9.6 %). The polar lipid profile consists of the major lipid phosphatidylethanolamine and moderate to minor amounts of phosphatidylserine, unidentified aminolipids, aminophospholipids, aminophosphoglycolipids, polar lipids lacking a functional group and an unidentified phospholipid and a glycolipid. In the polyamine pattern sym-homospermidine is predominant. On the basis of the results obtained, strain 9AT is proposed as a novel species of the genus Hymenobacter, for which the name Hymenobacter caeli sp. nov. is suggested. The type strain is 9AT (=CCM 8971T=LMG 32109T=DSM 111653T).

Frigoriflavimonas asaccharolytica gen. nov., sp. nov., a novel psychrophilic esterase and protease producing bacterium isolated from Antarctica.

The rod-shaped and Gram-stain-negative bacterial strain 16FT, isolated from an air sample collected at King George Island, maritime Antarctica, was investigated to determine its taxonomic status. Strain 16FT is strictly aerobic, catalase positive, oxidase positive and non-motile. Strain 16FT hydrolyses casein, lecithin, Tween 20, 60 and 80, but not aesculin, gelatin and starch. Growth of strain 16FT is observed at 0-20 °C (optimum 10 °C), pH 5.0-8.0 (optimum pH 6.0), and in the presence of 0-2.0% NaCl (optimum 0.5%). The predominant menaquinone is MK-6, and the major fatty acids comprise anteiso-C15:0 and iso-C15:0. The major polar lipids are phosphatidylethanolamine, ornithine lipid OL2, unidentified phospholipid PL1 and the unidentified lipids L3 and L6 lacking functional groups. The DNA G + C content based on the draft genome sequence is 32.3 mol%. Sequence analysis of the 16S rRNA gene indicates the highest similarity to Kaistella palustris 3A10T (95.4%), Kaistella chaponensis Sa 1147-06 T (95.2%), Kaistella antarctica AT1013T (95.1%), Kaistella carnis NCTC 13525 T (95.1%) and below 95.0% to other species with validly published names. Phylogenetic analysis based on 16S rRNA gene and whole-genome sequences places strain 16FT in a distinct branch, indicating a separate lineage within the family Weeksellaceae. Based on the data from our polyphasic approach, 16FT represents a novel species of a new genus, for which the name Frigoriflavimonas asaccharolytica gen. nov, sp. nov. is proposed. The type strain is 16FT (= CCM 8975 T = CGMCC No.1.16844 T).

IMG/M v.5.0: an integrated data management and comparative analysis system for microbial genomes and microbiomes.

The Integrated Microbial Genomes & Microbiomes system v.5.0 (IMG/M: https://img.jgi.doe.gov/m/) contains annotated datasets categorized into: archaea, bacteria, eukarya, plasmids, viruses, genome fragments, metagenomes, cell enrichments, single particle sorts, and metatranscriptomes. Source datasets include those generated by the DOE's Joint Genome Institute (JGI), submitted by external scientists, or collected from public sequence data archives such as NCBI. All submissions are typically processed through the IMG annotation pipeline and then loaded into the IMG data warehouse. IMG's web user interface provides a variety of analytical and visualization tools for comparative analysis of isolate genomes and metagenomes in IMG. IMG/M allows open access to all public genomes in the IMG data warehouse, while its expert review (ER) system (IMG/MER: https://img.jgi.doe.gov/mer/) allows registered users to access their private genomes and to store their private datasets in workspace for sharing and for further analysis. IMG/M data content has grown by 60% since the last report published in the 2017 NAR Database Issue. IMG/M v.5.0 has a new and more powerful genome search feature, new statistical tools, and supports metagenome binning.

Novel heavy metal resistance gene clusters are present in the genome of Cupriavidus neocaledonicus STM 6070, a new species of Mimosa pudica microsymbiont isolated from heavy-metal-rich mining site soil.

BACKGROUND: Cupriavidus strain STM 6070 was isolated from nickel-rich soil collected near Koniambo massif, New Caledonia, using the invasive legume trap host Mimosa pudica. STM 6070 is a heavy metal-tolerant strain that is highly effective at fixing nitrogen with M. pudica. Here we have provided an updated taxonomy for STM 6070 and described salient features of the annotated genome, focusing on heavy metal resistance (HMR) loci and heavy metal efflux (HME) systems. RESULTS: The 6,771,773 bp high-quality-draft genome consists of 107 scaffolds containing 6118 protein-coding genes. ANI values show that STM 6070 is a new species of Cupriavidus. The STM 6070 symbiotic region was syntenic with that of the M. pudica-nodulating Cupriavidus taiwanensis LMG 19424T. In contrast to the nickel and zinc sensitivity of C. taiwanensis strains, STM 6070 grew at high Ni2+ and Zn2+ concentrations. The STM 6070 genome contains 55 genes, located in 12 clusters, that encode HMR structural proteins belonging to the RND, MFS, CHR, ARC3, CDF and P-ATPase protein superfamilies. These HMR molecular determinants are putatively involved in arsenic (ars), chromium (chr), cobalt-zinc-cadmium (czc), copper (cop, cup), nickel (nie and nre), and silver and/or copper (sil) resistance. Seven of these HMR clusters were common to symbiotic and non-symbiotic Cupriavidus species, while four clusters were specific to STM 6070, with three of these being associated with insertion sequences. Within the specific STM 6070 HMR clusters, three novel HME-RND systems (nieIC cep nieBA, czcC2B2A2, and hmxB zneAC zneR hmxS) were identified, which constitute new candidate genes for nickel and zinc resistance. CONCLUSIONS: STM 6070 belongs to a new Cupriavidus species, for which we have proposed the name Cupriavidus neocaledonicus sp. nov.. STM6070 harbours a pSym with a high degree of gene conservation to the pSyms of M. pudica-nodulating C. taiwanensis strains, probably as a result of recent horizontal transfer. The presence of specific HMR clusters, associated with transposase genes, suggests that the selection pressure of the New Caledonian ultramafic soils has driven the specific adaptation of STM 6070 to heavy-metal-rich soils via horizontal gene transfer.

Ecological and genomic analyses of candidate phylum WPS-2 bacteria in an unvegetated soil.

Members of the bacterial candidate phylum WPS-2 (or Eremiobacterota) are abundant in several dry, bare soil environments. In a bare soil deposited by an extinct iron-sulfur spring, we found that WPS-2 comprised up to 24% of the bacterial community and up to 108 cells per g of soil based on 16S rRNA gene sequencing and quantification. A single genus-level cluster (Ca. Rubrimentiphilum) predominated in bare soils but was less abundant in adjacent forest. Nearly complete genomes of Ca. Rubrimentiphilum were recovered as single amplified genomes (SAGs) and metagenome-assembled genomes (MAGs). Surprisingly, given the abundance of WPS-2 in bare soils, the genomes did not indicate any capacity for autotrophy, phototrophy, or trace gas metabolism. Instead, they suggest a predominantly aerobic organoheterotrophic lifestyle, perhaps based on scavenging amino acids, nucleotides, and complex oligopeptides, along with lithotrophic capacity on thiosulfate. Network analyses of the entire community showed that some species of Chloroflexi, Actinobacteria, and candidate phylum AD3 (or Dormibacterota) co-occurred with Ca. Rubrimentiphilum and may represent ecological or metabolic partners. We propose that Ca. Rubrimentiphilum act as efficient heterotrophic scavengers. Combined with previous studies, these data suggest that the phylum WPS-2 includes bacteria with diverse metabolic capabilities.