Characterization of the Radiation Desiccation Response Regulon of the Radioresistant Bacterium Deinococcus radiodurans by Integrative Genomic Analyses.

Numerous genes are overexpressed in the radioresistant bacterium Deinococcus radiodurans after exposure to radiation or prolonged desiccation. It was shown that the DdrO and IrrE proteins play a major role in regulating the expression of approximately twenty genes. The transcriptional repressor DdrO blocks the expression of these genes under normal growth conditions. After exposure to genotoxic agents, the IrrE metalloprotease cleaves DdrO and relieves gene repression. At present, many questions remain, such as the number of genes regulated by DdrO. Here, we present the first ChIP-seq analysis performed at the genome level in Deinococcus species coupled with RNA-seq, which was achieved in the presence or not of DdrO. We also resequenced our laboratory stock strain of D. radiodurans R1 ATCC 13939 to obtain an accurate reference for read alignments and gene expression quantifications. We highlighted genes that are directly under the control of this transcriptional repressor and showed that the DdrO regulon in D. radiodurans includes numerous other genes than those previously described, including DNA and RNA metabolism proteins. These results thus pave the way to better understand the radioresistance pathways encoded by this bacterium and to compare the stress-induced responses mediated by this pair of proteins in diverse bacteria.

Reductive evolution and unique predatory mode in the CPR bacterium Vampirococcus lugosii.

The Candidate Phyla Radiation (CPR) constitutes a large group of mostly uncultured bacterial lineages with small cell sizes and limited biosynthetic capabilities. They are thought to be symbionts of other organisms, but the nature of this symbiosis has been ascertained only for cultured Saccharibacteria, which are epibiotic parasites of other bacteria. Here, we study the biology and the genome of Vampirococcus lugosii, which becomes the first described species of Vampirococcus, a genus of epibiotic bacteria morphologically identified decades ago. Vampirococcus belongs to the CPR phylum Absconditabacteria. It feeds on anoxygenic photosynthetic gammaproteobacteria, fully absorbing their cytoplasmic content. The cells divide epibiotically, forming multicellular stalks whose apical cells can reach new hosts. The genome is small (1.3 Mbp) and highly reduced in biosynthetic metabolism genes, but is enriched in genes possibly related to a fibrous cell surface likely involved in interactions with the host. Gene loss has been continuous during the evolution of Absconditabacteria, and generally most CPR bacteria, but this has been compensated by gene acquisition by horizontal gene transfer and de novo evolution. Our findings support parasitism as a widespread lifestyle of CPR bacteria, which probably contribute to the control of bacterial populations in diverse ecosystems.

Lysine-specific acetylated proteome from the archaeon Thermococcus gammatolerans reveals the presence of acetylated histones.

Thermococcus gammatolerans EJ3 is an extremophile archaeon which was revealed as one of the most radioresistant organisms known on Earth, withstanding up to 30 kGy gamma-ray radiations. While its theoretical proteome is rather small, T. gammatolerans may enhance its toolbox by post-translational modification of its proteins. Here, we explored its extent of Nε-acetylation of lysines. For this, we immunopurified with two acetylated-lysine antibodies the acetylated peptides resulting from a proteolysis of soluble proteins with trypsin. The comparison of acetylated proteomes of two archaea highlights some common acetylation patterns but only 4 out of 26 orthologous proteins found to be acetylated in both species, are acetylated on the same lysine site. We evidenced that histone B is acetylated in T. gammatolerans at least at two different sites (K27 and K36), and a peptide common at the C-terminus of histones A and B is also acetylated. We verified that acetylation of histones is a common trait among Thermococcales after recording data on Thermococcus kodakaraensis histones and identifying three acetylated sites. This discovery reinforces the strong evolutionary link between Archaea and Eukaryotes and should be an incentive for further investigation on the extent and role of acetylation of histones in Archaea. SIGNIFICANCE: Acetylation is an important post-translational modification of proteins that has been extensively described in Eukaryotes, and more recently in Bacteria. Here, we report for the first time ever that histones in Archaea are also modified by acetylation after a systematic survey of acetylated peptides in Thermococcus gammatolerans. Structural models of histones A and B indicates that acetylation of the identified modified residues may play an important role in histone assembly and/or interaction with DNA. The in-depth protein acetylome landscape in T. gammatolerans includes at least 181 unique protein sequences, some of them being modified on numerous residues. Proteins involved in metabolic processes, information storage and processing mechanisms are over-represented categories in this dataset, highlighting the ancient role of this protein post-translational modification in primitive cells.

