Pan-Genome Provides Insights into Vibrio Evolution and Adaptation to Deep-Sea Hydrothermal Vents.

This study delves into the genomic features of 10 Vibrio strains collected from deep-sea hydrothermal vents in the Pacific Ocean, providing insights into their evolutionary history and ecological adaptations. Through sequencing and pan-genome analysis involving 141 Vibrio species, we found that deep-sea strains exhibit larger genomes with unique gene distributions, suggesting adaptation to the vent environment. The phylogenomic reconstruction of the investigated isolates revealed the presence of 2 main clades: The first is monophyletic, consisting exclusively of Vibrio alginolyticus, while the second forms a monophyletic clade comprising both Vibrio antiquarius and Vibrio diabolicus species, which were previously isolated from deep-sea vents. All strains carry virulence and antibiotic resistance genes related to those found in human pathogenic Vibrio species which may play a wider ecological role other than host infection in these environments. In addition, functional genomic analysis identified genes potentially related to deep-sea survival and stress response, alongside candidate genes encoding for novel antimicrobial agents. Ultimately, the pan-genome we generated represents a valuable resource for future studies investigating the taxonomy, evolution, and ecology of Vibrio species.

Microbial eukaryotic predation pressure and biomass at deep-sea hydrothermal vents.

Deep-sea hydrothermal vent geochemistry shapes the foundation of the microbial food web by fueling chemolithoautotrophic microbial activity. Microbial eukaryotes (or protists) play a critical role in hydrothermal vent food webs as consumers and hosts of symbiotic bacteria, and as a nutritional source to higher trophic levels. We measured microbial eukaryotic cell abundance and predation pressure in low-temperature diffuse hydrothermal fluids at the Von Damm and Piccard vent fields along the Mid-Cayman Rise in the Western Caribbean Sea. We present findings from experiments performed under in situ pressure that show cell abundances and grazing rates higher than those done at 1 atmosphere (shipboard ambient pressure); this trend was attributed to the impact of depressurization on cell integrity. A relationship between the protistan grazing rate, prey cell abundance, and temperature of end-member hydrothermal vent fluid was observed at both vent fields, regardless of experimental approach. Our results show substantial protistan biomass at hydrothermally fueled microbial food webs, and when coupled with improved grazing estimates, suggest an important contribution of grazers to the local carbon export and supply of nutrient resources to the deep ocean.

Hydrogenimonas cancrithermarum sp. nov., a hydrogen- and thiosulfate-oxidizing mesophilic chemolithoautotroph isolated from diffuse-flow fluids on the East Pacific Rise, and an emended description of the genus Hydrogenimonas.

A novel mesophilic, hydrogen- and thiosulfate-oxidizing bacterium, strain ISO32T, was isolated from diffuse-flow hydrothermal fluids from the Crab Spa vent on the East Pacific Rise. Cells of ISO32T were rods, being motile by means of a single polar flagellum. The isolate grew at a temperature range between 30 and 55 °C (optimum, 43 °C), at a pH range between 5.3 and 7.6 (optimum, pH 5.8) and in the presence of 2.0-4.0 % NaCl (optimum, 2.5 %). The isolate was able to grow chemolithoautotrophically with molecular hydrogen, thiosulfate or elemental sulfur as the sole electron donor. Thiosulfate, elemental sulfur, nitrate and molecular oxygen were each used as a sole electron acceptor. Phylogenetic analysis of 16S rRNA gene sequences placed ISO32T in the genus Hydrogenimonas of the class Epsilonproteobacteria, with Hydrogenimonas thermophila EP1-55-1 %T as its closest relative (95.95 % similarity). On the basis of the phylogenetic, physiological and genomic characteristics, it is proposed that the organism represents a novel species within the genus Hydrogenimonas, Hydrogenimonas cancrithermarum sp. nov. The type strain is ISO32T (=JCM 39185T =KCTC 25252T). Furthermore, the genomic properties of members of the genus Hydrogenimonas are distinguished from those of members of other thermophilic genera in the orders Campylobacterales (Nitratiruptor and Nitrosophilus) and Nautiliales (Caminibacter, Nautilia and Lebetimonas), with larger genome sizes and lower 16S rRNA G+C content values. Comprehensive metabolic comparisons based on genomes revealed that genes responsible for the Pta-AckA pathway were observed exclusively in members of mesophilic genera in the order Campylobacterales and of the genus Hydrogenimonas. Our results indicate that the genus Hydrogenimonas contributes to elucidating the evolutionary history of Epsilonproteobacteria in terms of metabolism and transition from a thermophilic to a mesophilic lifestyle.

