Bacteria dominate the ammonia-oxidizing community in a hydrothermal vent site at the Mid-Atlantic Ridge of the South Atlantic Ocean

Ammonia oxidation is the first and rate-limiting step of nitrification, which is carried out by two groups of microorganisms: ammonia-oxidizing bacteria (AOB) and the recently discovered ammonia-oxidizing archaea (AOA). In this study, diversity and abundance of AOB and AOA were investigated in five rock samples from a deep-sea hydrothermal vent site at the Mid-Atlantic Ridge (MAR) of the South Atlantic Ocean. Both bacterial and archaeal ammonia monooxygenase subunit A (amoA) gene sequences obtained in this study were closely related to the sequences retrieved from deep-sea environments, indicating that AOB and AOA in this hydrothermal vent site showed typical deep ocean features. AOA were more diverse but less abundant than AOB. The ratios of AOA/AOB amoA gene abundance ranged from 1/3893 to 1/242 in all investigate samples, indicating that bacteria may be the major members responding to the aerobic ammonia oxidation in this hydrothermal vent site. Furthermore, diversity and abundance of AOA and AOB were significantly correlated with the contents of total nitrogen and total sulfur in investigated samples, suggesting that these two environmental factors exert strong influences on distribution of ammonia oxidizers in deep-sea hydrothermal vent environment.     

劳盆地深海热液喷口沉积物中细菌多样性研究

采用PCR-RFLP技术调查了劳盆地深海热液喷口两位点沉积物中的细菌多样性。结果表明, 在劳盆地深海热液喷口沉积物环境中细菌多样性十分丰富, 样品DY1中发现6个细菌类群, DY2中则存在4个细菌类群, 其中Gammaproteobacteria细菌亚群和Epsilonproteobacteria细菌亚群在两文库中均占据最大比例, 为沉积物样品中的优势菌群。另外, 在克隆文库中还发现了一些与数据库中的已知序列同源性较低的类群, 从而说明劳盆地深海热液喷口沉积物中存在特有的微生物种属。     

Novel bacterial and archaeal lineages from an in situ growth chamber deployed at a Mid-Atlantic Ridge hydrothermal vent

The phylogenetic diversity was determined for a microbial community obtained from an in situ growth chamber placed on a deep-sea hydrothermal vent on the Mid-Atlantic Ridge (23 degrees 22' N, 44 degrees 57' W). The chamber was deployed for 5 days, and the temperature within the chamber gradually decreased from 70 to 20 degrees C. Upon retrieval of the chamber, the DNA was extracted and the small-subunit rRNA genes (16S rDNA) were amplified by PCR using primers specific for the Archaea or Bacteria domain and cloned. Unique rDNA sequences were identified by restriction fragment length polymorphisms, and 38 different archaeal and bacterial phylotypes were identified from the 85 clones screened. The majority of the archaeal sequences were affiliated with the Thermococcales (71%) and Archaeoglobales (22%) orders. A sequence belonging to the Thermoplasmales confirms that thermoacidophiles may have escaped enrichment culturing attempts of deep-sea hydrothermal vent samples. Additional sequences that represented deeply rooted lineages in the low-temperature eurarchaeal (marine group II) and crenarchaeal clades were obtained. The majority of the bacterial sequences obtained were restricted to the Aquificales (18%), the epsilon subclass of the Proteobacteria (epsilon-Proteobacteria) (40%), and the genus Desulfurobacterium (25%). Most of the clones (28%) were confined to a monophyletic clade within the epsilon-Proteobacteria with no known close relatives. The prevalence of clones related to thermophilic microbes that use hydrogen as an electron donor and sulfur compounds (S(0), SO(4), thiosulfate) indicates the importance of hydrogen oxidation and sulfur metabolism at deep-sea hydrothermal vents. The presence of sequences that are related to sequences from hyperthermophiles, moderate thermophiles, and mesophiles suggests that the diversity obtained from this analysis may reflect the microbial succession that occurred in response to the shift in temperature and possible associated changes in the chemistry of the hydrothermal fluid.     