A Novel Microbialite-Associated Phototrophic Chloroflexi Lineage Exhibiting a Quasi-Clonal Pattern along Depth.

Chloroflexales (Chloroflexi) are typical members of the anoxygenic photosynthesizing component of microbial mats and have mostly been characterized from communities associated to hot springs. Here, we report the assembly of five metagenome-assembled genomes (MAGs) of a novel lineage of Chloroflexales found in mesophilic lithifying microbial mats (microbialites) in Lake Alchichica (Mexico). Genomic and phylogenetic analyses revealed that the bins shared 92% of their genes, and these genes were nearly identical despite being assembled from samples collected along a depth gradient (1-15 m depth). We tentatively name this lineage Candidatus Lithoflexus mexicanus. Metabolic predictions based on the MAGs suggest that these chlorosome-lacking mixotrophs share features in central carbon metabolism, electron transport, and adaptations to life under oxic and anoxic conditions, with members of two related lineages, Chloroflexineae and Roseiflexineae. Contrasting with the other diverse microbialite community members, which display much lower genomic conservation along the depth gradient, Ca. L. mexicanus MAGs exhibit remarkable similarity. This might reflect a particular flexibility to acclimate to varying light conditions with depth or the capacity to occupy a very specific spatial ecological niche in microbialites from different depths. Alternatively, Ca. L. mexicanus may also have the ability to modulate its gene expression as a function of the local environmental conditions during diel cycles in microbialites along the depth gradient.

Functional shifts in microbial mats recapitulate early Earth metabolic transitions.

Phototrophic microbial mats dominated terrestrial ecosystems for billions of years, largely causing, through cyanobacterial oxygenic photosynthesis, but also undergoing, the Great Oxidation Event approximately 2.5 billion years ago. Taking a space-for-time approach based on the universality of core metabolic pathways expressed at ecosystem level, we studied gene content and co-occurrence networks in high-diversity metagenomes from spatially close microbial mats along a steep redox gradient. The observed functional shifts suggest that anoxygenic photosynthesis was present but not predominant under early Precambrian conditions, being accompanied by other autotrophic processes. Our data also suggest that, in contrast to general assumptions, anoxygenic photosynthesis largely expanded in parallel with the subsequent evolution of oxygenic photosynthesis and aerobic respiration. Finally, our observations might represent space-for-time evidence that the Wood-Ljungdahl carbon fixation pathway dominated phototrophic mats in early ecosystems, whereas the Calvin cycle probably evolved from pre-existing variants before becoming the dominant contemporary form of carbon fixation.

High reactivity of deep biota under anthropogenic CO2 injection into basalt.

Basalts are recognized as one of the major habitats on Earth, harboring diverse and active microbial populations. Inconsistently, this living component is rarely considered in engineering operations carried out in these environments. This includes carbon capture and storage (CCS) technologies that seek to offset anthropogenic CO2 emissions into the atmosphere by burying this greenhouse gas in the subsurface. Here, we show that deep ecosystems respond quickly to field operations associated with CO2 injections based on a microbiological survey of a basaltic CCS site. Acidic CO2-charged groundwater results in a marked decrease (by ~ 2.5-4) in microbial richness despite observable blooms of lithoautotrophic iron-oxidizing Betaproteobacteria and degraders of aromatic compounds, which hence impact the aquifer redox state and the carbon fate. Host-basalt dissolution releases nutrients and energy sources, which sustain the growth of autotrophic and heterotrophic species whose activities may have consequences on mineral storage.

