Geomicrobiological exploration and characterization of a novel deep-sea hydrothermal system at the TOTO caldera in the Mariana Volcanic Arc

Novel hydrothermal activities accompanying effluent white smokers and elemental sulfur chimney structures at the north-east lava dome of the TOTO caldera depression in the Mariana Volcanic Arc have been explored and characterized by geochemical and microbiological surveys. White smoker hydrothermal fluids were observed in the potential hydrothermal activity centre of the field and represented the maximal temperature of 170 degrees C and the lowest pH of 1.6. The chimney structures, all consisting of elemental sulfur (sulfur chimney), were also unique to the TOTO caldera hydrothermal field. Microbial community structures in a sulfur chimney and its formation hydrothermal fluid with a high concentration of hydrogen sulfide (15 mM) have been investigated by culture-dependent and -independent analyses. 16S rRNA gene clone analysis and fluorescence in situ hybridization (FISH) analysis revealed that epsilon-Proteobacteria dominated the microbial communities in the sulfur chimney structure and formed a dense microbial mat covering the sulfur chimney surface. Archaeal phylotypes were consistently minor components in the communities and related to the genera Thermococcus, Pyrodictium, Aeropyrum, and the uncultivated archaeal group of 'deep-sea hydrothermal vent euryarchaeotal group'. Cultivation analysis suggested that the chemolithoautotrophs might play a significant ecological role as primary producers utilizing gas and sulfur compounds provided from hydrothermal fluids.     

Energy landscapes shape microbial communities in hydrothermal systems on the Arctic Mid-Ocean Ridge

Methods developed in geochemical modelling combined with recent advances in molecular microbial ecology provide new opportunities to explore how microbial communities are shaped by their chemical surroundings. Here, we present a framework for analyses of how chemical energy availability shape chemotrophic microbial communities in hydrothermal systems through an investigation of two geochemically different basalt-hosted hydrothermal systems on the Arctic Mid-Ocean Ridge: the Soria Moria Vent field (SMVF) and the Loki''s Castle Vent Field (LCVF). Chemical energy landscapes were evaluated through modelling of the Gibbs energy from selected redox reactions under different mixing ratios between seawater and hydrothermal fluids. Our models indicate that the sediment-influenced LCVF has a much higher potential for both anaerobic and aerobic methane oxidation, as well as aerobic ammonium and hydrogen oxidation, than the SMVF. The modelled energy landscapes were used to develop microbial community composition models, which were compared with community compositions in environmental samples inside or on the exterior of hydrothermal chimneys, as assessed by pyrosequencing of partial 16S rRNA genes. We show that modelled microbial communities based solely on thermodynamic considerations can have a high predictive power and provide a framework for analyses of the link between energy availability and microbial community composition.     

Distribution, phylogenetic diversity and physiological characteristics of epsilon-Proteobacteria in a deep-sea hydrothermal field

Epsilon-Proteobacteria is increasingly recognized as an ecologically significant group of bacteria, particularly in deep-sea hydrothermal environments. In this study, we studied the spatial distribution, diversity and physiological characteristics of the epsilon-Proteobacteria in various microbial habitats in the vicinity of a deep-sea hydrothermal vent occurring in the Iheya North field in the Mid-Okinawa Trough, by using culture-dependent and -independent approaches. The habitats studied were inside and outside hydrothermal plume, and annelid polychaete tubes. In addition, we deployed colonization devices near the vent emission. The polychaete tubes harboured physiologically and phylogenetically diverse microbial community. The in situ samplers were predominantly colonized by epsilon-Proteobacteria. Energy metabolism of epsilon-Proteobacteria isolates was highly versatile. Tree topology generated from the metabolic traits was significantly different (P = 0.000) from that of 16S rRNA tree, indicating current 16S rRNA gene-based analyses do not provide sufficient information to infer the physiological characteristics of epsilon-Proteobacteria. Nevertheless, culturability of epsilon-Proteobacteria in various microbial habitats differed among the phylogenetic subgroups. Members of Sulfurimonas were characterized by the robust culturability, and the other phylogenetic subgroups appeared to lose culturability in seawater, probably because of the sensitivity to oxygen. These results provide new insight into the ecophysiological characteristics of the deep-sea hydrothermal vent epsilon-Proteobacteria, which has never been assessed by comparative analysis of the 16S rRNA genes.     