Nitrosophilus kaiyonis sp. nov., a hydrogen-, sulfur- and thiosulfate-oxidizing chemolithoautotroph within "Campylobacteria" isolated from a deep-sea hydrothermal vent in the Mid-Okinawa Trough.

A novel bacterium, strain MOT50T, was isolated from the chimney structure at the Iheya North field in the Mid-Okinawa Trough. The cells were motile short rods with a single polar flagellum. Growth was observed between 40 and 65 â„ƒ (optimum, 52 â„ƒ), at pH values between 5.0 and 7.1 (optimum, pH 6.1) and in the presence of 2.0-4.0% NaCl (optimum, 2.5%). The isolates utilized molecular hydrogen, thiosulfate, or elemental sulfur as the sole electron donor. Thiosulfate, elemental sulfur, nitrate, and molecular oxygen are utilized as the sole electron acceptor. Ammonium is required as a nitrogen source. Thiosulfate, elemental sulfur, sulfate, or sulfite serves as a sulfur source for growth. The G + C content of the genomic DNA was 28.9%. Phylogenetic analysis based on the 16S rRNA gene sequences indicated that strain MOT50T belonged to the genus Nitrosophilus of the class "Campylobacteria", and its closest relative was Nitrosophilus labii HRV44T (97.20%). On the basis of the phylogenetic, physiological, and molecular characteristics, it is proposed that the organism represents a novel species within the genus Nitrosophilus, Nitrosophilus kaiyonis sp. nov. The type strain is MOT50T (= JCM 39187T = KCTC 25251T).

Global patterns of diversity and metabolism of microbial communities in deep-sea hydrothermal vent deposits.

BACKGROUND: When deep-sea hydrothermal fluids mix with cold oxygenated fluids, minerals precipitate out of solution and form hydrothermal deposits. These actively venting deep-sea hydrothermal deposits support a rich diversity of thermophilic microorganisms which are involved in a range of carbon, sulfur, nitrogen, and hydrogen metabolisms. Global patterns of thermophilic microbial diversity in deep-sea hydrothermal ecosystems have illustrated the strong connectivity between geological processes and microbial colonization, but little is known about the genomic diversity and physiological potential of these novel taxa. Here we explore this genomic diversity in 42 metagenomes from four deep-sea hydrothermal vent fields and a deep-sea volcano collected from 2004 to 2018 and document their potential implications in biogeochemical cycles. RESULTS: Our dataset represents 3635 metagenome-assembled genomes encompassing 511 novel and recently identified genera from deep-sea hydrothermal settings. Some of the novel bacterial (107) and archaeal genera (30) that were recently reported from the deep-sea Brothers volcano were also detected at the deep-sea hydrothermal vent fields, while 99 bacterial and 54 archaeal genera were endemic to the deep-sea Brothers volcano deposits. We report some of the first examples of medium- (≥ 50% complete, ≤ 10% contaminated) to high-quality (> 90% complete, < 5% contaminated) MAGs from phyla and families never previously identified, or poorly sampled, from deep-sea hydrothermal environments. We greatly expand the novel diversity of Thermoproteia, Patescibacteria (Candidate Phyla Radiation, CPR), and Chloroflexota found at deep-sea hydrothermal vents and identify a small sampling of two potentially novel phyla, designated JALSQH01 and JALWCF01. Metabolic pathway analysis of metagenomes provides insights into the prevalent carbon, nitrogen, sulfur, and hydrogen metabolic processes across all sites and illustrates sulfur and nitrogen metabolic "handoffs" in community interactions. We confirm that Campylobacteria and Gammaproteobacteria occupy similar ecological guilds but their prevalence in a particular site is driven by shifts in the geochemical environment. CONCLUSION: Our study of globally distributed hydrothermal vent deposits provides a significant expansion of microbial genomic diversity associated with hydrothermal vent deposits and highlights the metabolic adaptation of taxonomic guilds. Collectively, our results illustrate the importance of comparative biodiversity studies in establishing patterns of shared phylogenetic diversity and physiological ecology, while providing many targets for enrichment and cultivation of novel and endemic taxa. Video Abstract.