An Early-Branching Freshwater Cyanobacterium at the Origin of Plastids.

Photosynthesis evolved in eukaryotes by the endosymbiosis of a cyanobacterium, the future plastid, within a heterotrophic host. This primary endosymbiosis occurred in the ancestor of Archaeplastida, a eukaryotic supergroup that includes glaucophytes, red algae, green algae, and land plants [1-4]. However, although the endosymbiotic origin of plastids from a single cyanobacterial ancestor is firmly established, the nature of that ancestor remains controversial: plastids have been proposed to derive from either early- or late-branching cyanobacterial lineages [5-11]. To solve this issue, we carried out phylogenomic and supernetwork analyses of the most comprehensive dataset analyzed so far including plastid-encoded proteins and nucleus-encoded proteins of plastid origin resulting from endosymbiotic gene transfer (EGT) of primary photosynthetic eukaryotes, as well as wide-ranging genome data from cyanobacteria, including novel lineages. Our analyses strongly support that plastids evolved from deep-branching cyanobacteria and that the present-day closest cultured relative of primary plastids is Gloeomargarita lithophora. This species belongs to a recently discovered cyanobacterial lineage widespread in freshwater microbialites and microbial mats [12, 13]. The ecological distribution of this lineage sheds new light on the environmental conditions where the emergence of photosynthetic eukaryotes occurred, most likely in a terrestrial-freshwater setting. The fact that glaucophytes, the first archaeplastid lineage to diverge, are exclusively found in freshwater ecosystems reinforces this hypothesis. Therefore, not only did plastids emerge early within cyanobacteria, but the first photosynthetic eukaryotes most likely evolved in terrestrial-freshwater settings, not in oceans as commonly thought.

Description of Gloeomargarita lithophora gen. nov., sp. nov., a thylakoid-bearing, basal-branching cyanobacterium with intracellular carbonates, and proposal for Gloeomargaritales ord. nov.

A unicellular cyanobacterium, strain Alchichica-D10, was isolated from microbialites of the alkaline Lake Alchichica, Mexico. The cells were short rods (3.9±0.6 µm in length and 1.1±0.1 µm in width) forming biofilms of intense emerald green colour. They exhibited red autofluorescence under UV light excitation. UV-visible absorption spectra revealed that they contain chlorophyll a and phycocyanin, and electron microscopy showed the presence of thylakoids. The strain grew within a temperature range of 15-30 °C. Genomic DNA G+C content was 52.2 mol%. The most remarkable feature of this species was its granular cytoplasm, due to the presence of numerous intracellular spherical granules (16-26 per cell) with an average diameter of 270 nm. These granules, easily visible under scanning electron microscopy, were composed of amorphous carbonate containing Ca, Mg, Ba and Sr. A multi-gene phylogeny based on the analysis of 59 conserved protein markers supported robustly that this strain occupies a deep position in the cyanobacterial tree. Based on its phenotypic characters and phylogenetic position, strain Alchichica-D10 is considered to represent a new genus and novel species of cyanobacteria for which the name Gloeomargarita lithophora gen. nov., sp. nov. is proposed. The type strain is Alchichica-D10 (Culture Collection of Algae and Protozoa CCAP strain 1437/1; Collections de Cyanobactéries et Microalgues Vivantes of the Museum National d'Histoire Naturelle in Paris strain PMC 919.15). Furthermore, a new family, Gloeomargaritaceae, and a new order, Gloeoemargaritales, are proposed to accommodate this species under the International Code of Nomenclature for algae, fungi and plants.

Comparative metagenomics unveils functions and genome features of microbialite-associated communities along a depth gradient.