Comparison of microbial communities associated with three Atlantic ultramafic hydrothermal systems

The distribution of Archaea and methanogenic, methanotrophic and sulfate-reducing communities in three Atlantic ultramafic-hosted hydrothermal systems (Rainbow, Ashadze, Lost City) was compared using 16S rRNA gene and functional gene (mcrA, pmoA and dsrA) clone libraries. The overall archaeal community was diverse and heterogeneously distributed between the hydrothermal sites and the types of samples analyzed (seawater, hydrothermal fluid, chimney and sediment). The Lost City hydrothermal field, characterized by high alkaline warm fluids (pH>11; T<95 °C), harbored a singular archaeal diversity mostly composed of unaffiliated Methanosarcinales. The archaeal communities associated with the recently discovered Ashadze 1 site, one of the deepest active hydrothermal fields known (4100 m depth), showed significant differences between the two different vents analyzed and were characterized by putative extreme halophiles. Sequences related to the rarely detected Nanoarchaeota phylum and Methanopyrales order were also retrieved from the Rainbow and Ashadze hydrothermal fluids. However, the methanogenic Methanococcales was the most widely distributed hyper/thermophilic archaeal group among the hot and acidic ultramafic-hosted hydrothermal system environments. Most of the lineages detected are linked to methane and hydrogen cycling, suggesting that in ultramafic-hosted hydrothermal systems, large methanogenic and methanotrophic communities could be fuelled by hydrothermal fluids highly enriched in methane and hydrogen.     

冲绳海槽热液区可培养硫氧化细菌多样性及其硫氧化特性

冲绳海槽热液区独特的地质环境孕育了特殊的生物群落,硫氧化细菌作为生物地球化学循环的重要参与者在热液生态系统中发挥着至关重要的作用。【目的】通过硫氧化菌株的分离培养揭示冲绳海槽热液区可培养硫氧化细菌的多样性和硫氧化活性。【方法】采用多种培养基对冲绳海槽热液区不同沉积物样品中的硫氧化细菌进行富集培养和分离纯化;利用16S rRNA基因序列确定硫氧化细菌的分类地位并进行系统发育分析;采用碘量法对典型硫氧化菌株硫氧化活性进行检测。【结果】本研究从冲绳海槽热液区样品中共分离鉴定85株硫氧化细菌,分属于α-变形菌纲、γ-变形菌纲、放线菌门和厚壁菌门,优势属为氢弧菌属(Hydrogenovibrio)、拉布伦氏菌属(Labrenzia)、深海海旋菌属(Thalassospira)和海杆状菌属(Marinobacter)。硫氧化活性检测结果表明,7株典型硫氧化菌株对硫代硫酸钠的降解活性介于31%–100%之间,其中泰坦尼克号盐单胞菌SOB56 (Halomonas titanicae SOB56)、南极海杆状菌SOB93(Marinobacter antarcticus SOB93)、印度硫氧化粗杆菌SOB107 (Thioclava indica SOB107)和嗜温氢弧菌CJG136 (Hydrogenovibrio thermophiles CJG136)可以完全降解硫代硫酸钠。【结论】冲绳海槽热液区可培养硫氧化细菌的多样性丰富,为研究该热液区的硫循环过程提供了实验材料和理论基础,多种硫氧化活性菌株的获得极大地丰富了菌种资源,为探究深海热液区硫循环的能量代谢途径和分子机制奠定基础。     

Geochemical and microbiological evidence for a hydrogen-based,hyperthermophilic subsurface lithoautotrophic microbial ecosystem (HyperSLiME) beneath an active deep-sea hydrothermal field