Physiological and metabolic responses of chemolithoautotrophic NO 3 - reducers to high hydrostatic pressure.

We investigated the impact of pressure on thermophilic, chemolithoautotrophic NO 3 - reducing bacteria of the phyla Campylobacterota and Aquificota isolated from deep-sea hydrothermal vents. Batch incubations at 5 and 20 MPa resulted in decreased NO 3 - consumption, lower cell concentrations, and overall slower growth in Caminibacter mediatlanticus (Campylobacterota) and Thermovibrio ammonificans (Aquificota), relative to batch incubations near standard pressure (0.2 MPa) conditions. Nitrogen isotope fractionation effects from chemolithoautotrophic NO 3 - reduction by both microorganisms were, on the contrary, maintained under all pressure conditions. Comparable chemolithoautotrophic NO 3 - reducing activities between previously reported natural hydrothermal vent fluid microbial communities dominated by Campylobacterota at 25 MPa and Campylobacterota laboratory isolates at 0.2 MPa, suggest robust similarities in cell-specific NO 3 - reduction rates and doubling times between microbial populations and communities growing maximally under similar temperature conditions. Physiological and metabolic comparisons of our results with other studies of pressure effects on anaerobic chemolithoautotrophic processes (i.e., microbial S0 -oxidation coupled to Fe(III) reduction and hydrogenotrophic methanogenesis) suggest that anaerobic chemolithoautotrophs relying on oxidation-reduction (redox) reactions that yield higher Gibbs energies experience larger shifts in cell-specific respiration rates and doubling times at increased pressures. Overall, our results advance understanding of the role of pressure, its relationship with temperature and redox conditions, and their effects on seafloor chemolithoautotrophic NO 3 - reduction and other anaerobic chemolithoautotrophic processes.

Metagenomics Reveals Dominant Unusual Sulfur Oxidizers Inhabiting Active Hydrothermal Chimneys From the Southwest Indian Ridge.

The deep-sea hydrothermal vents (DSHVs) in the Southwest Indian Ridge (SWIR) are formed by specific geological settings. However, the community structure and ecological function of the microbial inhabitants on the sulfide chimneys of active hydrothermal vents remain largely unknown. In this study, our analyses of 16S rRNA gene amplicons and 16S rRNA metagenomic reads showed the dominance of sulfur-oxidizing Ectothiorhodospiraceae, Thiomicrorhabdus, Sulfurimonas, and Sulfurovum on the wall of two active hydrothermal chimneys. Compared with the inactive hydrothermal sediments of SWIR, the active hydrothermal chimneys lacked sulfur-reducing bacteria. The metabolic potentials of the retrieved 82 metagenome-assembled genomes (MAGs) suggest that sulfur oxidation might be conducted by Thiohalomonadales (classified as Ectothiorhodospiraceae based on 16S rRNA gene amplicons), Sulfurovaceae, Hyphomicrobiaceae, Thiotrichaceae, Thiomicrospiraceae, and Rhodobacteraceae. For CO2 fixation, the Calvin-Benson-Bassham and reductive TCA pathways were employed by these bacteria. In Thiohalomonadales MAGs, we revealed putative phytochrome, carotenoid precursor, and squalene synthesis pathways, indicating a possible capacity of Thiohalomonadales in adaptation to dynamics redox conditions and the utilization of red light from the hot hydrothermal chimneys for photolithotrophic growth. This study, therefore, reveals unique microbiomes and their genomic features in the active hydrothermal chimneys of SWIR, which casts light on ecosystem establishment and development in hydrothermal fields and the deep biosphere.