Vertical distribution of bacterial and archaeal communities along discrete layers of a deep-sea cold sediment sample at the East Pacific Rise (∼13°N)

The community structure and vertical distribution of prokaryotes in a deep-sea (ca. 3,191 m) cold sediment sample (ca. 43 cm long) collected at the East Pacific Rise (EPR) approximately 13 degrees N were studied with 16SrDNA-based molecular analyses. Total community DNA was extracted from each of four discrete layers EPRDS-1, -2, -3 and -4 (from top to bottom) and 16S rDNA were amplified by PCR. Cluster analysis of DGGE profiles revealed that the bacterial communities shifted sharply between EPRDS-1 and EPRDS-2 in similarity coefficient at merely 49%. Twenty-three sequences retrieved from DGGE bands fell into 11 groups based on BLAST and bootstrap analysis. The dominant groups in the bacterial communities were Chloroflexi, Gamma proteobacteria, Actinobacterium and unidentified bacteria, with their corresponding percentages varying along discrete layers. Pairwise Fst (F-statistics) values between the archaeal clone libraries indicated that the archaeal communities changed distinctly between EPRDS-2 and EPRDS-3. Sequences from the archaeal libraries were divided to eight groups. Crenarchaea Marine Group I (MGI) was prevalent in EPRDS-1 at 83%, while Uncultured Crenarchaea group II B (UCII B) abounded in EPRDS-4 at 61%. Our results revealed that the vertically stratified distribution of prokaryotic communities might be in response to the geochemical settings and suggested that the sampling area was influenced by hydrothermalism. The copresence of members related to hydrothermalism and cold deep-sea environments in the microbial community indicated that the area might be a transitional region from hydrothermal vents to cold deep-sea sediments.     

Microbial Community of a Hydrothermal Mud Vent Underneath the Deep-Sea Anoxic Brine Lake Urania (Eastern Mediterranean)

The composition of a metabolically active prokaryotic community thriving in hydrothermal mud fluids of the deep-sea hypersaline anoxic Western Urania Basin was characterized using rRNA-based phylogenetic analysis of a clone library. The physiologically active prokaryotic assemblage in this extreme environment showed a great genetic diversity. Most members of the microbial community appeared to be affiliated to yet uncultured organisms from similar ecosystems, i.e., deep-sea hypersaline basins and hydrothermal vents. The bacterial clone library was dominated by phylotypes affiliated with the epsilon-Proteobacteria subdivision recognized as an ecologically significant group of bacteria inhabiting deep-sea hydrothermal environments. Almost 18% of all bacterial clones were related to delta-Proteobacteria, suggesting that sulfate reduction is one of the dominant metabolic processes occurring in warm mud fluids. The remaining bacterial phylotypes were related to alpha- and beta-Proteobacteria, Actinobacteria, Bacteroides, Deinococcus-Thermus, KB1 and OP-11 candidate divisions. Moreover, a novel monophyletic clade, deeply branched with unaffiliated 16S rDNA clones was also retrieved from deep-sea sediments and halocline of Urania Basin. Archaeal diversity was much lower and detected phylotypes included organisms affiliated exclusively with the Euryarchaeota. More than 96% of the archaeal clones belonged to the MSBL-1 candidate order recently found in hypersaline anoxic environments, such as endoevaporitic microbial mats, Mediterranean deep-sea mud volcanoes and anoxic basins. Two phylotypes, represented by single clones were related to uncultured groups DHVE-1 and ANME-1. Thus, the hydrothermal mud of hypersaline Urania Basin seems to contain new microbial diversity. The prokaryotic community was significantly different from that occurring in the upper layers of the Urania Basin since 60% of all bacterial and 40% of all archaeal phylotypes were obtained only from mud fluids. The uniqueness of the composition of the active prokaryotic community could be explained by the complex environmental conditions at the site. The interaction of oxygenated warm mud fluids with the cold hypersaline brine of the Urania Basin seems to simultaneously select for various metabolic processes, such as aerobic and anaerobic heterotrophy, sulfide- and methane-dependent chemotrophy along with anaerobic oxidation of methane, sulfate- and metal-reduction.     