Subsurface microbial communities supported by geologically and abiologically derived hydrogen and carbon dioxide from the Earths interior are of great interest, not only with regard to the nature of primitive life on Earth, but as potential analogs for extraterrestrial life. Here, for the first time, we present geochemical and microbiological evidence pointing to the existence of hyperthermophilic subsurface lithoautotrophic microbial ecosystem (HyperSLiME) dominated by hyperthermophilic methanogens beneath an active deep-sea hydrothermal field in the Central Indian Ridge. Geochemical and isotopic analyses of gaseous components in the hydrothermal fluids revealed heterogeneity of both concentration and carbon isotopic compositions of methane between the main hydrothermal vent (0.08 mM and –13.8 PDB, respectively) and the adjacent divergent vent site (0.2 mM and –18.5 PDB, respectively), representing potential subsurface microbial methanogenesis, at least in the divergent vent emitting more ¹³C-depleted methane. Extremely high abundance of magmatic energy sources such as hydrogen (2.5 mM) in the fluids also encourages a hydrogen-based, lithoautotrophic microbial activity. Both cultivation and cultivation-independent molecular analyses suggested the predominance of Methanococcales members in the superheated hydrothermal emissions and chimney interiors along with the other major microbial components of Thermococcales members. These results imply that a HyperSLiME, consisting of methanogens and fermenters, occurs in this tectonically active subsurface zone, strongly supporting the existence of hydrogen-driven subsurface microbial communities.     

Microbial Community Stratification Controlled by the Subseafloor Fluid Flow and Geothermal Gradient at the Iheya North Hydrothermal Field in the Mid-Okinawa Trough (Integrated Ocean Drilling Program Expedition 331)

The impacts of lithologic structure and geothermal gradient on subseafloor microbial communities were investigated at a marginal site of the Iheya North hydrothermal field in the Mid-Okinawa Trough. Subsurface marine sediments composed of hemipelagic muds and volcaniclastic deposits were recovered through a depth of 151 m below the seafloor at site C0017 during Integrated Ocean Drilling Program Expedition 331. Microbial communities inferred from 16S rRNA gene clone sequencing in low-temperature hemipelagic sediments were mainly composed of members of the Chloroflexi and deep-sea archaeal group. In contrast, 16S rRNA gene sequences of marine group I Thaumarchaeota dominated the microbial phylotype communities in the coarse-grained pumiceous gravels interbedded between the hemipelagic sediments. Based on the physical properties of sediments such as temperature and permeability, the porewater chemistry, and the microbial phylotype compositions, the shift in the physical properties of the sediments is suggested to induce a potential subseafloor recharging flow of oxygenated seawater in the permeable zone, leading to the generation of variable chemical environments and microbial communities in the subseafloor habitats. In addition, the deepest section of sediments under high-temperature conditions (∼90°C) harbored the sequences of an uncultivated archaeal lineage of hot water crenarchaeotic group IV that may be associated with the high-temperature hydrothermal fluid flow. These results indicate that the subseafloor microbial community compositions and functions at the marginal site of the hydrothermal field are highly affected by the complex fluid flow structure, such as recharging seawater and underlying hydrothermal fluids, coupled with the lithologic transition of sediments.     

Sources of organic nitrogen at the serpentinite‐hosted Lost City hydrothermal field

The reaction of ultramafic rocks with water during serpentinization at moderate temperatures results in alkaline fluids with high concentrations of reduced chemical compounds such as hydrogen and methane. Such environments provide unique habitats for microbial communities capable of utilizing these reduced compounds in present‐day and, possibly, early Earth environments. However, these systems present challenges to microbial communities as well, particularly due to high fluid pH and possibly the availability of essential nutrients such as nitrogen. Here we investigate the source and cycling of organic nitrogen at an oceanic serpentinizing environment, the Lost City hydrothermal field (30°N, Mid‐Atlantic Ridge). Total hydrolizable amino acid (THAA) concentrations in the fluids range from 736 to 2300 nm and constitute a large fraction of the dissolved organic carbon (2.5–15.1%). The amino acid distributions, and the relative concentrations of these compounds across the hydrothermal field, indicate they most likely derived from chemolithoautotrophic production. Previous studies have identified the presence of numerous nitrogen fixation genes in the fluids and the chimneys. Organic nitrogen in actively venting chimneys has δ¹⁵N values as low as 0.1‰ which is compatible with biological nitrogen fixation. Total hydrolizable amino acids in the chimneys are enriched in ¹³C by 2–7‰ compared to bulk organic matter. The distribution and absolute δ¹³C_THAA values are compatible with a chemolithoautotrophic source, an attribution also supported by molar organic C/N ratios in most active chimneys (4.1–5.5) which are similar to those expected for microbial communities. In total, these data indicate nitrogen is readily available to microbial communities at Lost City.     