Modern microbialites are often used as analogs of Precambrian stromatolites; therefore, studying the metabolic interplay within their associated microbial communities can help formulating hypotheses on their formation and long-term preservation within the fossil record. We performed a comparative metagenomic analysis of microbialite samples collected at two sites and along a depth gradient in Lake Alchichica (Mexico). The community structure inferred from single-copy gene family identification and long-contig (>10 kb) assignation, consistently with previous rRNA gene surveys, showed a wide prokaryotic diversity dominated by Alphaproteobacteria, Gammaproteobacteria, Cyanobacteria, and Bacteroidetes, while eukaryotes were largely dominated by green algae or diatoms. Functional analyses based on RefSeq, COG and SEED assignations revealed the importance of housekeeping functions, with an overrepresentation of genes involved in carbohydrate metabolism, as compared with other metabolic capacities. The search for genes diagnostic of specific metabolic functions revealed the important involvement of Alphaproteobacteria in anoxygenic photosynthesis and sulfide oxidation, and Cyanobacteria in oxygenic photosynthesis and nitrogen fixation. Surprisingly, sulfate reduction appeared negligible. Comparative analyses suggested functional similarities among various microbial mat and microbialite metagenomes as compared with soil or oceans, but showed differences in microbial processes among microbialite types linked to local environmental conditions.

Metagenome-based diversity analyses suggest a significant contribution of non-cyanobacterial lineages to carbonate precipitation in modern microbialites.

Cyanobacteria are thought to play a key role in carbonate formation due to their metabolic activity, but other organisms carrying out oxygenic photosynthesis (photosynthetic eukaryotes) or other metabolisms (e.g., anoxygenic photosynthesis, sulfate reduction), may also contribute to carbonate formation. To obtain more quantitative information than that provided by more classical PCR-dependent methods, we studied the microbial diversity of microbialites from the Alchichica crater lake (Mexico) by mining for 16S/18S rRNA genes in metagenomes obtained by direct sequencing of environmental DNA. We studied samples collected at the Western (AL-W) and Northern (AL-N) shores of the lake and, at the latter site, along a depth gradient (1, 5, 10, and 15 m depth). The associated microbial communities were mainly composed of bacteria, most of which seemed heterotrophic, whereas archaea were negligible. Eukaryotes composed a relatively minor fraction dominated by photosynthetic lineages, diatoms in AL-W, influenced by Si-rich seepage waters, and green algae in AL-N samples. Members of the Gammaproteobacteria and Alphaproteobacteria classes of Proteobacteria, Cyanobacteria, and Bacteroidetes were the most abundant bacterial taxa, followed by Planctomycetes, Deltaproteobacteria (Proteobacteria), Verrucomicrobia, Actinobacteria, Firmicutes, and Chloroflexi. Community composition varied among sites and with depth. Although cyanobacteria were the most important bacterial group contributing to the carbonate precipitation potential, photosynthetic eukaryotes, anoxygenic photosynthesizers and sulfate reducers were also very abundant. Cyanobacteria affiliated to Pleurocapsales largely increased with depth. Scanning electron microscopy (SEM) observations showed considerable areas of aragonite-encrusted Pleurocapsa-like cyanobacteria at microscale. Multivariate statistical analyses showed a strong positive correlation of Pleurocapsales and Chroococcales with aragonite formation at macroscale, and suggest a potential causal link. Despite the previous identification of intracellularly calcifying cyanobacteria in Alchichica microbialites, most carbonate precipitation seems extracellular in this system.

Bacterial gene import and mesophilic adaptation in archaea.

It is widely believed that the archaeal ancestor was hyperthermophilic, but during archaeal evolution, several lineages - including haloarchaea and their sister methanogens, the Thaumarchaeota, and the uncultured Marine Group II and Marine Group III Euryarchaeota (MGII/III) - independently adapted to lower temperatures. Recent phylogenomic studies suggest that the ancestors of these lineages were recipients of massive horizontal gene transfer from bacteria. Many of the acquired genes, which are often involved in metabolism and cell envelope biogenesis, were convergently acquired by distant mesophilic archaea. In this Opinion article, we explore the intriguing hypothesis that the import of these bacterial genes was crucial for the adaptation of archaea to mesophilic lifestyles.

