Comparative metagenomics of microbial communities inhabiting deep-sea hydrothermal vent chimneys with contrasting chemistries

Deep-sea hydrothermal vent chimneys harbor a high diversity of largely unknown microorganisms. Although the phylogenetic diversity of these microorganisms has been described previously, the adaptation and metabolic potential of the microbial communities is only beginning to be revealed. A pyrosequencing approach was used to directly obtain sequences from a fosmid library constructed from a black smoker chimney 4143-1 in the Mothra hydrothermal vent field at the Juan de Fuca Ridge. A total of 308 034 reads with an average sequence length of 227 bp were generated. Comparative genomic analyses of metagenomes from a variety of environments by two-way clustering of samples and functional gene categories demonstrated that the 4143-1 metagenome clustered most closely with that from a carbonate chimney from Lost City. Both are highly enriched in genes for mismatch repair and homologous recombination, suggesting that the microbial communities have evolved extensive DNA repair systems to cope with the extreme conditions that have potential deleterious effects on the genomes. As previously reported for the Lost City microbiome, the metagenome of chimney 4143-1 exhibited a high proportion of transposases, implying that horizontal gene transfer may be a common occurrence in the deep-sea vent chimney biosphere. In addition, genes for chemotaxis and flagellar assembly were highly enriched in the chimney metagenomes, reflecting the adaptation of the organisms to the highly dynamic conditions present within the chimney walls. Reconstruction of the metabolic pathways revealed that the microbial community in the wall of chimney 4143-1 was mainly fueled by sulfur oxidation, putatively coupled to nitrate reduction to perform inorganic carbon fixation through the Calvin–Benson–Bassham cycle. On the basis of the genomic organization of the key genes of the carbon fixation and sulfur oxidation pathways contained in the large genomic fragments, both obligate and facultative autotrophs appear to be present and contribute to biomass production.     

Characterizing the distribution and rates of microbial sulfate reduction at Middle Valley hydrothermal vents

Few studies have directly measured sulfate reduction at hydrothermal vents, and relatively little is known about how environmental or ecological factors influence rates of sulfate reduction in vent environments. A better understanding of microbially mediated sulfate reduction in hydrothermal vent ecosystems may be achieved by integrating ecological and geochemical data with metabolic rate measurements. Here we present rates of microbially mediated sulfate reduction from three distinct hydrothermal vents in the Middle Valley vent field along the Juan de Fuca Ridge, as well as assessments of bacterial and archaeal diversity, estimates of total biomass and the abundance of functional genes related to sulfate reduction, and in situ geochemistry. Maximum rates of sulfate reduction occurred at 90 °C in all three deposits. Pyrosequencing and functional gene abundance data revealed differences in both biomass and community composition among sites, including differences in the abundance of known sulfate-reducing bacteria. The abundance of sequences for Thermodesulfovibro-like organisms and higher sulfate reduction rates at elevated temperatures suggests that Thermodesulfovibro-like organisms may have a role in sulfate reduction in warmer environments. The rates of sulfate reduction presented here suggest that—within anaerobic niches of hydrothermal deposits—heterotrophic sulfate reduction may be quite common and might contribute substantially to secondary productivity, underscoring the potential role of this process in both sulfur and carbon cycling at vents.     

Spatial Differences in East Scotia Ridge Hydrothermal Vent Food Webs: Influences of Chemistry,Microbiology and Predation on Trophodynamics