The influence of ultramafic rocks on microbial communities at the Logatchev hydrothermal field, located 15 degrees N on the Mid-Atlantic Ridge

The ultramafic-hosted Logatchev hydrothermal field (LHF) on the Mid-Atlantic Ridge is characterized by high hydrogen and methane contents in the subseafloor, which support a specialized microbial community of phylogenetically diverse, hydrogen-oxidizing chemolithoautotrophs. We compared the prokaryotic communities of three sites located in the LHF and encountered a predominance of archaeal sequences affiliated with methanogenic Methanococcales at all three. However, the bacterial composition varied in accordance with differences in fluid chemistry between the three sites investigated. An increase in hydrogen seemed to coincide with the diversification of hydrogen-oxidizing bacteria. This might indicate that the host rock indirectly selects this specific group of bacteria. However, next to hydrogen availability further factors are evident (e.g. mixing of hot reduced hydrothermal fluids with cold oxygenated seawater), which have a significant impact on the distribution of microorganisms.     

Spatial distribution of viruses associated with planktonic and attached microbial communities in hydrothermal environments

Viruses play important roles in marine surface ecosystems, but little is known about viral ecology and virus-mediated processes in deep-sea hydrothermal microbial communities. In this study, we examined virus-like particle (VLP) abundances in planktonic and attached microbial communities, which occur in physical and chemical gradients in both deep and shallow submarine hydrothermal environments (mixing waters between hydrothermal fluids and ambient seawater and dense microbial communities attached to chimney surface areas or macrofaunal bodies and colonies). We found that viruses were widely distributed in a variety of hydrothermal microbial habitats, with the exception of the interior parts of hydrothermal chimney structures. The VLP abundance and VLP-to-prokaryote ratio (VPR) in the planktonic habitats increased as the ratio of hydrothermal fluid to mixing water increased. On the other hand, the VLP abundance in attached microbial communities was significantly and positively correlated with the whole prokaryotic abundance; however, the VPRs were always much lower than those for the surrounding hydrothermal waters. This is the first report to show VLP abundance in the attached microbial communities of submarine hydrothermal environments, which presented VPR values significantly lower than those in planktonic microbial communities reported before. These results suggested that viral lifestyles (e.g., lysogenic prevalence) and virus interactions with prokaryotes are significantly different among the planktonic and attached microbial communities that are developing in the submarine hydrothermal environments.     

Abundance of Zetaproteobacteria within crustal fluids in back‐arc hydrothermal fields of the Southern Mariana Trough

To extend knowledge of subseafloor microbial communities within the oceanic crust, the abundance, diversity and composition of microbial communities in crustal fluids at back‐arc hydrothermal fields of the Southern Mariana Trough (SMT) were investigated using culture‐independent molecular techniques based on 16S rRNA gene sequences. Seafloor drilling was carried out at two hydrothermal fields, on‐ and off‐ridge of the back‐arc spreading centre of the SMT. 16S rRNA gene clone libraries for bacterial and archaeal communities were constructed from the fluid samples collected from the boreholes. Phylotypes related to Thiomicrospira in the Gammaproteobacteria (putative sulfide‐oxidizers) and Mariprofundus in the Zetaproteobacteria (putative iron‐oxidizers) were recovered from the fluid samples. A number of unique archaeal phylotypes were also recovered. Fluorescence in situ hybridization (FISH) analysis indicated the presence of active bacterial and archaeal populations in the fluids. The Zetaproteobacteria accounted for up to 32% of the total prokaryotic cell number as shown by FISH analysis using a specific probe designed in this study. Our results lead to the hypothesis that the Zetaproteobacteria play a role in iron oxidation within the oceanic crust.     