Diversity of Mixotrophic Neutrophilic Thiosulfate- and Iron-Oxidizing Bacteria from Deep-Sea Hydrothermal Vents.

At deep-sea hydrothermal vents, sulfur oxidation and iron oxidation are of the highest importance to microbial metabolisms, which are thought to contribute mainly in chemolithoautotrophic groups. In this study, 17 mixotrophic neutrophilic thiosulfate- and iron-oxidizing bacteria were isolated from hydrothermal fields on the Carlsberg Ridge in the Indian Ocean, nine to the γ-proteobacteria (Halomonas (4), Pseudomonas (2), Marinobacter (2), and Rheinheimera (1)), seven to the α-proteobacteria (Thalassospira, Qipengyuania, Salipiger, Seohaeicola, Martelella, Citromicrobium, and Aurantimonas), and one to the Actinobacteria (Agromyces), as determined by their 16S rRNA and genome sequences. The physiological characterization of these isolates revealed wide versatility in electron donors (Fe(II) and Mn(II), or thiosulfate) and a variety of lifestyles as lithotrophic or heterotrophic, microaerobic, or anaerobic. As a representative strain, Pseudomonas sp. IOP_13 showed its autotrophic gowth from 105 cells/ml to 107 cells/ml;carbon dioxide fixation capacity with the δ13CVPDB in the biomass increased from -27.42‱ to 3460.06‱; the thiosulfate-oxidizing ability with produced SO42- increased from 60 mg/L to 287 mg/L; and the iron-oxidizing ability with Fe(II) decreased from 10 mM to 5.2 mM. In addition, iron-oxide crust formed outside the cells. Gene coding for energy metabolism involved in possible iron, manganese, and sulfur oxidation, and denitrification was identified by their genome analysis. This study sheds light on the function of the mixotrophic microbial community in the iron/manganese/sulfur cycles and the carbon fixation of the hydrothermal fields.

Genus-Specific Carbon Fixation Activity Measurements Reveal Distinct Responses to Oxygen among Hydrothermal Vent Campylobacteria.

Molecular surveys of low temperature deep-sea hydrothermal vent fluids have shown that Campylobacteria (previously Epsilonproteobacteria) often dominate the microbial community and that three genera, Arcobacter, Sulfurimonas, and Sulfurovum, frequently coexist. In this study, we used replicated radiocarbon incubations of deep-sea hydrothermal fluids to investigate activity of each genus under three experimental conditions. To quantify genus-specific radiocarbon incorporation, we used newly designed oligonucleotide probes for Arcobacter, Sulfurimonas, and Sulfurovum to quantify their activity using catalyzed-reporter deposition fluorescence in situ hybridization (CARD-FISH) combined with fluorescence-activated cell sorting. All three genera actively fixed CO2 in short-term (∼ 20 h) incubations, but responded differently to the additions of nitrate and oxygen. Oxygen additions had the largest effect on community composition, and caused a pronounced shift in community composition at the amplicon sequence variant (ASV) level after only 20 h of incubation. The effect of oxygen on carbon fixation rates appeared to depend on the initial starting community. The presented results support the hypothesis that these chemoautotrophic genera possess functionally redundant core metabolic capabilities, but also reveal finer-scale differences in growth likely reflecting adaptation of physiologically-distinct phylotypes to varying oxygen concentrations in situ. Overall, our study provides new insights into how oxygen controls community composition and total chemoautotrophic activity, and underscores how quickly deep-sea vent microbial communities respond to disturbances. IMPORTANCE Sulfidic environments worldwide are often dominated by sulfur-oxidizing, carbon-fixing Campylobacteria. Environmental factors associated with this group's dominance are now understood, but far less is known about the ecology and physiology of members of subgroups of chemoautotrophic Campylobacteria. In this study, we used a novel method to differentiate the genus-specific chemoautotrophic activity of three subtypes of Campylobacteria. In combination with evidence from microscopic counts, chemical consumption/production during incubations, and DNA-based measurements, our data show that oxygen concentration affects both community composition and chemoautotrophic function in situ. These results help us better understand factors controlling microbial diversity at deep-sea hydrothermal vents, and provide first-order insights into the ecophysiological differences between these distinct microbial taxa.