The hydrothermal vents on the East Scotia Ridge are the first to be explored in the Antarctic and are dominated by large peltospiroid gastropods, stalked barnacles (Vulcanolepas sp.) and anomuran crabs (Kiwa sp.) but their food webs are unknown. Vent fluid and macroconsumer samples were collected at three vent sites (E2, E9N and E9S) at distances of tens of metres to hundreds of kilometres apart with contrasting vent fluid chemistries to describe trophic interactions and identify potential carbon fixation pathways using stable isotopes. δ¹³C of dissolved inorganic carbon from vent fluids ranged from −4.6‰ to 0.8‰ at E2 and from −4.4‰ to 1.5‰ at E9. The lowest macroconsumer δ¹³C was observed in peltospiroid gastropods (−30.0‰ to −31.1‰) and indicated carbon fixation via the Calvin-Benson-Bassham (CBB) cycle by endosymbiotic gamma-Proteobacteria. Highest δ¹³C occurred in Kiwa sp. (−19.0‰ to −10.5‰), similar to that of the epibionts sampled from their ventral setae. Kiwa sp. δ¹³C differed among sites, which were attributed to spatial differences in the epibiont community and the relative contribution of carbon fixed via the reductive tricarboxylic acid (rTCA) and CBB cycles assimilated by Kiwa sp. Site differences in carbon fixation pathways were traced into higher trophic levels e.g. a stichasterid asteroid that predates on Kiwa sp. Sponges and anemones at the periphery of E2 assimilated a proportion of epipelagic photosynthetic primary production but this was not observed at E9N. Differences in the δ¹³C and δ³⁴S values of vent macroconsumers between E2 and E9 sites suggest the relative contributions of photosynthetic and chemoautotrophic carbon fixation (rTCA v CBB) entering the hydrothermal vent food webs vary between the sites.     

Growth and Phylogenetic Properties of Novel Bacteria Belonging to the Epsilon Subdivision of the Proteobacteria Enriched from Alvinella pompejana and Deep-Sea Hydrothermal Vents

Recent molecular characterizations of microbial communities from deep-sea hydrothermal sites indicate the predominance of bacteria belonging to the epsilon subdivision of Proteobacteria (epsilon Proteobacteria). Here, we report the first enrichments and characterizations of four epsilon Proteobacteria that are directly associated with Alvinella pompejana, a deep sea hydrothermal vent polychete, or with hydrothermal vent chimney samples. These novel bacteria were moderately thermophilic sulfur-reducing heterotrophs growing on formate as the energy and carbon source. In addition, two of them (Am-H and Ex-18.2) could grow on sulfur lithoautrotrophically using hydrogen as the electron donor. Optimal growth temperatures of the bacteria ranged from 41 to 45°C. Phylogenetic analysis of the small-subunit ribosomal gene of the two heterotrophic bacteria demonstrated 95% similarity to Sulfurospirillum arcachonense, an epsilon Proteobacteria isolated from an oxidized marine surface sediment. The autotrophic bacteria grouped within a deeply branching clade of the epsilon Proteobacteria, to date composed only of uncultured bacteria detected in a sample from a hydrothermal vent along the mid-Atlantic ridge. A molecular survey of various hydrothermal vent environments demonstrated the presence of two of these bacteria (Am-N and Am-H) in more than one geographic location and habitat. These results suggest that certain epsilon Proteobacteria likely fill important niches in the environmental habitats of deep-sea hydrothermal vents, where they contribute to overall carbon and sulfur cycling at moderate thermophilic temperatures.     

Assessing the influence of physical,geochemical and biological factors on anaerobic microbial primary productivity within hydrothermal vent chimneys

Chemosynthetic primary production supports hydrothermal vent ecosystems, but the extent of that productivity and its governing factors have not been well constrained. To better understand anaerobic primary production within massive vent deposits, we conducted a series of incubations at 4, 25, 50 and 90 °C using aggregates recovered from hydrothermal vent structures. We documented in situ geochemistry, measured autochthonous organic carbon stable isotope ratios and assessed microbial community composition and functional gene abundances in three hydrothermal vent chimney structures from Middle Valley on the Juan de Fuca Ridge. Carbon fixation rates were greatest at lower temperatures and were comparable among chimneys. Stable isotope ratios of autochthonous organic carbon were consistent with the Calvin–Benson–Bassham cycle being the predominant mode of carbon fixation for all three chimneys. Chimneys exhibited marked differences in vent fluid geochemistry and microbial community composition, with structures being differentially dominated by gamma (γ) or epsilon (ε) proteobacteria. Similarly, qPCR analyses of functional genes representing different carbon fixation pathways showed striking differences in gene abundance among chimney structures. Carbon fixation rates showed no obvious correlation with observed in situ vent fluid geochemistry, community composition or functional gene abundance. Together, these data reveal that (i) net anaerobic carbon fixation rates among these chimneys are elevated at lower temperatures, (ii) clear differences in community composition and gene abundance exist among chimney structures, and (iii) tremendous spatial heterogeneity within these environments likely confounds efforts to relate the observed rates to in situ microbial and geochemical factors. We also posit that microbes typically thought to be mesophiles are likely active and growing at cooler temperatures, and that their activity at these temperatures comprises the majority of endolithic anaerobic primary production in hydrothermal vent chimneys.     