Archaeal Diversity and Distribution along Thermal and Geochemical Gradients in Hydrothermal Sediments at the Yonaguni Knoll IV Hydrothermal Field in the Southern Okinawa Trough

A variety of archaeal lineages have been identified using culture-independent molecular phylogenetic surveys of microbial habitats occurring in deep-sea hydrothermal environments such as chimney structures, sediments, vent emissions, and chemosynthetic macrofauna. With the exception of a few taxa, most of these archaea have not yet been cultivated, and their physiological and metabolic traits remain unclear. In this study, phylogenetic diversity and distribution profiles of the archaeal genes encoding small subunit (SSU) rRNA, methyl coenzyme A (CoA) reductase subunit A, and the ammonia monooxygenase large subunit were characterized in hydrothermally influenced sediments at the Yonaguni Knoll IV hydrothermal field in the Southern Okinawa Trough. Sediment cores were collected at distances of 0.5, 2, or 5 m from a vent emission (90°C). A moderate temperature gradient extends both horizontally and vertically (5 to 69°C), indicating the existence of moderate mixing between the hydrothermal fluid and the ambient sediment pore water. The mixing of reductive hot hydrothermal fluid and cold ambient sediment pore water establishes a wide spectrum of physical and chemical conditions in the microbial habitats that were investigated. Under these different physico-chemical conditions, variability in archaeal phylotype composition was observed. The relationship between the physical and chemical parameters and the archaeal phylotype composition provides important insight into the ecophysiological requirements of uncultivated archaeal lineages in deep-sea hydrothermal vent environments, giving clues for approximating culture conditions to be used in future culturing efforts.Deep-sea hydrothermal activity results in diverse physical and chemical environments for the resident microbial communities. Using cultivation techniques and culture-independent molecular analyses, diverse lineages of archaea and bacteria have so far been observed from chimney structures, retrieved in situ colonization systems settled in or on the hydrothermal conduit, microbial mats, sediments, and chemosynthetic macrofaunal bodies (19, 35, 62). Especially in the domain Archaea, most of lineages derived from hydrothermal environments have not yet been cultivated, and little is known about their physiological and metabolic traits.Environmental conditions of the habitat for a particular uncultivated archaeal lineage permit us to speculate about the physiological and metabolic traits of the archaea. For instance, the acidophilic and thermophilic archaeon “Aciduliprofundum boonei,” representing the previously uncultivated deep-sea hydrothermal vent euryarchaeotic group I (DHVEG I) subgroup 2 (DHVE2), has been isolated from a chimney habitat in the Lau Basin (49). In fact, before the cultivation of A. boonei, the DHVE2 was assumed to consist of thermophilic and acidophilic heterotrophs because their habitats had similar characteristics (13, 48, 60, 68). In order to elucidate the distribution patterns of the functionally unknown microbial components in response to the dynamically varying physico-chemical conditions, hydrothermally influenced sediments are considered better study targets than hot vent chimney structures to determine the eco-physiological roles of uncultivated microbes. This is because, unlike vent chimneys, sedimentary habitats affected by subseafloor hydrothermal fluid are expected to have more moderate physico-chemical gradients from mixing of hydrothermal fluid and ambient seawater due to the relatively lower heat convection and hydrothermal fluid penetration. Several studies have already examined the phylogenetic diversity of archaea and bacteria in hydrothermal sediments from the Guaymas Basin (7, 66), the Rainbow vent field in the Mid-Atlantic Ridge (39), and the Iheya Ridge and the Yonaguni Knoll IV in the Okinawa Trough (14, 57). However, only the relationship between the distribution pattern of microbial components and the physico-chemical conditions of these environments has been addressed.The Yonaguni Knoll IV hydrothermal field located at the southern end of the Okinawa Trough is characterized as having thick sediment, several Cl-enriched black smoker sites, and numerous vapor-enriched clear fluid sites (25, 56). The geochemical characterization of these hydrothermal fluids revealed that hydrothermal fluids undergo phase separation under the seafloor (25, 56). Furthermore, the emission of liquid CO₂ droplets has been reported, and occurrence of subseafloor CO₂ hydrate is assumed to have arisen in response to pore water chemistry in the sediments at liquid CO₂ emission sites (14, 25). According to pore water chemistry, it seems likely that these vapor-enriched hydrothermal fluids permeate the sediments around hydrothermal vent sites, and the subseafloor formation-dissociation processes of gas hydrates produce a variety of hydrothermally affected sedimentary habitats (25).In this study, we focused on the “abyss vent” site, which is characterized by 90°C hydrothermal emissions that discharge directly from the seafloor sediments (56). Sediment cores (>25 cm in length) were taken at horizontal distances of 0.5, 2, and 5 m from the hydrothermal emission while the in situ temperature of sediments was measured simultaneously. Vent fluids and interstitial water chemistry of the sediments were characterized along vertical and horizontal gradients of subseafloor mixing zones. Microbial distributions, particularly of archaea, were ascertained by culture-independent molecular analyses targeting the small subunit (SSU) rRNA gene and, mcrA (gene for methyl coenzyme A [CoA] reductase subunit A) and archaeal amoA (gene for ammonia monooxygenase large subunit). Molecular analyses for the functional genes, mcrA and amoA, are expected to indicate diversity and abundance of methanogens, anaerobic methanotrophs, and archaeal ammonium oxidizers that utilize hydrogen, methane, and ammonium, respectively, in hydrothermal fluids as electron donors. In addition, we inferred the phylogenetic diversity and distribution patterns of the bacterial SSU rRNA genes that provide insight into the potential metabolic characteristics and microbial ecosystems in each habitat.     