Protistan grazing impacts microbial communities and carbon cycling at deep-sea hydrothermal vents.

Microbial eukaryotes (or protists) in marine ecosystems are a link between primary producers and all higher trophic levels, and the rate at which heterotrophic protistan grazers consume microbial prey is a key mechanism for carbon transport and recycling in microbial food webs. At deep-sea hydrothermal vents, chemosynthetic bacteria and archaea form the base of a food web that functions in the absence of sunlight, but the role of protistan grazers in these highly productive ecosystems is largely unexplored. Here, we pair grazing experiments with a molecular survey to quantify protistan grazing and to characterize the composition of vent-associated protists in low-temperature diffuse venting fluids from Gorda Ridge in the northeast Pacific Ocean. Results reveal protists exert higher predation pressure at vents compared to the surrounding deep seawater environment and may account for consuming 28 to 62% of the daily stock of prokaryotic biomass within discharging hydrothermal vent fluids. The vent-associated protistan community was more species rich relative to the background deep sea, and patterns in the distribution and co-occurrence of vent microbes provide additional insights into potential predator-prey interactions. Ciliates, followed by dinoflagellates, Syndiniales, rhizaria, and stramenopiles, dominated the vent protistan community and included bacterivorous species, species known to host symbionts, and parasites. Our findings provide an estimate of protistan grazing pressure within hydrothermal vent food webs, highlighting the important role that diverse protistan communities play in deep-sea carbon cycling.

Geothermobacter hydrogeniphilus sp. nov., a mesophilic, iron(III)-reducing bacterium from seafloor/subseafloor environments in the Pacific Ocean, and emended description of the genus Geothermobacter.

A novel mesophilic, anaerobic, mixotrophic bacterium, with designated strains EPR-MT and HR-1, was isolated from a semi-extinct hydrothermal vent at the East Pacific Rise and from an Fe-mat at Lo'ihi Seamount, respectively. The cells were Gram-negative, pleomorphic rods of about 2.0 µm in length and 0.5 µm in width. Strain EPR-MT grew between 25 and 45 °C (optimum, 37.5-40 °C), 10 and 50 g l-1 NaCl (optimum, 15-20 g l-1) and pH 5.5 and 8.6 (optimum, pH 6.4). Strain HR-1 grew between 20 and 45 °C (optimum, 37.5-40 °C), 10 and 50 g l-1 NaCl (optimum, 15-25 g l-1) and pH 5.5 and 8.6 (optimum, pH 6.4). Shortest generation times with H2 as the primary electron donor, CO2 as the carbon source and ferric citrate as terminal electron acceptor were 6.7 and 5.5 h for EPR-MT and HR-1, respectively. Fe(OH)3, MnO2, AsO4 3-, SO4 2-, SeO4 2-, S2O3 2-, S0 and NO3 - were also used as terminal electron acceptors. Acetate, yeast extract, formate, lactate, tryptone and Casamino acids also served as both electron donors and carbon sources. G+C content of the genomic DNA was 59.4 mol% for strain EPR-MT and 59.2 mol% for strain HR-1. Phylogenetic and phylogenomic analyses indicated that both strains were closely related to each other and to Geothermobacter ehrlichii, within the class δ-Proteobacteria (now within the class Desulfuromonadia). Based on phylogenetic and phylogenomic analyses in addition to physiological and biochemical characteristics, both strains were found to represent a novel species within the genus Geothermobacter, for which the name Geothermobacter hydrogeniphilus sp. nov. is proposed. Geothermobacter hydrogeniphilus is represented by type strain EPR-MT (=JCM 32109T=KCTC 15831T=ATCC TSD-173T) and strain HR-1 (=JCM 32110=KCTC 15832).

