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A series of basaltic eruptions from 1963 to 1967 off the southern coast of Iceland produced the oceanic island of Surtsey. Investigations of this volcanic system provide a time-lapse window into the real-time alteration of basaltic tephra through interactions with meteoric water in a subaerial tuff cone and with seawater in submarine deposits. In 1979, a 181 m core was recovered from a borehole (SE-01) on the eastern flank of the Surtur vent. In 2017, the ICDP-supported SUSTAIN drilling project drilled two vertical cored boreholes (SE-02a, SE-02b) to 151 and 187 m below surface (m b.s.) parallel to the 1979 borehole, and an additional angled cored borehole (SE-03) to 354 measured depth. These newly recovered cores, in comparison with the 1979 core, have promoted research into alteration processes within the volcano over the half century since its eruption. The scientific drilling undertaken in both 1979 and 2017 provides data critical to investigating mechanisms and rates of mineralogical change in basalt, evolving material and magnetic properties, and the characterization of basalt-hosted microbial communities. Previous research, including mineralogical analyses and geophysical downhole logging, reveals a weakly altered region at ~143-155 m b.s. that corresponds with a submarine zone of cool seawater inflow. The purpose of this study is to better understand processes in this zone by examining SE-02b drill core samples taken at 141.6 m b.s. (83-86 °C) with mineralogical analyses and at 148 m b.s. (83-84 °C) with magnetic analyses and microbial community analyses. Mapping of the weakly-consolidated basaltic tuff at micrometer-scale using synchrotron X-ray micro-diffraction and micro-fluorescence studies shows that the basalt is primarily composed of fresh sideromelane glass, volcanic crystals, and open voids. Olivine and labradorite are the principal volcanic minerals; they have begun to alter to lizardite and aluminous tobermorite, respectively. The basaltic glass has begun to alter to nanocrystalline clinochlore and smectitic clay mineral, mainly nontronite and montmorillonite. The abundance of fresh glass, however, confirms a weakly altered region of the volcano. Uniaxial and cubic single domain titanomagnetite is the principal magnetic remanence carrier in the glass, whereas the magnetic minerals in more highly altered zones of lapilli tuff, only a few meters distant, are more oxidized and exhibit different magnetic anisotropies, consistent with the growth of secondary titanomaghemite. The properties of magnetic remanence remained relatively stable in the cool seawater inflow zone but changed very rapidly during fluid-rock interactions at higher hydrothermal temperatures. The microbial community detected in the drill core sample at 148 m b.s. from SE-02b is dominated by taxa generally found in seawater such as Psychromonas, Glaciecola, Marinomonas and suggests a possible infiltration of microbial taxa from the seawater to the submarine deposit. This anomalously permeable, poorly-consolidated horizon provides a strong contrast to the characteristics of the well-lithified lapilli tuff deposits and demonstrates the potential for substantial variability in mineralogical, magnetic and microbial submarine processes in other Surtseyan volcanoes and seamount structures.

Publisher's version (útgefin grein), One of the main objectives for astrobiology is to unravel and explore the habitability of environments beyond Earth, paying special attention to Mars. If the combined environmental stress factors on Mars are compatible with life or if they were less harsh in the past, to investigate the traces of past or present life is critical to understand its potential habitability. Essential for this research is the characterization of Mars analogue environments on Earth through the development of techniques for biomarker detection in them. Biosensing techniques based on fluorescence sandwich microarray immunoassays (FSMI) have shown to be a powerful tool to detect biosignatures and depict the microbial profiles of different environments. In this study, we described the microbial biomarker profile of five anoxic Mars analogues sites using the Life Detector Chip (LDChip), an antibody microarray for multiple microbial marker detection. Furthermore, we contributed to new targets by developing a new 26-polyclonal antibodies microarray using crude extracts from anaerobic sampling sites, halophilic microorganisms, and anaerobic isolates obtained in the framework of the European Mars Analogues for Space Exploration (MASE) project. The new subset of antibodies was characterized and implemented into a microarray platform (MASE-Chip) for microbial marker searching in salty and anaerobic environments., MASE is supported by European Community’s Seventh Framework Program (FP7/2007–2013) under Grant Agreement N° 607297.