Sulfur metabolizing microbes dominate microbial communities in andesite-hosted shallow-sea hydrothermal systems

To determine microbial community composition, community spatial structure and possible key microbial processes in the shallow-sea hydrothermal vent systems off NE Taiwan's coast, we examined the bacterial and archaeal communities of four samples collected from the water column extending over a redoxocline gradient of a yellow and four from a white hydrothermal vent. Ribosomal tag pyrosequencing based on DNA and RNA showed statistically significant differences between the bacterial and archaeal communities of the different hydrothermal plumes. The bacterial and archaeal communities from the white hydrothermal plume were dominated by sulfur-reducing Nautilia and Thermococcus, whereas the yellow hydrothermal plume and the surface water were dominated by sulfide-oxidizing Thiomicrospira and Euryarchaeota Marine Group II, respectively. Canonical correspondence analyses indicate that methane (CH(4)) concentration was the only statistically significant variable that explains all community cluster patterns. However, the results of pyrosequencing showed an essential absence of methanogens and methanotrophs at the two vent fields, suggesting that CH(4) was less tied to microbial processes in this shallow-sea hydrothermal system. We speculated that mixing between hydrothermal fluids and the sea or meteoric water leads to distinctly different CH(4) concentrations and redox niches between the yellow and white vents, consequently influencing the distribution patterns of the free-living Bacteria and Archaea. We concluded that sulfur-reducing and sulfide-oxidizing chemolithoautotrophs accounted for most of the primary biomass synthesis and that microbial sulfur metabolism fueled microbial energy flow and element cycling in the shallow hydrothermal systems off the coast of NE Taiwan.     

Rapid Microbial Production of Filamentous Sulfur Mats at Hydrothermal Vents

During recent oceanographic cruises to Pacific hydrothermal vent sites (9°N and the Guaymas Basin), the rapid microbial formation of filamentous sulfur mats by a new chemoautotrophic, hydrogen sulfide-oxidizing bacterium was documented in both in situ and shipboard experiments. Observations suggest that formation of these sulfur mats may be a factor in the initial colonization of hydrothermal surfaces by macrofaunal Alvinella worms. This novel metabolic capability, previously shown to be carried out by a coastal strain in H₂S continuous-flow reactors, may be an important, heretofore unconsidered, source of microbial organic matter production at deep-sea hydrothermal vents.     

Sulfur oxidizers dominate carbon fixation at a biogeochemical hot spot in the dark ocean