Microorganisms from deep-sea hydrothermal vents.

With a rich variety of chemical energy sources and steep physical and chemical gradients, hydrothermal vent systems offer a range of habitats to support microbial life. Cultivation-dependent and independent studies have led to an emerging view that diverse microorganisms in deep-sea hydrothermal vents live their chemolithoautotrophic, heterotrophic, or mixotrophic life with versatile metabolic strategies. Biogeochemical processes are mediated by microorganisms, and notably, processes involving or coupling the carbon, sulfur, hydrogen, nitrogen, and metal cycles in these unique ecosystems. Here, we review the taxonomic and physiological diversity of microbial prokaryotic life from cosmopolitan to endemic taxa and emphasize their significant roles in the biogeochemical processes in deep-sea hydrothermal vents. According to the physiology of the targeted taxa and their needs inferred from meta-omics data, the media for selective cultivation can be designed with a wide range of physicochemical conditions such as temperature, pH, hydrostatic pressure, electron donors and acceptors, carbon sources, nitrogen sources, and growth factors. The application of novel cultivation techniques with real-time monitoring of microbial diversity and metabolic substrates and products are also recommended. Supplementary Information: The online version contains supplementary material available at 10.1007/s42995-020-00086-4.

Allopatric and Sympatric Drivers of Speciation in Alviniconcha Hydrothermal Vent Snails.

Despite significant advances in our understanding of speciation in the marine environment, the mechanisms underlying evolutionary diversification in deep-sea habitats remain poorly investigated. Here, we used multigene molecular clocks and population genetic inferences to examine processes that led to the emergence of the six extant lineages of Alviniconcha snails, a key taxon inhabiting deep-sea hydrothermal vents in the Indo-Pacific Ocean. We show that both allopatric divergence through historical vicariance and ecological isolation due to niche segregation contributed to speciation in this genus. The split between the two major Alviniconcha clades (separating A. boucheti and A. marisindica from A. kojimai, A. hessleri, and A. strummeri) probably resulted from tectonic processes leading to geographic separation, whereas the splits between co-occurring species might have been influenced by ecological factors, such as the availability of specific chemosynthetic symbionts. Phylogenetic origin of the sixth species, Alviniconcha adamantis, remains uncertain, although its sister position to other extant Alviniconcha lineages indicates a possible ancestral relationship. This study lays a foundation for future genomic studies aimed at deciphering the roles of local adaptation, reproductive biology, and host-symbiont compatibility in speciation of these vent-restricted snails.

Metabolic Potential of As-yet-uncultured Archaeal Lineages of Candidatus Hydrothermarchaeota Thriving in Deep-sea Metal Sulfide Deposits.

Candidatus Hydrothermarchaeota, formally called Marine Benthic Group E, has often been detected in iron- and sulfur-rich marine environments, such as hydrothermal vents and cold seeps. However, their ecology and physiology remain unclear. Cultivated representatives of this group are still lacking and only several metagenome-assembled genomes (MAGs) and single-amplified genomes (SAGs) are available from two deep-sea hydrothermal areas, the Juan de Fuca Ridge (JdFR) and Guaymas Basin (GB), in the north-east Pacific. We herein report four MAGs of Ca. Hydrothermarchaeota recovered from hydrothermally-inactive metal sulfide deposits at the Southern Mariana Trough (SMT) in the north-west Pacific. A phylogenetic analysis indicated that the MAGs of the SMT were distinct from those of the JdFR and GB at the genus or potentially family level. Ca. Hydrothermarchaeota MAGs from the SMT commonly possessed putative genes for carboxydotrophic and hydrogenotrophic respiration using oxidized chemical species of sulfur as electron acceptors and also for carbon fixation, as reported previously in MAGs/SAGs from the JdFR and GB. This result strongly supports Ca. Hydrothermarchaeota containing anaerobic chemolithoautotrophs using carbon monoxide and/or hydrogen as electron donors. A comparative genome analysis highlighted differences in the capability of nitrogen fixation between MAGs from the SMT and the other fields, which are consistent with environmental differences in the availability of nitrogen sources for assimilation between the fields. Based on the wide distribution in various areas, abundance, and metabolic potential of Ca. Hydrothermarchaeota, they may play a role in the biogeochemical cycling of carbon, nitrogen, sulfur, and iron in marine environments, particularly in deep-sea hydrothermal fields.