Archaeal diversity associated with in situ samplers deployed on hydrothermal vents on the East Pacific Rise (13 degrees N)

To evaluate possible compositional changes in archaeal communities at a deep-sea hydrothermal vent field scale, we examined five different samples obtained after deploying in situ collectors for different times on three spatially separated venting sulphide structures on the East Pacific Rise (13 degrees N). Direct cell counts and whole-cell hybridizations with fluorescently labelled 16S rRNA-based oligonucleotide probes revealed that the relative abundance of archaeal populations represented from 14 to 33% of the prokaryotic community. 16S rDNA sequence analysis of the archaeal clone libraries indicated that a large percentage of clones were closely related to known archaeal isolates recovered from similar habitats. Among the 24 different phylotypes identified, Thermococcales-related sequences were dominant in all the libraries that also included representative genera of orders Methanopyrales, Methanococcales, Archaeoglobales and Desulfurococcales. The presence of most of these phylogenetic groups was confirmed in enrichment cultures performed at temperatures from 60 to 90 degrees C. Additional sequences with no known cultivated relatives grouped with the Marine group I Crenarchaeota, Korarchaeota and Deep-sea Hydrothermal Vent Euryarchaeota (DHVE) within which a novel lineage was identified. Furthermore, the archaeal community composition was distinct from vent to vent within the same vent field and varied within short time scales. This study provides new insights into microbial diversity and distribution at deep-sea hydrothermal vents.     

Genetic diversity of archaea in deep-sea hydrothermal vent environments.

Molecular phylogenetic analysis of naturally occurring archaeal communities in deep-sea hydrothermal vent environments was carried out by PCR-mediated small subunit rRNA gene (SSU rDNA) sequencing. As determined through partial sequencing of rDNA clones amplified with archaea-specific primers, the archaeal populations in deep-sea hydrothermal vent environments showed a great genetic diversity, and most members of these populations appeared to be uncultivated and unidentified organisms. In the phylogenetic analysis, a number of rDNA sequences obtained from deep-sea hydrothermal vents were placed in deep lineages of the crenarchaeotic phylum prior to the divergence of cultivated thermophilic members of the crenarchaeota or between thermophilic members of the euryarchaeota and members of the methanogen-halophile clade. Whole cell in situ hybridization analysis suggested that some microorganisms of novel phylotypes predicted by molecular phylogenetic analysis were likely present in deep-sea hydrothermal vent environments. These findings expand our view of the genetic diversity of archaea in deep-sea hydrothermal vent environments and of the phylogenetic organization of archaea.     

Archaeal diversity in tidal flat sediment as revealed by 16S rDNA analysis

During the past ten years, Archaea have been recognized as a widespread and significant component of marine picoplankton assemblages. More recently, the presence of novel archaeal phylogenetic lineages has been discovered in coastal marine environments, freshwater lakes, polar seas, and deep-sea hydrothermal vents. Therefore, we conducted an investigation into the archaeal community existing in tidal flat sediment collected from Ganghwa Island, Korea. Phylogenetic analysis of archaeal 16S rDNA amplified directly from tidal flat sediment DNA revealed the presence of two major lineages, belonging to the Crenarchaeota (53.9%) and Euryarchaeota (46.1%) phyla. A total of 102 clones were then sequenced and analyzed by comprehensive phylogenetic analysis. The sequences determined in our samples were found to be closely related to the sequences of clones which had been previously obtained from a variety of marine environments. Archaeal clones exhibited higher similarities (83.25-100%) to sequences from other environments in the public database than did those (75.22-98.46%) of previously reported bacterial clones obtained from tidal flat sediment. The results of our study suggest that the archaeal community in tidal flat sediment is remarkably diverse.     

Fine-scale vertical distribution of bacteria in the East Pacific deep-sea sediments determined via 16S rRNA gene T-RFLP and clone library analyses