Diversity and comparative genomics of Microviridae in Sphagnum- dominated peatlands.

Microviridae, a family of bacteria-infecting ssDNA viruses, is one of the still poorly characterized bacteriophage groups, even though it includes phage PhiX174, one of the main models in virology for genomic and capsid structure studies. Recent studies suggest that they are diverse and well represented in marine and freshwater virioplankton as well as in human microbiomes. However, their diversity, abundance, and ecological role are completely unknown in soil ecosystems. Here we present the comparative analysis of 17 completely assembled Microviridae genomes from 12 viromes of a Sphagnum-dominated peatland. Phylogenetic analysis of the conserved major capsid protein sequences revealed the affiliation to Gokushovirinae and Pichovirinae as well as to two newly defined subfamilies, the Aravirinae and Stokavirinae. Additionally, two new distinct prophages were identified in the genomes of Parabacteroides merdae and Parabacteroides distasonis representing a potential new subfamily of Microviridae. The differentiation of the subfamilies was confirmed by gene order and similarity analysis. Relative abundance analysis using the affiliation of the major capsid protein (VP1) revealed that Gokushovirinae, followed by Aravirinae, are the most abundant Microviridae in 11 out of 12 peat viromes. Sequences matching the Gokushovirinae and Aravirinae VP1 matching sequences, respectively, accounted for up to 4.19 and 0.65% of the total number of sequences in the corresponding virome, respectively. In this study we provide new genome information of Microviridae and pave the way toward quantitative estimations of Microviridae subfamilies.

Pangenome evidence for extensive interdomain horizontal transfer affecting lineage core and shell genes in uncultured planktonic thaumarchaeota and euryarchaeota.

Horizontal gene transfer (HGT) is an important force in evolution, which may lead, among other things, to the adaptation to new environments by the import of new metabolic functions. Recent studies based on phylogenetic analyses of a few genome fragments containing archaeal 16S rRNA genes and fosmid-end sequences from deep-sea metagenomic libraries have suggested that marine planktonic archaea could be affected by high HGT frequency. Likewise, a composite genome of an uncultured marine euryarchaeote showed high levels of gene sequence similarity to bacterial genes. In this work, we ask whether HGT is frequent and widespread in genomes of these marine archaea, and whether HGT is an ancient and/or recurrent phenomenon. To answer these questions, we sequenced 997 fosmid archaeal clones from metagenomic libraries of deep-Mediterranean waters (1,000 and 3,000 m depth) and built comprehensive pangenomes for planktonic Thaumarchaeota (Group I archaea) and Euryarchaeota belonging to the uncultured Groups II and III Euryarchaeota (GII/III-Euryarchaeota). Comparison with available reference genomes of Thaumarchaeota and a composite marine surface euryarchaeote genome allowed us to define sets of core, lineage-specific core, and shell gene ortholog clusters for the two archaeal lineages. Molecular phylogenetic analyses of all gene clusters showed that 23.9% of marine Thaumarchaeota genes and 29.7% of GII/III-Euryarchaeota genes had been horizontally acquired from bacteria. HGT is not only extensive and directional but also ongoing, with high HGT levels in lineage-specific core (ancient transfers) and shell (recent transfers) genes. Many of the acquired genes are related to metabolism and membrane biogenesis, suggesting an adaptive value for life in cold, oligotrophic oceans. We hypothesize that the acquisition of an important amount of foreign genes by the ancestors of these archaeal groups significantly contributed to their divergence and ecological success.

Insights into bacteriophage T5 structure from analysis of its morphogenesis genes and protein components.