Bacteria and archaea in the dark ocean (>200 m) comprise 0.3–1.3 billion tons of actively cycled marine carbon. Many of these microorganisms have the genetic potential to fix inorganic carbon (autotrophs) or assimilate single-carbon compounds (methylotrophs). We identified the functions of autotrophic and methylotrophic microorganisms in a vent plume at Axial Seamount, where hydrothermal activity provides a biogeochemical hot spot for carbon fixation in the dark ocean. Free-living members of the SUP05/Arctic96BD-19 clade of marine gamma-proteobacterial sulfur oxidizers (GSOs) are distributed throughout the northeastern Pacific Ocean and dominated hydrothermal plume waters at Axial Seamount. Marine GSOs expressed proteins for sulfur oxidation (adenosine phosphosulfate reductase, sox (sulfur oxidizing system), dissimilatory sulfite reductase and ATP sulfurylase), carbon fixation (ribulose-1,5-bisphosphate carboxylase oxygenase (RuBisCO)), aerobic respiration (cytochrome c oxidase) and nitrogen regulation (PII). Methylotrophs and iron oxidizers were also active in plume waters and expressed key proteins for methane oxidation and inorganic carbon fixation (particulate methane monooxygenase/methanol dehydrogenase and RuBisCO, respectively). Proteomic data suggest that free-living sulfur oxidizers and methylotrophs are among the dominant primary producers in vent plume waters in the northeastern Pacific Ocean.     

Desulfotomaculum and Methanobacterium spp. Dominate a 4- to 5-Kilometer-Deep Fault

Alkaline, sulfidic, 54 to 60°C, 4 to 53 million-year-old meteoric water emanating from a borehole intersecting quartzite-hosted fractures >3.3 km beneath the surface supported a microbial community dominated by a bacterial species affiliated with Desulfotomaculum spp. and an archaeal species related to Methanobacterium spp. The geochemical homogeneity over the 650-m length of the borehole, the lack of dividing cells, and the absence of these microorganisms in mine service water support an indigenous origin for the microbial community. The coexistence of these two microorganisms is consistent with a limiting flux of inorganic carbon and SO₄²⁻ in the presence of high pH, high concentrations of H₂ and CH₄, and minimal free energy for autotrophic methanogenesis. Sulfide isotopic compositions were highly enriched, consistent with microbial SO₄²⁻ reduction under hydrologic isolation. An analogous microbial couple and similar abiogenic gas chemistry have been reported recently for hydrothermal carbonate vents of the Lost City near the Mid-Atlantic Ridge (D. S. Kelly et al., Science 307:1428-1434, 2005), suggesting that these features may be common to deep subsurface habitats (continental and marine) bearing this geochemical signature. The geochemical setting and microbial communities described here are notably different from microbial ecosystems reported for shallower continental subsurface environments.     

The uptake and excretion of partially oxidized sulfur expands the repertoire of energy resources metabolized by hydrothermal vent symbioses

Symbiotic associations between animals and chemoautotrophic bacteria crowd around hydrothermal vents. In these associations, symbiotic bacteria use chemical reductants from venting fluid for the energy to support autotrophy, providing primary nutrition for the host. At vents along the Eastern Lau Spreading Center, the partially oxidized sulfur compounds (POSCs) thiosulfate and polysulfide have been detected in and around animal communities but away from venting fluid. The use of POSCs for autotrophy, as an alternative to the chemical substrates in venting fluid, could mitigate competition in these communities. To determine whether ESLC symbioses could use thiosulfate to support carbon fixation or produce POSCs during sulfide oxidation, we used high-pressure, flow-through incubations to assess the productivity of three symbiotic mollusc genera—the snails Alviniconcha spp. and Ifremeria nautilei, and the mussel Bathymodiolus brevior—when oxidizing sulfide and thiosulfate. Via the incorporation of isotopically labelled inorganic carbon, we found that the symbionts of all three genera supported autotrophy while oxidizing both sulfide and thiosulfate, though at different rates. Additionally, by concurrently measuring their effect on sulfur compounds in the aquaria with voltammetric microelectrodes, we showed that these symbioses excreted POSCs under highly sulfidic conditions, illustrating that these symbioses could represent a source for POSCs in their habitat. Furthermore, we revealed spatial disparity in the rates of carbon fixation among the animals in our incubations, which might have implications for the variability of productivity in situ. Together, these results re-shape our thinking about sulfur cycling and productivity by vent symbioses, demonstrating that thiosulfate may be an ecologically important energy source for vent symbioses and that they also likely impact the local geochemical regime through the excretion of POSCs.     