Although the deep-sea sediments harbor diverse and novel bacteria with important ecological and environmental functions, a comprehensive view of their community characteristics is still lacking, considering the vast area and volume of the deep-sea sedimentary environments. Sediment bacteria vertical distribution and community structure were studied of the E272 site in the East Pacific Ocean with the molecular methods of 16S rRNA gene T-RFLP (terminal restriction fragment length polymorphism) and clone library analyses. Layered distribution of the bacterial assemblages was detected by both methods, indicating that the shallow sediments (40 cm in depth) harbored a diverse and distinct bacterial composition with fine-scale spatial heterogeneity. Substantial bacterial diversity was detected and nine major bacterial lineages were obtained, including Acidobacteria, Actinobacteria, Bacteroidetes, Chloroflexi, Nitrospirae, Planctomycetes, Proteobacteria, and the candidate divisions OP8 and TM6. Three subdivisions of the Proteobacteria presented in our libraries, including the α-, γ- and δ-Proteobacteria. Most of our sequences have low similarity with known bacterial 16S rRNA genes, indicating that these sequences may represent as-yet-uncultivated novel bacteria. Most of our sequences were related to the GenBank nearest neighboring sequences retrieved from marine sediments, especially from deep-sea methane seep, gas hydrate or mud volcano environments. Several sequences were related to the sequences recovered from the deep-sea hydrothermal vent or basalt glasses-bearing sediments, indicating that our deep-sea sampling site might be influenced to certain degree by the nearby hydrothermal field of the East Pacific Rise at 13°N. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Population structure and phylogenetic characterization of marine benthic Archaea in deep-sea sediments.

During the past few years Archaea have been recognized as a widespread and significant component of marine picoplankton assemblages and, more recently, the presence of novel archaeal phylogenetic lineages has been reported in coastal marine benthic environments. We investigated the relative abundance, vertical distribution, phylogenetic composition, and spatial variability of Archaea in deep-sea sediments collected from several stations in the Atlantic Ocean. Quantitative oligonucleotide hybridization experiments indicated that the relative abundance of archaeal 16S rRNA in deep-sea sediments (1500 m deep) ranged from about 2.5 to 8% of the total prokaryotic rRNA. Clone libraries of PCR-amplified archaeal rRNA genes (rDNA) were constructed from 10 depth intervals obtained from sediment cores collected at depths of 1,500, 2,600, and 4,500 m. Phylogenetic analysis of rDNA sequences revealed the presence of a complex archaeal population structure, whose members could be grouped into discrete phylogenetic lineages within the two kingdoms, Crenarchaeota and Euryarchaeota. Comparative denaturing gradient gel electrophoresis profile analysis of archaeal 16S rDNA V3 fragments revealed a significant depth-related variability in the composition of the archaeal population.     

Changes in northern Gulf of Mexico sediment bacterial and archaeal communities exposed to hypoxia

Biogeochemical changes in marine sediments during coastal water hypoxia are well described, but less is known about underlying changes in microbial communities. Bacterial and archaeal communities in Louisiana continental shelf (LCS) hypoxic zone sediments were characterized by pyrosequencing 16S rRNA V4‐region gene fragments obtained by PCR amplification of community genomic DNA with bacterial‐ or archaeal‐specific primers. Duplicate LCS sediment cores collected during hypoxia had higher concentrations of Fe(II), and dissolved inorganic carbon, phosphate, and ammonium than cores collected when overlying water oxygen concentrations were normal. Pyrosequencing yielded 158 686 bacterial and 225 591 archaeal sequences from 20 sediment samples, representing five 2‐cm depth intervals in the duplicate cores. Bacterial communities grouped by sampling date and sediment depth in a neighbor‐joining analysis using Chao–Jaccard shared species values. Redundancy analysis indicated that variance in bacterial communities was mainly associated with differences in sediment chemistry between oxic and hypoxic water column conditions. Gammaproteobacteria (26.5%) were most prominent among bacterial sequences, followed by Firmicutes (9.6%), and Alphaproteobacteria (5.6%). Crenarchaeotal, thaumarchaeotal, and euryarchaeotal lineages accounted for 57%, 27%, and 16% of archaeal sequences, respectively. In Thaumarchaeota Marine Group I, sequences were 96–99% identical to the Nitrosopumilus maritimus SCM1 sequence, were highest in surficial sediments, and accounted for 31% of archaeal sequences when waters were normoxic vs. 13% of archaeal sequences when waters were hypoxic. Redundancy analysis showed Nitrosopumilus‐related sequence abundance was correlated with high solid‐phase Fe(III) concentrations, whereas most of the remaining archaeal clusters were not. In contrast, crenarchaeotal sequences were from phylogenetically diverse lineages, differed little in relative abundance between sampling times, and increased to high relative abundance with sediment depth. These results provide further evidence that marine sediment microbial community composition can be structured according to sediment chemistry and suggest the expansion of hypoxia in coastal waters may alter sediment microbial communities involved in carbon and nitrogen cycling.     