Bacteriophage T5 represents a large family of lytic Siphoviridae infecting Gram-negative bacteria. The low-resolution structure of T5 showed the T=13 geometry of the capsid and the unusual trimeric organization of the tail tube, and the assembly pathway of the capsid was established. Although major structural proteins of T5 have been identified in these studies, most of the genes encoding the morphogenesis proteins remained to be identified. Here, we combine a proteomic analysis of T5 particles with a bioinformatic study and electron microscopic immunolocalization to assign function to the genes encoding the structural proteins, the packaging proteins, and other nonstructural components required for T5 assembly. A head maturation protease that likely accounts for the cleavage of the different capsid proteins is identified. Two other proteins involved in capsid maturation add originality to the T5 capsid assembly mechanism: the single head-to-tail joining protein, which closes the T5 capsid after DNA packaging, and the nicking endonuclease responsible for the single-strand interruptions in the T5 genome. We localize most of the tail proteins that were hitherto uncharacterized and provide a detailed description of the tail tip composition. Our findings highlight novel variations of viral assembly strategies and of virion particle architecture. They further recommend T5 for exploring phage structure and assembly and for deciphering conformational rearrangements that accompany DNA transfer from the capsid to the host cytoplasm.

Genome-wide transcriptional response of the archaeon Thermococcus gammatolerans to cadmium.

Thermococcus gammatolerans, the most radioresistant archaeon known to date, is an anaerobic and hyperthermophilic sulfur-reducing organism living in deep-sea hydrothermal vents. Knowledge of mechanisms underlying archaeal metal tolerance in such metal-rich ecosystem is still poorly documented. We showed that T. gammatolerans exhibits high resistance to cadmium (Cd), cobalt (Co) and zinc (Zn), a weaker tolerance to nickel (Ni), copper (Cu) and arsenate (AsO(4)) and that cells exposed to 1 mM Cd exhibit a cellular Cd concentration of 67 µM. A time-dependent transcriptomic analysis using microarrays was performed at a non-toxic (100 µM) and a toxic (1 mM) Cd dose. The reliability of microarray data was strengthened by real time RT-PCR validations. Altogether, 114 Cd responsive genes were revealed and a substantial subset of genes is related to metal homeostasis, drug detoxification, re-oxidization of cofactors and ATP production. This first genome-wide expression profiling study of archaeal cells challenged with Cd showed that T. gammatolerans withstands induced stress through pathways observed in both prokaryotes and eukaryotes but also through new and original strategies. T. gammatolerans cells challenged with 1 mM Cd basically promote: 1) the induction of several transporter/permease encoding genes, probably to detoxify the cell; 2) the upregulation of Fe transporters encoding genes to likely compensate Cd damages in iron-containing proteins; 3) the induction of membrane-bound hydrogenase (Mbh) and membrane-bound hydrogenlyase (Mhy2) subunits encoding genes involved in recycling reduced cofactors and/or in proton translocation for energy production. By contrast to other organisms, redox homeostasis genes appear constitutively expressed and only a few genes encoding DNA repair proteins are regulated. We compared the expression of 27 Cd responsive genes in other stress conditions (Zn, Ni, heat shock, γ-rays), and showed that the Cd transcriptional pattern is comparable to other metal stress transcriptional responses (Cd, Zn, Ni) but not to a general stress response.

Complete-fosmid and fosmid-end sequences reveal frequent horizontal gene transfers in marine uncultured planktonic archaea.