Metagenomic comparison of two Thiomicrospira lineages inhabiting contrasting deep-sea hydrothermal environments

Background

The most widespread bacteria in oxic zones of carbonate chimneys at the serpentinite-hosted Lost City hydrothermal field, Mid-Atlantic Ridge, belong to the Thiomicrospira group of sulfur-oxidizing chemolithoautotrophs. It is unclear why Thiomicrospira-like organisms thrive in these chimneys considering that Lost City hydrothermal fluids are notably lacking in hydrogen sulfide and carbon dioxide.

Methodology/Principal Findings

Here we describe metagenomic sequences obtained from a Lost City carbonate chimney that are highly similar to the genome of Thiomicrospira crunogena XCL-2, an isolate from a basalt-hosted hydrothermal vent in the Pacific Ocean. Even though T. crunogena and Lost City Thiomicrospira inhabit different types of hydrothermal systems in different oceans, their genomic contents are highly similar. For example, sequences encoding the sulfur oxidation and carbon fixation pathways (including a carbon concentration mechanism) of T. crunogena are also present in the Lost City metagenome. Comparative genomic analyses also revealed substantial genomic changes that must have occurred since the divergence of the two lineages, including large genomic rearrangements, gene fusion events, a prophage insertion, and transposase activity.

Conclusions/Significance

Our results show significant genomic similarity between Thiomicrospira organisms inhabiting different kinds of hydrothermal systems in different oceans, suggesting that these organisms are widespread and highly adaptable. These data also indicate genomic processes potentially associated with the adaptation of these lineages into strikingly different habitats.     

Pathways of carbon and energy metabolism of the epibiotic community associated with the deep-sea hydrothermal vent shrimp Rimicaris exoculata

Background

The shrimp Rimicaris exoculata dominates the faunal biomass at many deep-sea hydrothermal vent sites at the Mid-Atlantic Ridge. In its enlarged gill chamber it harbors a specialized epibiotic bacterial community for which a nutritional role has been proposed.

Methodology/Principal Findings

We analyzed specimens from the Snake Pit hydrothermal vent field on the Mid-Atlantic Ridge by complementing a 16S rRNA gene survey with the analysis of genes involved in carbon, sulfur and hydrogen metabolism. In addition to Epsilon- and Gammaproteobacteria, the epibiotic community unexpectedly also consists of Deltaproteobacteria of a single phylotype, closely related to the genus Desulfocapsa. The association of these phylogenetic groups with the shrimp was confirmed by fluorescence in situ hybridization. Based on functional gene analyses, we hypothesize that the Gamma- and Epsilonproteobacteria are capable of autotrophic growth by oxidizing reduced sulfur compounds, and that the Deltaproteobacteria are also involved in sulfur metabolism. In addition, the detection of proteobacterial hydrogenases indicates the potential for hydrogen oxidation in these communities. Interestingly, the frequency of these phylotypes in 16S rRNA gene clone libraries from the mouthparts differ from that of the inner lining of the gill chamber, indicating potential functional compartmentalization.

Conclusions

Our data show the specific association of autotrophic bacteria with Rimicaris exoculata from the Snake Pit hydrothermal vent field, and suggest that autotrophic carbon fixation is contributing to the productivity of the epibiotic community with the reductive tricarboxylic acid cycle as one important carbon fixation pathway. This has not been considered in previous studies of carbon fixation and stable carbon isotope composition of the shrimp and its epibionts. Furthermore, the co-occurrence of sulfur-oxidizing and sulfur-reducing epibionts raises the possibility that both may be involved in the syntrophic exchange of sulfur compounds, which could increase the overall efficiency of this epibiotic community.     