Post-Drilling Changes in Seabed Landscape and Megabenthos in a Deep-Sea Hydrothermal System,the Iheya North Field,Okinawa Trough

There has been an increasing interest in seafloor exploitation such as mineral mining in deep-sea hydrothermal fields, but the environmental impact of anthropogenic disturbance to the seafloor is poorly known. In this study, the effect of such anthropogenic disturbance by scientific drilling operations (IODP Expedition 331) on seabed landscape and megafaunal habitation was surveyed for over 3 years using remotely operated vehicle video observation in a deep-sea hydrothermal field, the Iheya North field, in the Okinawa Trough. We focused on observations from a particular drilling site (Site C0014) where the most dynamic change of landscape and megafaunal habitation was observed among the drilling sites of IODP Exp. 331. No visible hydrothermal fluid discharge had been observed at the sedimentary seafloor at Site C0014, where Calyptogena clam colonies were known for more than 10 years, before the drilling event. After drilling commenced, the original Calyptogena colonies were completely buried by the drilling deposits. Several months after the drilling, diffusing high-temperature hydrothermal fluid began to discharge from the sedimentary subseafloor in the area of over 20 m from the drill holes, ‘artificially’ creating a new hydrothermal vent habitat. Widespread microbial mats developed on the seafloor with the diffusing hydrothermal fluids and the galatheid crab Shinkaia crosnieri endemic to vents dominated the new vent community. The previously soft, sedimentary seafloor was hardened probably due to barite/gypsum mineralization or silicification, becoming rough and undulated with many fissures after the drilling operation. Although the effects of the drilling operation on seabed landscape and megafaunal composition are probably confined to an area of maximally 30 m from the drill holes, the newly established hydrothermal vent ecosystem has already lasted 2 years and is like to continue to exist until the fluid discharge ceases and thus the ecosystem in the area has been altered for long-term.     