The marine genus Pseudoalteromonas is known for its versatile biotechnological potential with respect to the production of antimicrobials and enzymes of industrial interest. We have sequenced the genomes of three Pseudoalteromonas sp. strains isolated from different deep sea sponges on the Illumina MiSeq platform. The isolates have been screened for various industrially important enzymes and comparative genomics has been applied to investigate potential relationships between the isolates and their host organisms, while comparing them to free-living Pseudoalteromonas spp. from shallow and deep sea environments. The genomes of the sponge associated Pseudoalteromonas strains contained much lower levels of potential eukaryotic-like proteins which are known to be enriched in symbiotic sponge associated microorganisms, than might be expected for true sponge symbionts. While all the Pseudoalteromonas shared a large distinct subset of genes, nonetheless the number of unique and accessory genes is quite large and defines the pan-genome as open. Enzymatic screens indicate that a vast array of enzyme activities is expressed by the isolates, including β-galactosidase, β-glucosidase, and protease activities. A β-glucosidase gene from one of the Pseudoalteromonas isolates, strain EB27 was heterologously expressed in Escherichia coli and, following biochemical characterization, the recombinant enzyme was found to be cold-adapted, thermolabile, halotolerant, and alkaline active.

A molecular biological analysis of Icelandic volcanic rocks of different compositions and glassiness revealed the presence of Actinobacteria as an abundant phylum. In outcrops of basaltic glass they were the dominant bacterial phylum. A diversity of Actinobacteria were cultured from the rocks on rock-agar plates showing that they are capable of growing on rock-derived nutrient sources and that many of the taxa identified by molecular methods are viable, potentially active members of the community. Laboratory batch-culture experiments using a Streptomyces isolate showed that it was capable of enhancing the release of major elements from volcanic rocks, including weathered basaltic glass, crystalline basalt and komatiite, when provided with a carbon source. Actinobacteria of a variety of other sub-orders were also capable of enhancing volcanic rock weathering, measured as Si release. However, most strains did not significantly increase the weathering of the silica-rich rock, obsidian. These data show that Actinobacteria can contribute to volcanic rock weathering and, therefore, the carbonate-silicate cycle. Given their ancient lineage, it is likely they have played a role in rock weathering for over two billion years.

Icelandic streams with mean annual temperatures of less than 5 °C, which receive the cationic products of basaltic rock weathering, were found to host mats of iron-cycling microorganisms. We investigated two representative sites. Iron-oxidizing Gallionella and iron-reducing Geobacter species were present. The mats host a high bacterial diversity as determined by culture-independent methods. β-Proteobacteria, Actinobacteria, α-Proteobacteria, and Bacteroidetes were abundant microbial taxa. The mat contained a high number of phototroph sequences. The carbon compounds in the mat displayed broad G and D bands with Raman spectroscopy. This signature becomes incorporated into the weathered oxidized surface layer of the basaltic rocks and was observed on rocks that no longer host mats. The presence of iron-oxidizing taxa in the stream microbial mats, and the lack of them in previously studied volcanic rocks in Iceland that have intermittently been exposed to surface water flows, can be explained by the kinetic limitations to the extraction of reduced iron from rocks. This type of ecosystem illustrates key factors that control the distribution of chemolithotrophs in cold volcanic environments. The data show that one promising sample type for which the hypothesis of the existence of past life on Mars can be tested is the surface of volcanic rocks that, previously, were situated within channels carved by flowing water. Our results also show that the carbonaceous signatures of life, if life had occurred, could be found in or on these rocks.

Nine thermophilic strains of aerobic, non-sporulating, heterotrophic bacteria were isolated after enrichment of chimney material sampled from a deep-sea hydrothermal field at a depth of 2634 m on the East-Pacific Rise (1 °N). The bacteria stained Gram-negative. They were rod-shaped and measured approximately 0.5 μm in width and 1.5–3.5 μm in length. They grew at 55–80 °C, pH 6–8 and 1–6 % NaCl. Optimal growth was observed at 70–75 °C, pH 7.0 and 1–3 % NaCl. The organisms were identified as members of the genus Rhodothermus, having a 16S rRNA gene similarity of 98.1 % with Rhodothermus marinus DSM 4252T. The novel isolates differed morphologically, physiologically and chemotaxonomically from R. marinus, e.g. in lack of pigmentation, response to hydrostatic pressure, maximum growth temperature and DNA G+C content. DNA–DNA hybridization revealed a reassociation value of 37.2 % between strain PRI 2902T and R. marinus DSM 4252T, which strongly suggested that they represent different species. Furthermore, AFLP fingerprinting separated the novel strains from R. marinus reference strains. It is therefore concluded that the strains described here should be classified as representatives of a novel species for which the name Rhodothermus profundi sp. nov. is proposed; the type strain is PRI 2902T (=DSM 22212T =JCM 15944T).