Culture-Dependent and -Independent Characterization of Microbial Communities Associated with a Shallow Submarine Hydrothermal System Occurring within a Coral Reef off Taketomi Island, Japan

Microbial communities in a shallow submarine hydrothermal system near Taketomi Island, Japan, were investigated using cultivation-based and molecular techniques. The main hydrothermal activity occurred in a craterlike basin (depth, ~23 m) on the coral reef seafloor. The vent fluid (maximum temperature, >52°C) contained 175 μM H₂S and gas bubbles mainly composed of CH₄ (69%) and N₂ (29%). A liquid serial dilution cultivation technique targeting a variety of metabolism types quantified each population in the vent fluid and in a white microbial mat located near the vent. The most abundant microorganisms cultivated from both the fluid and the mat were autotrophic sulfur oxidizers, including mesophilic Thiomicrospira spp. and thermophilic Sulfurivirga caldicuralii. Methane oxidizers were the second most abundant organisms in the fluid; one novel type I methanotroph exhibited optimum growth at 37°C, and another novel type I methanotroph exhibited optimum growth at 45°C. The number of hydrogen oxidizers cultivated only from the mat was less than the number of sulfur and methane oxidizers, although a novel mesophilic hydrogen-oxidizing member of the Epsilonproteobacteria was isolated. Various mesophilic to hyperthermophilic heterotrophs, including sulfate-reducing Desulfovibrio spp., iron-reducing Deferribacter sp., and sulfur-reducing Thermococcus spp., were also cultivated. Culture-independent 16S rRNA gene clone analysis of the vent fluid and mat revealed highly diverse archaeal communities. In the bacterial community, S. caldicuralii was identified as the predominant phylotype in the fluid (clonal frequency, 25%). Both bacterial clone libraries indicated that there were bacterial communities involved in sulfur, hydrogen, and methane oxidation and sulfate reduction. Our results indicate that there are unique microbial communities that are sustained by active chemosynthetic primary production rather than by photosynthetic production in a shallow hydrothermal system where sunlight is abundant.     

Carbon Dioxide Fixation by Metallosphaera yellowstonensis and Acidothermophilic Iron-Oxidizing Microbial Communities from Yellowstone National Park

The fixation of inorganic carbon has been documented in all three domains of life and results in the biosynthesis of diverse organic compounds that support heterotrophic organisms. The primary aim of this study was to assess carbon dioxide fixation in high-temperature Fe(III)-oxide mat communities and in pure cultures of a dominant Fe(II)-oxidizing organism (Metallosphaera yellowstonensis strain MK1) originally isolated from these environments. Protein-encoding genes of the complete 3-hydroxypropionate/4-hydroxybutyrate (3-HP/4-HB) carbon dioxide fixation pathway were identified in M. yellowstonensis strain MK1. Highly similar M. yellowstonensis genes for this pathway were identified in metagenomes of replicate Fe(III)-oxide mats, as were genes for the reductive tricarboxylic acid cycle from Hydrogenobaculum spp. (Aquificales). Stable-isotope (¹³CO₂) labeling demonstrated CO₂ fixation by M. yellowstonensis strain MK1 and in ex situ assays containing live Fe(III)-oxide microbial mats. The results showed that strain MK1 fixes CO₂ with a fractionation factor of ∼2.5‰. Analysis of the ¹³C composition of dissolved inorganic C (DIC), dissolved organic C (DOC), landscape C, and microbial mat C showed that mat C is from both DIC and non-DIC sources. An isotopic mixing model showed that biomass C contains a minimum of 42% C of DIC origin, depending on the fraction of landscape C that is present. The significance of DIC as a major carbon source for Fe(III)-oxide mat communities provides a foundation for examining microbial interactions that are dependent on the activity of autotrophic organisms (i.e., Hydrogenobaculum and Metallosphaera spp.) in simplified natural communities.     