Occurrence and recent long-distance dispersal of deep-sea hydrothermal vent shrimps

Deep-sea hydrothermal vents and methane seeps are extreme environments that have a high concentration of hydrogen sulphide. However, abundant unique invertebrates including shrimps of the family Bresiliidae have been found in such environments. The bresiliid shrimps are believed to have radiated in the Miocene (less than 20 Myr); however, the period when and the mechanisms by which they dispersed across the hydrothermal vents and cold seeps in oceans worldwide have not been clarified. In the present study, we collected the deep-sea blind shrimp Alvinocaris longirostris from the hydrothermal vent site in the Okinawa Trough and carried out the first investigation of the 18S rRNA gene of a bresiliid shrimp. The phylogenetic analysis revealed that the bresiliid shrimp is situated at an intermediate lineage within the infraorder Caridea and shows monophyly with palaemonid shrimps, which live in shallow sea and freshwater. Furthermore, the mitochondrial cytochrome oxidase I (COI) gene sequences were analysed to determine the phylogenetic relationship with known bresiliid shrimps. A. longirostris of the Okinawa Trough had two haplotypes of the COI gene, one of which was identical to the Alvinocaris sp. of the cold seeps in Sagami Bay. These results indicate that a long-distance dispersal of A. longirostris occurred possibly within the last 100,000 years.     

Metabolically active microbial communities in marine sediment under high-CO2 and low-pH extremes

Sediment-hosting hydrothermal systems in the Okinawa Trough maintain a large amount of liquid, supercritical and hydrate phases of CO₂ in the seabed. The emission of CO₂ may critically impact the geochemical, geophysical and ecological characteristics of the deep-sea sedimentary environment. So far it remains unclear whether microbial communities that have been detected in such high-CO₂ and low-pH habitats are metabolically active, and if so, what the biogeochemical and ecological consequences for the environment are. In this study, RNA-based molecular approaches and radioactive tracer-based respiration rate assays were combined to study the density, diversity and metabolic activity of microbial communities in CO₂-seep sediment at the Yonaguni Knoll IV hydrothermal field of the southern Okinawa Trough. In general, the number of microbes decreased sharply with increasing sediment depth and CO₂ concentration. Phylogenetic analyses of community structure using reverse-transcribed 16S ribosomal RNA showed that the active microbial community became less diverse with increasing sediment depth and CO₂ concentration, indicating that microbial activity and community structure are sensitive to CO₂ venting. Analyses of RNA-based pyrosequences and catalyzed reporter deposition-fluorescence in situ hybridization data revealed that members of the SEEP-SRB2 group within the Deltaproteobacteria and anaerobic methanotrophic archaea (ANME-2a and -2c) were confined to the top seafloor, and active archaea were not detected in deeper sediments (13–30 cm in depth) characterized by high CO₂. Measurement of the potential sulfate reduction rate at pH conditions of 3–9 with and without methane in the headspace indicated that acidophilic sulfate reduction possibly occurs in the presence of methane, even at very low pH of 3. These results suggest that some members of the anaerobic methanotrophs and sulfate reducers can adapt to the CO₂-seep sedimentary environment; however, CO₂ and pH in the deep-sea sediment were found to severely impact the activity and structure of the microbial community.     

Short-term microbial and physico-chemical variability in low-temperature hydrothermal fluids near 5°S on the Mid-Atlantic Ridge

This study examines the representativeness of low-temperature hydrothermal fluid samples with respect to their chemical and microbiological characteristics. Within this scope, we investigated short-term temporal chemical and microbial variability of the hydrothermal fluids. For this purpose we collected three fluid samples consecutively from the same spot at the Clueless field near 5°S on the southern Mid-Atlantic Ridge over a period of 50 min. During sampling, the temperature was monitored online. We measured fluid chemical parameters, characterized microbial community compositions and used statistical analyses to determine significant differences between the samples. Overall, the three fluid samples are more closely related to each other than to any other tested habitat. Therefore, on a broad scale, the three collected fluid samples can be regarded as habitat representatives. However, small differences are apparent between all samples. One of the Clueless samples even displayed significant differences ( P -value < 0.01) to the other two Clueless samples. Our data suggest that the observed variations in fluid chemical and microbial compositions are not reflecting sampling artefacts but are related to short-term fluid variability due to dynamic subseafloor fluid mixing. Recorded temporal changes in fact reflect spatial heterogeneity found in the subsurface as the fluid flows through distinctive pathways. While conservative elements (Cl, Si, Na and K) indicate variable degrees of fluid-seawater mixing, reactive components, including Fe(II), O₂ and H₂S, show that chemical and microbial reactions within the mixing zone further modify the emanating fluids on short-time scales. Fluids entrain microorganisms, which modify the chemical microenvironment within the subsurface biotopes. This is the first study focusing on short-term microbial variability linked to chemical changes in hydrothermal fluids.     