Gaidropsarus mauli a new species of three-bearded rockling (Gadiformes, Gadidae) from the Lucky Strike hydrothermal vent field (Mid-Atlantic Ridge) and the Biscay Slope (Northeastern Atlantic).

Gaidropsarus mauli, new species, is described from the Lucky Strike Hydrothermal vent site (Mid-Atlantic Ridge) and from the Bay of Biscay. It is distinguished from congeners by a combination of characters such as the number of vertebrae, the size of the first dorsal-fin ray, the profile of the head and the shape of the snout, in dorsal view, the size and the position of the eyes, the length of the pelvic fins, the shape of the pectoral fins, and the length of the lateral line. A comparison with the other 13 valid species of the genus is presented.

Primary productivity below the seafloor at deep-sea hot springs.

Below the seafloor at deep-sea hot springs, mixing of geothermal fluids with seawater supports a potentially vast microbial ecosystem. Although the identity of subseafloor microorganisms is largely known, their effect on deep-ocean biogeochemical cycles cannot be predicted without quantitative measurements of their metabolic rates and growth efficiency. Here, we report on incubations of subseafloor fluids under in situ conditions that quantitatively constrain subseafloor primary productivity, biomass standing stock, and turnover time. Single-cell-based activity measurements and 16S rRNA-gene analysis showed that Campylobacteria dominated carbon fixation and that oxygen concentration and temperature drove niche partitioning of closely related phylotypes. Our data reveal a very active subseafloor biosphere that fixes carbon at a rate of up to 321 µg C⋅L-1⋅d-1, turns over rapidly within tens of hours, rivals the productivity of chemosynthetic symbioses above the seafloor, and significantly influences deep-ocean biogeochemical cycling.

Identification and characterization of a novel alkalistable and salt-tolerant esterase from the deep-sea hydrothermal vent of the East Pacific Rise.

A novel esterase gene selected from metagenomic sequences of deep-sea hydrothermal vents was successfully expressed in Escherichia coli. The recombinant protein (est-OKK), which belongs to the lipolytic enzyme family V, exhibited high activity toward pNP-esters with short acyl chains and especially p-nitrophenyl butyrate. Site-mutagenesis results confirmed that est-OKK contains the nonclassical catalytic tetrad predicted by alignment and computational modeling. The est-OKK protein is a moderately thermophilic enzyme that is relatively thermostable, and highly salt-tolerant, which remained stable in 3 mol/L NaCl for 6 hr. The est-OKK protein showed the considerable alkalistability, displayed optimal activity at pH 9.0 and maintained approximately 70% of its residual activity after incubation at pH 10 for 4 hr. Furthermore, the est-OKK activity was strongly resistant to a variety of metal ions such as Co2+ , Zn2+ , Fe2+ , Na+ , and K+ ; nonionic detergents such as Tween-20, Tween-80; and organic solvents such as acetone and isopropanol. Taken together, the novel esterase with unique characteristics may give us a new insight into the family V of lipolytic enzymes, and could be a highly valuable candidate for biotechnological applications such as organic synthesis reactions or food and pharmaceutical industries.

Insertion sequences enrichment in extreme Red sea brine pool vent.