Microbial communities in iron-silica-rich microbial mats at deep-sea hydrothermal fields of the Southern Mariana Trough

The abundance, diversity and composition of bacterial and archaeal communities in the microbial mats at deep-sea hydrothermal fields were investigated, using culture-independent 16S rRNA and functional gene analyses combined with mineralogical analysis. Microbial mats were collected at two hydrothermal areas on the ridge of the back-arc spreading centre in the Southern Mariana Trough. Scanning electron microscope and energy dispersive X-ray spectroscopic (SEM-EDS) analyses revealed that the mats were mainly composed of amorphous silica and contained numerous filamentous structures of iron hydroxides. Direct cell counting with SYBR Green I staining showed that the prokaryotic cell densities were more than 10⁸ cells g⁻¹. Quantitative polymerase chain reaction (Q-PCR) analysis revealed that Bacteria are more abundant than Archaea in the microbial communities. Furthermore, zetaproteobacterial cells accounted for 6% and 22% of the prokaryotic cells in each mat estimated by Q-PCR with newly designed primers and TaqMan probe. Phylotypes related to iron-oxidizers, methanotrophs/methylotrophs, ammonia-oxidizers and sulfate-reducers were found in the 16S rRNA gene clone libraries constructed from each mat sample. A variety of unique archaeal 16S rRNA gene phylotypes, several pmoA , dsrAB and archaeal amoA gene phylotypes were also recovered from the microbial mats. Our results provide insights into the diversity and abundance of microbial communities within microbial mats in deep-sea hydrothermal fields.     

Population Structure and Phylogenetic Characterization of Marine Benthic Archaea in Deep-Sea Sediments

During the past few years Archaea have been recognized as a widespread and significant component of marine picoplankton assemblages and, more recently, the presence of novel archaeal phylogenetic lineages has been reported in coastal marine benthic environments. We investigated the relative abundance, vertical distribution, phylogenetic composition, and spatial variability of Archaea in deep-sea sediments collected from several stations in the Atlantic Ocean. Quantitative oligonucleotide hybridization experiments indicated that the relative abundance of archaeal 16S rRNA in deep-sea sediments (1500 m deep) ranged from about 2.5 to 8% of the total prokaryotic rRNA. Clone libraries of PCR-amplified archaeal rRNA genes (rDNA) were constructed from 10 depth intervals obtained from sediment cores collected at depths of 1,500, 2,600, and 4,500 m. Phylogenetic analysis of rDNA sequences revealed the presence of a complex archaeal population structure, whose members could be grouped into discrete phylogenetic lineages within the two kingdoms, Crenarchaeota and Euryarchaeota. Comparative denaturing gradient gel electrophoresis profile analysis of archaeal 16S rDNA V3 fragments revealed a significant depth-related variability in the composition of the archaeal population.     

Archaeal diversity and community development in deep-sea hydrothermal vents

Over the past 35 years, researchers have explored deep-sea hydrothermal vent environments around the globe and studied a number of archaea, their unique metabolic and physiological properties, and their vast phylogenetic diversity. Although the pace of discovery of new archaeal taxa, phylotypes and phenotypes in deep-sea hydrothermal vents has slowed recently, bioinformatics and interdisciplinary geochemistry-microbiology approaches are providing new information on the diversity and community composition of archaea living in deep-sea vents. Recent investigations have revealed that archaea could have originated and dispersed from ancestral communities endemic to hydrothermal vents into other biomes on Earth, and the community structure and productivity of chemolithotrophic archaea are controlled primarily by variations in the geochemical composition of hydrothermal fluids.     