The extent of horizontal gene transfer (HGT) among marine pelagic prokaryotes and the role that HGT may have played in their adaptation to this particular environment remain open questions. This is partly due to the paucity of cultured species and genomic information for many widespread groups of marine bacteria and archaea. Molecular studies have revealed a large diversity and relative abundance of marine planktonic archaea, in particular of Thaumarchaeota (also known as group I Crenarchaeota) and Euryarchaeota of groups II and III, but only one species (the thaumarchaeote Candidatus Nitrosopumilus maritimus) has been isolated in pure culture so far. Therefore, metagenomics remains the most powerful approach to study these environmental groups. To investigate the impact of HGT in marine archaea, we carried out detailed phylogenetic analyses of all open reading frames of 21 archaeal 16S rRNA gene-containing fosmids and, to extend our analysis to other genomic regions, also of fosmid-end sequences of 12 774 fosmids from three different deep-sea locations (South Atlantic and Adriatic Sea at 1000 m depth, and Ionian Sea at 3000 m depth). We found high HGT rates in both marine planktonic Thaumarchaeota and Euryarchaeota, with remarkable converging values estimated from complete-fosmid and fosmid-end sequence analysis (25 and 21% of the genes, respectively). Most HGTs came from bacterial donors (mainly from Proteobacteria, Firmicutes and Chloroflexi) but also from other archaea and eukaryotes. Phylogenetic analyses showed that in most cases HGTs are shared by several representatives of the studied groups, implying that they are ancient and have been conserved over relatively long evolutionary periods. This, together with the functions carried out by these acquired genes (mostly related to energy metabolism and transport of metabolites across membranes), suggests that HGT has played an important role in the adaptation of these archaea to the cold and nutrient-depleted deep marine environment.

Comparative metagenomics of bathypelagic plankton and bottom sediment from the Sea of Marmara.

To extend comparative metagenomic analyses of the deep-sea, we produced metagenomic data by direct 454 pyrosequencing from bathypelagic plankton (1000 m depth) and bottom sediment of the Sea of Marmara, the gateway between the Eastern Mediterranean and the Black Seas. Data from small subunit ribosomal RNA (SSU rRNA) gene libraries and direct pyrosequencing of the same samples indicated that Gamma- and Alpha-proteobacteria, followed by Bacteroidetes, dominated the bacterial fraction in Marmara deep-sea plankton, whereas Planctomycetes, Delta- and Gamma-proteobacteria were the most abundant groups in high bacterial-diversity sediment. Group I Crenarchaeota/Thaumarchaeota dominated the archaeal plankton fraction, although group II and III Euryarchaeota were also present. Eukaryotes were highly diverse in SSU rRNA gene libraries, with group I (Duboscquellida) and II (Syndiniales) alveolates and Radiozoa dominating plankton, and Opisthokonta and Alveolates, sediment. However, eukaryotic sequences were scarce in pyrosequence data. Archaeal amo genes were abundant in plankton, suggesting that Marmara planktonic Thaumarchaeota are ammonia oxidizers. Genes involved in sulfate reduction, carbon monoxide oxidation, anammox and sulfatases were over-represented in sediment. Genome recruitment analyses showed that Alteromonas macleodii 'surface ecotype', Pelagibacter ubique and Nitrosopumilus maritimus were highly represented in 1000 m-deep plankton. A comparative analysis of Marmara metagenomes with ALOHA deep-sea and surface plankton, whale carcasses, Peru subsurface sediment and soil metagenomes clustered deep-sea Marmara plankton with deep-ALOHA plankton and whale carcasses, likely because of the suboxic conditions in the deep Marmara water column. The Marmara sediment clustered with the soil metagenome, highlighting the common ecological role of both types of microbial communities in the degradation of organic matter and the completion of biogeochemical cycles.

Genome analysis and genome-wide proteomics of Thermococcus gammatolerans, the most radioresistant organism known amongst the Archaea.

BACKGROUND: Thermococcus gammatolerans was isolated from samples collected from hydrothermal chimneys. It is one of the most radioresistant organisms known amongst the Archaea. We report the determination and annotation of its complete genome sequence, its comparison with other Thermococcales genomes, and a proteomic analysis. RESULTS: T. gammatolerans has a circular chromosome of 2.045 Mbp without any extra-chromosomal elements, coding for 2,157 proteins. A thorough comparative genomics analysis revealed important but unsuspected genome plasticity differences between sequenced Thermococcus and Pyrococcus species that could not be attributed to the presence of specific mobile elements. Two virus-related regions, tgv1 and tgv2, are the only mobile elements identified in this genome. A proteogenome analysis was performed by a shotgun liquid chromatography-tandem mass spectrometry approach, allowing the identification of 10,931 unique peptides corresponding to 951 proteins. This information concurrently validates the accuracy of the genome annotation. Semi-quantification of proteins by spectral count was done on exponential- and stationary-phase cells. Insights into general catabolism, hydrogenase complexes, detoxification systems, and the DNA repair toolbox of this archaeon are revealed through this genome and proteome analysis. CONCLUSIONS: This work is the first archaeal proteome investigation done at the stage of primary genome annotation. This archaeon is shown to use a large variety of metabolic pathways even under a rich medium growth condition. This proteogenomic study also indicates that the high radiotolerance of T. gammatolerans is probably due to proteins that remain to be characterized rather than a larger arsenal of known DNA repair enzymes.