Mobile genetic elements are major agents of genome diversification and evolution. Limited studies addressed their characteristics, including abundance, and role in extreme habitats. One of the rare natural habitats exposed to multiple-extreme conditions, including high temperature, salinity and concentration of heavy metals, are the Red Sea brine pools. We assessed the abundance and distribution of different mobile genetic elements in four Red Sea brine pools including the world's largest known multiple-extreme deep-sea environment, the Red Sea Atlantis II Deep. We report a gradient in the abundance of mobile genetic elements, dramatically increasing in the harshest environment of the pool. Additionally, we identified a strong association between the abundance of insertion sequences and extreme conditions, being highest in the harshest and deepest layer of the Red Sea Atlantis II Deep. Our comparative analyses of mobile genetic elements in secluded, extreme and relatively non-extreme environments, suggest that insertion sequences predominantly contribute to polyextremophiles genome plasticity.

Sulfurovum riftiae sp. nov., a mesophilic, thiosulfate-oxidizing, nitrate-reducing chemolithoautotrophic epsilonproteobacterium isolated from the tube of the deep-sea hydrothermal vent polychaete Riftia pachyptila.

An anaerobic, nitrate-reducing, sulfur- and thiosulfate-oxidizing bacterium, designated strain 1812ET, was isolated from the vent polychaete Riftia pachyptila, which was collected from a deep-sea hydrothermal vent on the East Pacific Rise. Cells were Gram-stain-negative rods, measuring approximately 1.05±0.11 µm by 0.40±0.05 µm. Strain 1812ET grew at 25 - -45 °C (optimum 35 °C), with 1.5-4.0 % (w/v) NaCl (optimum 3.0 %) and at pH 5.0-8.0 (optimum pH 6.0). The generation time under optimal conditions was 3 h. Strain 1812ET was an anaerobic chemolithotroph that grew with either sulfur or thiosulfate as the energy source and carbon dioxide as the sole carbon source. Nitrate was used as a sole terminal electron acceptor. The predominant fatty acids were C16 : 1ω7c, C18 : 1ω7c and C16 : 0. The major polar lipids were phosphatidylethanolamine, diphosphatidylglycerol and phosphatidylglycerol. The major respiratory quinone was menaquinone MK-6 and the G+C content of the genomic DNA was 47.4 mol%. Phylogenetic analysis of the 16S rRNA gene of strain 1812ET showed that the isolate belonged to the Epsilonproteobacteria, and its closest relatives were Sulfurovum lithotrophicum 42BKTT and Sulfurovum aggregans Monchim 33T (98.3 and 95.7 % sequence similarity, respectively). DNA-DNA relatedness between strain 1812ET and the type strain of S. lithotrophicum was 29.7 %, demonstrating that the two strains are not members of the same species. Based on the phylogenetic, molecular, chemotaxonomic and physiological evidence, strain 1812ET represents a novel species within the genus Sulfurovum, for which the name Sulfurovum riftiae sp. nov. is proposed. The type strain is 1812ET (=DSM 101780T=JCM 30810T).

Identity and mechanisms of alkane-oxidizing metalloenzymes from deep-sea hydrothermal vents.

Six aerobic alkanotrophs (organism that can metabolize alkanes as their sole carbon source) isolated from deep-sea hydrothermal vents were characterized using the radical clock substrate norcarane to determine the metalloenzyme and reaction mechanism used to oxidize alkanes. The organisms studied were Alcanivorax sp. strains EPR7 and MAR14, Marinobacter sp. strain EPR21, Nocardioides sp. strains EPR26w, EPR28w, and Parvibaculum hydrocarbonoclasticum strain EPR92. Each organism was able to grow on n-alkanes as the sole carbon source and therefore must express genes encoding an alkane-oxidizing enzyme. Results from the oxidation of the radical-clock diagnostic substrate norcarane demonstrated that five of the six organisms (EPR7, MAR14, EPR21, EPR26w, and EPR28w) used an alkane hydroxylase functionally similar to AlkB to catalyze the oxidation of medium-chain alkanes, while the sixth organism (EPR92) used an alkane-oxidizing cytochrome P450 (CYP)-like protein to catalyze the oxidation. DNA sequencing indicated that EPR7 and EPR21 possess genes encoding AlkB proteins, while sequencing results from EPR92 confirmed the presence of a gene encoding CYP-like alkane hydroxylase, consistent with the results from the norcarane experiments.