Chemoautotrophic Carbon Fixation Rates and Active Bacterial Communities in Intertidal Marine Sediments

Chemoautotrophy has been little studied in typical coastal marine sediments, but may be an important component of carbon recycling as intense anaerobic mineralization processes in these sediments lead to accumulation of high amounts of reduced compounds, such as sulfides and ammonium. We studied chemoautotrophy by measuring dark-fixation of ¹³C-bicarbonate into phospholipid derived fatty acid (PLFA) biomarkers at two coastal sediment sites with contrasting sulfur chemistry in the Eastern Scheldt estuary, the Netherlands. At one site where free sulfide accumulated in the pore water right to the top of the sediment, PLFA labeling was restricted to compounds typically found in sulfur and ammonium oxidizing bacteria. At the other site, with no detectable free sulfide in the pore water, a very different PLFA labeling pattern was found with high amounts of label in branched i- and a-PLFA besides the typical compounds for sulfur and ammonium oxidizing bacteria. This suggests that other types of chemoautotrophic bacteria were also active, most likely Deltaproteobacteria related to sulfate reducers. Maximum rates of chemoautotrophy were detected in first 1 to 2 centimeters of both sediments and chemosynthetic biomass production was high ranging from 3 to 36 mmol C m⁻² d⁻¹. Average dark carbon fixation to sediment oxygen uptake ratios were 0.22±0.07 mol C (mol O₂)⁻¹, which is in the range of the maximum growth yields reported for sulfur oxidizing bacteria indicating highly efficient growth. Chemoautotrophic biomass production was similar to carbon mineralization rates in the top of the free sulfide site, suggesting that chemoautotrophic bacteria could play a crucial role in the microbial food web and labeling in eukaryotic poly-unsaturated PLFA was indeed detectable. Our study shows that dark carbon fixation by chemoautotrophic bacteria is a major process in the carbon cycle of coastal sediments, and should therefore receive more attention in future studies on sediment biogeochemistry and microbial ecology.     

Bacterial Communities Associated with Subsurface Geochemical Processes in Continental Serpentinite Springs

Reactions associated with the geochemical process of serpentinization can generate copious quantities of hydrogen and low-molecular-weight organic carbon compounds, which may provide energy and nutrients to sustain subsurface microbial communities independently of the photosynthetically supported surface biosphere. Previous microbial ecology studies have tested this hypothesis in deep sea hydrothermal vents, such as the Lost City hydrothermal field. This study applied similar methods, including molecular fingerprinting and tag sequencing of the 16S rRNA gene, to ultrabasic continental springs emanating from serpentinizing ultramafic rocks. These molecular surveys were linked with geochemical measurements of the fluids in an interdisciplinary approach designed to distinguish potential subsurface organisms from those derived from surface habitats. The betaproteobacterial genus Hydrogenophaga was identified as a likely inhabitant of transition zones where hydrogen-enriched subsurface fluids mix with oxygenated surface water. The Firmicutes genus Erysipelothrix was most strongly correlated with geochemical factors indicative of subsurface fluids and was identified as the most likely inhabitant of a serpentinization-powered subsurface biosphere. Both of these taxa have been identified in multiple hydrogen-enriched subsurface habitats worldwide, and the results of this study contribute to an emerging biogeographic pattern in which Betaproteobacteria occur in near-surface mixing zones and Firmicutes are present in deeper, anoxic subsurface habitats.     

Quantitative PCR Analysis of Functional Genes in Iron-Rich Microbial Mats at an Active Hydrothermal Vent System (Lō'ihi Seamount,Hawai'i)

The chemolithotrophic Zetaproteobacteria represent a novel class of Proteobacteria which oxidize Fe(II) to Fe(III) and are the dominant bacterial population in iron-rich microbial mats. Zetaproteobacteria were first discovered at Lō''ihi Seamount, located 35 km southeast off the big island of Hawai''i, which is characterized by low-temperature diffuse hydrothermal venting. Novel nondegenerate quantitative PCR (qPCR) assays for genes associated with microbial nitrogen fixation, denitrification, arsenic detoxification, Calvin-Benson-Bassham (CBB), and reductive tricarboxylic acid (rTCA) cycles were developed using selected microbial mat community-derived metagenomes. Nitrogen fixation genes were not detected, but all other functional genes were present. This suggests that arsenic detoxification and denitrification processes are likely cooccurring in addition to two modes of carbon fixation. Two groups of microbial mat community types were identified by terminal restriction fragment length polymorphism (T-RFLP) and were further described based on qPCR data for zetaproteobacterial abundance and carbon fixation mode preference. qPCR variance was associated with mat morphology but not with temperature or sample site. Geochemistry data were significantly associated with sample site and mat morphology. Together, these qPCR assays constitute a functional gene signature for iron microbial mat communities across a broad array of temperatures, mat types, chemistries, and sampling sites at Lō''ihi Seamount.     