The diversity of microbial communities inhabiting two terrestrial volcanic glasses of contrasting mineralogy and age was characterised. Basaltic glass from a

Bacteria play an important role in rock weathering and yet their diversity and potential activity in the terrestrial rock weathering environment is poorly understood. Culture and culture-independent methods (16S rDNA) were used to investigate the populations of bacteria inhabiting a basaltic glass/palagonite subglacial (hyaloclastite) deposit subject to weathering in Iceland. The rock hosts a diverse microbial community. The 16S rDNA clones were dominated by Actinobacteria, Proteobacteria, Bacteroidetes and Acidobacteria. Representatives of Gemmatimonadetes and Verrucomicrobia were present. Isolation of organisms on basalt/palagonite yielded only two isolates, an actinobacterium and a Bacteroidetes, showing that the active species, at least in the time scale of laboratory cultivation, are a small proportion of the total diversity. Firmicutes and Actinobacteria were isolated when basalt/palagonite was supplemented with an organic source. Many of the isolates demonstrated tolerance to transition metals (Cr,...

By combining low nutrient enrichments and molecular methods, a high diversity of new amylase genes was detected in a neutral sulphide-rich hot spring in Iceland. Enrichments based on hot spring water and low concentrations of starch were used to select slow-growing, starch-degrading microorganisms. Six enrichments had in total 17 bacterial types detected by 16S rRNA analysis, mostly related to the Thermus-Deinococcus group, green non-sulphur bacteria, gram positives, and uncultivated new candidate divisions. No Archaea were found. The apparent 16S rRNA species composition of the enrichments was very different from that of the microbial mat in the same hot spring. DNA samples obtained from 4 enrichments and from hot spring biomass were screened by PCR for amylase genes in glycoside-hydrolase family 13. Degenerate primers, based on conserved amino acid sequences from multiple␣alignments of family 13, enabled the detection of 18 amylase sequence types in the enrichments, including α-amylases, α-glucosidases, 1,4-α-glucan branching enzymes, cyclomaltodextrin hydrolases, maltogenic amylases and neopullulanases, and unspecified family 13 glycoside-hydrolases. Only one unique neopullulanase sequence, also found in most of the enrichments, was detected in the hot spring biomass DNA. The results suggest that the enrichment method combined with sequence-based screening is an efficient way to access the silent, i.e. not detectable, gene diversity in natural environments.

Kopf, Anna ... et. al.-- 5 pages, 1 figure.-- This manuscript is NOAA-GLERL contribution number 1763, Ocean Sampling Day was initiated by the EU-funded Micro B3 (Marine Microbial Biodiversity, Bioinformatics, Biotechnology) project to obtain a snapshot of the marine microbial biodiversity and function of the world’s oceans. It is a simultaneous global mega-sequencing campaign aiming to generate the largest standardized microbial data set in a single day. This will be achievable only through the coordinated efforts of an Ocean Sampling Day Consortium, supportive partnerships and networks between sites. This commentary outlines the establishment, function and aims of the Consortium and describes our vision for a sustainable study of marine microbial communities and their embedded functional traits, This work was supported by the Micro B3 project, which is funded from the European Union’s Seventh Framework Programme (FP7; Joint Call OCEAN.2011‐2: Marine microbial diversity – new insights into marine ecosystems functioning and its biotechnological potential) under the grant agreement no 287589

A thermophilic, anaerobic, piezophilic, chemo-organotrophic sulfur-reducing bacterium, designated as KA3T, was isolated from a deep-sea hydrothermal chimney sample collected at a depth of 2630 m on the East-Pacific Rise (13 degrees N). When grown under elevated hydrostatic pressure, the cells are rod-shaped with a sheath-like outer structure, motile, have a mean length of 1-1.5 microm and stain Gram-negative. They appear singly or in short chains. When grown at lower, or atmospheric, pressures, the cells elongate and become twisted. Growth is enhanced by hydrostatic pressure; the optimal pressure for growth is 40 MPa (26 MPa pressure at sampling site). The temperature range for growth is 45-70 degrees C, the optimum being around 65 degrees C (doubling time is approximately 20 min at 40 MPa). Growth is observed from pH 5 to pH 8, the optimum being at pH 6. The salinity range for growth is 10-50 g NaCl l(-1), the optimum being at 30 g l(-1). The isolate is able to grow on a broad spectrum of carbohydrates or complex proteinaceous substrates, and growth is stimulated by L-cystine and elemental sulfur. The G+C content of the genomic DNA is 29 +/- 1 mol%. According to phylogenetic analysis of the 16S rDNA gene, the strain is placed within the order Thermotogales, in the bacterial domain. On the basis of 16S rDNA sequence comparisons and morphological, physiological and genotypic characteristics, it is proposed that the isolate be described as a novel species of the genus Marinitoga, with Marinitoga piezophila sp. nov. as the type species. The type strain is KA3T (= DSM 14283T = JCM 11233T).