Design of 16S rRNA-targeted oligonucleotide probes for detecting cultured and uncultured archaeal lineages in high-temperature environments

In order to facilitate the evaluation of archaeal community diversity and distribution in high-temperature environments, 14 16S rRNA oligonucleotide probes were designed. Adequate hybridization and wash conditions of the probes encompassing most known hyperthermophilic Archaea, members of the orders Thermococcales, Desulfurococcales and Sulfolobales, of the families Methanocaldococcaceae, Pyrodictiaceae and Thermoproteaceae, of the genera Archaeoglobus, Methanopyrus and Ignicoccus, and of the as yet uncultured lineages Korarchaeota, Crenarchaeota marine group I, deep-sea hydrothermal vent euryarchaeotic group 2 (DHVE 2), and deep-sea hydrothermal vent euryarchaeotic group 8 (DHVE 8) were determined by dot-blot hybridization from target and non-target reference organisms and environmental clones. The oligonucleotide probes were also used to evaluate the archaeal community composition in nine deep-sea hydrothermal vent samples. All probes, except those targeting members of Sulfolobales, Thermoproteaceae, Pyrodictiaceae and Korarchaeota, gave positive hybridization signals when hybridized against 16S rDNA amplification products obtained from hydrothermal DNA extracts. The results confirmed the widespread occurrence of Thermococcales, Desulfurococcales, Methanocaldococcaceae and Archaeoglobus in deep-sea hydrothermal vents, and extended the known ecological habitats of uncultured lineages. Despite their wide coverage, the probes were unable to resolve the archaeal communities associated with hydrothermally influenced sediments, suggesting that these samples may contain novel lineages. This suite of oligonucleotide probes may represent an efficient tool for rapid qualitative and quantitative characterization of archaeal communities. Their application would help to provide new insights in the future into the composition, distribution and abundance of Archaea in high-temperature environments.     

Microbial communities from methane hydrate-bearing deep marine sediments in a forearc basin

Microbial communities in cores obtained from methane hydrate-bearing deep marine sediments (down to more than 300 m below the seafloor) in the forearc basin of the Nankai Trough near Japan were characterized with cultivation-dependent and -independent techniques. Acridine orange direct count data indicated that cell numbers generally decreased with sediment depth. Lipid biomarker analyses indicated the presence of viable biomass at concentrations greater than previously reported for terrestrial subsurface environments at similar depths. Archaeal lipids were more abundant than bacterial lipids. Methane was produced from both acetate and hydrogen in enrichments inoculated with sediment from all depths evaluated, at both 10 and 35 degrees C. Characterization of 16S rRNA genes amplified from the sediments indicated that archaeal clones could be discretely grouped within the Euryarchaeota and Crenarchaeota domains. The bacterial clones exhibited greater overall diversity than the archaeal clones, with sequences related to the Bacteroidetes, Planctomycetes, Actinobacteria, Proteobacteria, and green nonsulfur groups. The majority of the bacterial clones were either members of a novel lineage or most closely related to uncultured clones. The results of these analyses suggest that the microbial community in this environment is distinct from those in previously characterized methane hydrate-bearing sediments.     

Phylogenetic Diversity and Ecological Pattern of Ammonia-oxidizing Archaea in the Surface Sediments of the Western Pacific

The phylogenetic diversity of ammonia-oxidizing archaea (AOA) was surveyed in the surface sediments from the northern part of the South China Sea (SCS). The distribution pattern of AOA in the western Pacific was discussed through comparing the SCS with other areas in the western Pacific including Changjiang Estuary and the adjacent East China Sea where high input of anthropogenic nitrogen was evident, the tropical West Pacific Continental Margins close to the Philippines, the deep-sea methane seep sediments in the Okhotsk Sea, the cold deep sea of Northeastern Japan Sea, and the hydrothermal field in the Southern Okinawa Trough. These various environments provide a wide spectrum of physical and chemical conditions for a better understanding of the distribution pattern and diversities of AOA in the western Pacific. Under these different conditions, the distinct community composition between shallow and deep-sea sediments was clearly delineated based on the UniFrac PCoA and Jackknife Environmental Cluster analyses. Phylogenetic analyses showed that a few ammonia-oxidizing archaeal subclades in the marine water column/sediment clade and endemic lineages were indicative phylotypes for some environments. Higher phylogenetic diversity was observed in the Philippines while lower diversity in the hydrothermal vent habitat. Water depth and possibly with other environmental factors could be the main driving forces to shape the phylogenetic diversity of AOA observed, not only in the SCS but also in the whole western Pacific. The multivariate regression tree analysis also supported this observation consistently. Moreover, the functions of current and other climate factors were also discussed in comparison of phylogenetic diversity. The information collectively provides important insights into the ecophysiological requirements of uncultured ammonia-oxidizing archaeal lineages in the western Pacific Ocean.     