Strain-level genomic variation in natural populations of Lebetimonas from an erupting deep-sea volcano

Chemolithoautotrophic Epsilonproteobacteria are ubiquitous in sulfidic, oxygen-poor habitats, including hydrothermal vents, marine oxygen minimum zones, marine sediments and sulfidic caves and have a significant role in cycling carbon, hydrogen, nitrogen and sulfur in these environments. The isolation of diverse strains of Epsilonproteobacteria and the sequencing of their genomes have revealed that this group has the metabolic potential to occupy a wide range of niches, particularly at dynamic deep-sea hydrothermal vents. We expand on this body of work by examining the population genomics of six strains of Lebetimonas, a vent-endemic, thermophilic, hydrogen-oxidizing Epsilonproteobacterium, from a single seamount in the Mariana Arc. Using Lebetimonas as a model for anaerobic, moderately thermophilic organisms in the warm, anoxic subseafloor environment, we show that genomic content is highly conserved and that recombination is limited between closely related strains. The Lebetimonas genomes are shaped by mobile genetic elements and gene loss as well as the acquisition of novel functional genes by horizontal gene transfer, which provide the potential for adaptation and microbial speciation in the deep sea. In addition, these Lebetimonas genomes contain two operons of nitrogenase genes with different evolutionary origins. Lebetimonas expressed nifH during growth with nitrogen gas as the sole nitrogen source, thus providing the first evidence of nitrogen fixation in any Epsilonproteobacteria from deep-sea hydrothermal vents. In this study, we provide a comparative overview of the genomic potential within the Nautiliaceae as well as among more distantly related hydrothermal vent Epsilonproteobacteria to broaden our understanding of microbial adaptation and diversity in the deep sea.     

Intense but variable autotrophic activity in a rapidly flushed shallow‐water hydrothermal plume (Kueishantao Islet,Taiwan)

Shallow‐water hydrothermal plumes concomitantly host both photosynthetic and chemoautotrophic organisms in a single biotope. Yet, rate measurements to quantify the contributions of different autotrophic activity types are scarce. Herein, we measured the light and dark dissolved inorganic carbon (DIC) uptake rates in the plume water of the Kueishantao hydrothermal field using the ¹³C‐labeling approach. Seventy percent of the plume‐water samples had chemoautotrophy as the dominant mode of carbon fixation, with the dark DIC uptake rates (up to 18.6 mg C/m³/h) within the range of the primary production in productive inner‐shelf waters. When considered alongside the geochemical and microbiological observations, the rate data reveal the distribution of different trophic activities in the hydrothermal plume. The autotrophic activity at the initial phase of plume dispersal is low. This is explained by the short response time the chemoautotrophs have to the stimulation from vent‐fluid discharge, and the harmful effects of hydrothermal substances on phytoplankton. As plume dispersal and mixing continue, chemoautotrophic activities begin to rise and peak in waters that have low‐to‐moderate Si(OH)₄ content. Toward the plume margin, chemoautotrophy declines to background levels, whereas photosynthesis by phytoplankton regains importance. Our results also provide preliminary indication to the loci of enhanced heterotrophy in the plume. Results of artificial mixing experiments suggest that previously formed plume water is the primary source of microbial inoculum for new plume water. This self‐inoculation mechanism, in combination with the intense DIC uptake, helps to sustain a distinct planktonic autotrophic community in this rapidly flushed hydrothermal plume.