Comparative analysis of genome fragments of Acidobacteria from deep Mediterranean plankton.

Acidobacteria constitute a still poorly studied phylum that is well represented in soils. Recent studies suggest that members of this phylum may be also abundant in deep-sea plankton, but their relative abundance and ecological role in this ecosystem are completely unknown. A recent screening of three metagenomic deep-sea libraries of bathypelagic plankton from the South Atlantic (1000 m depth), the Adriatic (1000 m depth) and the Ionian (3000 m depth) seas in the Mediterranean revealed an unexpected relative proportion of acidobacterial fosmids, which affiliated to the Solibacterales (Group 3), to the Group 11 and, most frequently, to the Group 6 of this diverse phylum. Here, we present the comparative analysis of 11 acidobacterial genome fragments containing the rrn operon from these Mediterranean libraries. A highly conserved syntenic region spanning up to 30 kb and containing up to 25 open reading frames was shared by Group 6 Acidobacteria. Synteny was also partially conserved in distantly related acidobacterial genome fragments derived from a metagenomic soil library, indicating a remarkable conservation of this genomic region within these Acidobacteria. A search for Acidobacteria-specific hits in directly comparable, available fosmid-end sequences from soil and marine metagenomic libraries showed a significant increase of their relative proportion in plankton libraries as a function of increasing depth reaching, at high depth, levels nearly comparable to those of soil. Thus, our results suggest that Acidobacteria are abundant and represent a significant proportion of the microbial community in the deep-sea ecosystem.

Nanoarchaea: representatives of a novel archaeal phylum or a fast-evolving euryarchaeal lineage related to Thermococcales?

BACKGROUND: Cultivable archaeal species are assigned to two phyla -- the Crenarchaeota and the Euryarchaeota -- by a number of important genetic differences, and this ancient split is strongly supported by phylogenetic analysis. The recently described hyperthermophile Nanoarchaeum equitans, harboring the smallest cellular genome ever sequenced (480 kb), has been suggested as the representative of a new phylum -- the Nanoarchaeota -- that would have diverged before the Crenarchaeota/Euryarchaeota split. Confirming the phylogenetic position of N. equitans is thus crucial for deciphering the history of the archaeal domain. RESULTS: We tested the placement of N. equitans in the archaeal phylogeny using a large dataset of concatenated ribosomal proteins from 25 archaeal genomes. We indicate that the placement of N. equitans in archaeal phylogenies on the basis of ribosomal protein concatenation may be strongly biased by the coupled effect of its above-average evolutionary rate and lateral gene transfers. Indeed, we show that different subsets of ribosomal proteins harbor a conflicting phylogenetic signal for the placement of N. equitans. A BLASTP-based survey of the phylogenetic pattern of all open reading frames (ORFs) in the genome of N. equitans revealed a surprisingly high fraction of close hits with Euryarchaeota, notably Thermococcales. Strikingly, a specific affinity of N. equitans and Thermococcales was strongly supported by phylogenies based on a subset of ribosomal proteins, and on a number of unrelated molecular markers. CONCLUSION: We suggest that N. equitans may more probably be the representative of a fast-evolving euryarchaeal lineage (possibly related to Thermococcales) than the representative of a novel and early diverging archaeal phylum.