Bacterial diversity of water and sediment in the Changjiang estuary and coastal area of the East China Sea

The Changjiang estuary and the coastal area of the East China Sea (ECS) represent important interfaces of terrestrial and marine environments. This study included analyses of water and sediments collected during different seasons in these regions to determine the composition of microbial assemblages by means of 16S rRNA gene clone libraries. We retrieved 1946 sequences and 779 distinct operational taxonomic units from 36 clone libraries. Shannon–Weaver diversity index values and rarefaction analysis indicated that bacterial diversity in the sediment samples was much higher than in the water samples. Proteobacteria (72.9%) was the most abundant phylum, followed by Firmicutes (6.4%), Bacteroidetes (4.6%) and Actinobacteria (4.1%). In the water, clone sequences related to Alphaproteobacteria were the most abundant, whereas in the sediment samples, sequences affiliated with Gammaproteobacteria were predominant. Principal coordinate analysis showed that water samples collected from the Changjiang estuary and the ECS clustered separately. However, this spatial pattern could not be observed in sediment samples, which were mainly distinguished from one another by the season. Bacterial diversity in the Changjiang estuary was higher than that in the ECS, which may be the result of the mixing of bacterial communities from the Changjiang River, the estuary and the coastal ocean.     

Marine subsurface eukaryotes: the fungal majority

Studies on the microbial communities of deep subsurface sediments have indicated the presence of Bacteria and Archaea throughout the sediment column. Microbial eukaryotes could also be present in deep-sea subsurface sediments; either bacterivorous protists or eukaryotes capable of assimilating buried organic carbon. DNA- and RNA-based clone library analyses are used here to examine the microbial eukaryotic diversity and identify the potentially active members in deep-sea sediment cores of the Peru Margin and the Peru Trench. We compared surface communities with those much deeper in the same cores, and compared cores from different sites. Fungal sequences were most often recovered from both DNA- and RNA-based clone libraries, with variable overall abundances of different sequence types and different dominant clone types in the RNA-based and the DNA-based libraries. Surficial sediment communities were different from each other and from the deep subsurface samples. Some fungal sequences represented potentially novel organisms as well as ones with a cosmopolitan distribution in terrestrial, fresh and salt water environments. Our results indicate that fungi are the most consistently detected eukaryotes in the marine sedimentary subsurface; further, some species may be specifically adapted to the deep subsurface and may play important roles in the utilization and recycling of nutrients.     

Analysis of dissimilatory sulfite reductase and 16S rRNA gene fragments from deep-sea hydrothermal sites of the Suiyo Seamount, Izu-Bonin Arc, Western Pacific

This study describes the occurrence of unique dissimilatory sulfite reductase (DSR) genes at a depth of 1,380 m from the deep-sea hydrothermal vent field at the Suiyo Seamount, Izu-Bonin Arc, Western Pacific, Japan. The DSR genes were obtained from microbes that grew in a catheter-type in situ growth chamber deployed for 3 days on a vent and from the effluent water of drilled holes at 5 degrees C and natural vent fluids at 7 degrees C. DSR clones SUIYOdsr-A and SUIYOdsr-B were not closely related to cultivated species or environmental clones. Moreover, samples of microbial communities were examined by PCR-denaturing gradient gel electrophoresis (DGGE) analysis of the 16S rRNA gene. The sequence analysis of 16S rRNA gene fragments obtained from the vent catheter after a 3-day incubation revealed the occurrence of bacterial DGGE bands affiliated with the Aquificae and gamma- and epsilon-Proteobacteria as well as the occurrence of archaeal phylotypes affiliated with the Thermococcales and of a unique archaeon sequence that clustered with "Nanoarchaeota." The DGGE bands obtained from drilled holes and natural vent fluids from 7 to 300 degrees C were affiliated with the delta-Proteobacteria, genus Thiomicrospira, and Pelodictyon. The dominant DGGE bands retrieved from the effluent water of casing pipes at 3 and 4 degrees C were closely related to phylotypes obtained from the Arctic Ocean. Our results suggest the presence of microorganisms corresponding to a unique DSR lineage not detected previously from other geothermal environments.