Phylogenetic diversity of sulfate-reducing prokaryotes in active deep-sea hydrothermal vent chimney structures

The phylogenetic diversity of sulfate-reducing prokaryotes occurring in active deep-sea hydrothermal vent chimney structures was characterized based on the deduced amino acid sequence analysis of the polymerase chain reaction-amplified dissimilatory sulfite reductase (DSR) gene. The DSR genes were successfully amplified from microbial assemblages of the chimney structures, derived from three geographically and geologically distinct deep-sea hydrothermal systems in the Central Indian Ridge (CIR), in the Izu-Bonin Arc (IBA), and the Okinawa Trough (OT), respectively. Phylogenetic analysis revealed seven major phylogenetic groups. More than half of the clones from the CIR chimney structure were related to DSR amino acid sequences of the hyperthermophilic archaeal members of the genus Archaeoglobus, and those of environmental DSR clones within the class Thermodesulfobacteria. From the OT chimney structure, a different group was obtained, which comprised a novel, deep lineage associated with the DSRs of the thermophilic sulfate-reducing bacterium Thermodesulfovibrio. Most of the DSR clones from the IBA chimney structure were phylogenetically associated with the delta-proteobacterial sulfate-reducing bacteria represented by the genus Desulfobulbus. Sequence analysis of DSR clones demonstrated a diverse sulfate-reducing prokaryotic community in the active deep-sea hydrothermal chimney structures.     

Halophilic Microorganisms Are Responsible for the Rosy Discolouration of Saline Environments in Three Historical Buildings with Mural Paintings

A number of mural paintings and building materials from monuments located in central and south Europe are characterized by the presence of an intriguing rosy discolouration phenomenon. Although some similarities were observed among the bacterial and archaeal microbiota detected in these monuments, their origin and nature is still unknown. In order to get a complete overview of this biodeterioration process, we investigated the microbial communities in saline environments causing the rosy discolouration of mural paintings in three Austrian historical buildings using a combination of culture-dependent and -independent techniques as well as microscopic techniques. The bacterial communities were dominated by halophilic members of Actinobacteria, mainly of the genus Rubrobacter. Representatives of the Archaea were also detected with the predominating genera Halobacterium, Halococcus and Halalkalicoccus. Furthermore, halophilic bacterial strains, mainly of the phylum Firmicutes, could be retrieved from two monuments using special culture media. Inoculation of building materials (limestone and gypsum plaster) with selected isolates reproduced the unaesthetic rosy effect and biodeterioration in the laboratory.     

First Comparative Analysis of the Community Structures and Carbon Metabolic Pathways of the Bacteria Associated with Alvinocaris longirostris in a Hydrothermal Vent of Okinawa Trough

Alvinocaris longirostris is a species of shrimp existing in the hydrothermal fields of Okinawa Trough. To date the structure and function of the microbial community associated with A. longirostris are essentially unknown. In this study, by employment of the techniques of high through-put sequencing and clone library construction and analysis, we compared for the first time the community structures and metabolic profiles of microbes associated with the gill and gut of A. longirostris in a hydrothermal field of Okinawa Trough. Fourteen phyla were detected in the gill and gut communities, of which 11 phyla were shared by both tissues. Proteobacteria made up a substantial proportion in both tissues, while Firmicutes was abundant only in gut. Although gill and gut communities were similar in bacterial diversities, the bacterial community structures in these two tissues were significantly different. Further, we discovered for the first time the existence in the gill and gut communities of A. longirostris the genes (cbbM and aclB) encoding the key enzymes of Calvin-Benson-Bassham (CBB) cycle and the reductive tricarboxylic acid (rTCA) cycle, and that both cbbM and aclB were significantly more abundant in gill than in gut. Taken together, these results provide the first evidence that at least two carbon fixation pathways are present in both the gill and the gut communities of A. longirostris, and that the communities in different tissues likely differ in autotrophic productivity.