A study of the mineralogy, chemical composition and structure of poorly-crystalline saponite precipitated from a submarine hot spring in Eyjafjordur, northern Iceland is reported. Special emphasis was placed on the microstructures of the minerals and a possible connection with biological activity during their precipitation. The microstructures of the minerals were found to be very similar to specific clay minerals precipitated from geothermal vents in oceanic rift zones. The composition of the minerals was, however, found to be similar to magnesium silicate scales formed in geothermal installations in Iceland where geothermal waters were mixed with cold fresh waters. High contents of organic substances were found in the clay mineral samples as compared to geothermal precipitates from other localities. Microstructural features of the layer silicates in one of the samples suggest that a gelatinous substance was a precursor of the saponite clay. The organic matter content appears to be greater when the precipitates are more crystalline.

Little is understood regarding the phylogeny and metabolic capabilities of the earliest colonists of volcanic rocks, yet these data are essential for understanding how life becomes established in, and interacts with the planetary crust, ultimately contributing to critical zone processes and soil formation. Here we report the use of molecular and culture-dependent methods to determine the composition of pioneer microbial communities colonising the basaltic Fimmvörðuháls lava flow at Eyjafjallajökull, Iceland, formed in 2010. Our data show that three to five months post eruption, the lava was colonized by a low-diversity microbial community dominated by Betaproteobacteria, primarily taxa related to nonphototrophic diazotrophs such as Herbaspirillum spp., and chemolithotrophs such as Thiobacillus. Although successfully cultured following enrichment, phototrophs were not abundant members of the Fimmvörðuháls communities, as revealed by molecular analysis, and phototrophy is therefore not likely to be a dominant biogeochemical process in these early successional basalt communities. These results contrast with older Icelandic lava of comparable mineralogy, in which phototrophs comprised a significant fraction of microbial communities and the non-phototrophic community fractions were dominated by Acidobacteria and Actinobacteria.

Microbial community profiles of recently formed hot spring systems ranging in temperatures from 57°C to 100°C and pH values from 2 to 4 in Hveragerði (Iceland) were analyzed with PhyloChip G3 technology. In total, 1173 bacterial operational taxonomic units (OTUs) spanning 576 subfamilies and 38 archaeal OTUs covering 32 subfamilies were observed. As expected, the hyperthermophilic (100°C) spring system exhibited both low microbial biomass and diversity when compared to thermophilic (60°C) springs. Ordination analysis revealed distinct bacterial and archaeal diversity in geographically distinct hot springs. Slight variations in temperature (from 57°C to 64°C) within the interconnected pools led to a marked fluctuation in microbial abundance and diversity. Correlation and PERMANOVA tests provided evidence that temperature was the key environmental factor responsible for microbial community dynamics, while pH, H_(2)S, and SO_2 influenced the abundance of specific microbial groups. When archaeal community composition was analyzed, the majority of detected OTUs correlated negatively with temperature, and few correlated positively with pH. Key Words: Microbial diversity—PhyloChip G3—Acidophilic—Thermophilic—Hot springs—Iceland. Astrobiology 14, xxx–xxx.

Alanine dehydrogenase (AlaDH: EC 1.4.1.1), malate dehydrogenase (MDH: EC 1.1.1.37), and glutamate dehydrogenase (EC 1.4.1.2), all NAD+ dependent, were detected in extracts from a psychrophilic bacterium, strain PA-43, isolated from a sea urchin off the Icelandic coast. Characterization tests suggested that the strain had a close relationship to Vibrio, but sequencing of part of the 16S rDNA gene placed the bacterium among Shewanella species in a constructed phylogenetic tree. The bacterium had an optimum growth temperature of 16.5 degrees C, and maximum dehydrogenase expression was obtained in a rich medium supplemented with NaCl. Both AlaDH and MDH were purified to homogeneity. AlaDH is a hexamer, with an approximate relative molecular mass of 260,000, whereas MDH is dimeric, with an apparent relative molecular mass of approximately 70,000. Both enzymes were thermolabile, and the optimum temperatures for activity were shifted toward lower temperatures than those found in the same enzymes from mesophiles, 37 degrees C for MDH and approximately 47 degrees C for AlaDH. The pH optima for AlaDH in the forward and reverse reactions were 10.5 and 9, respectively, whereas those for MDH were 10-10.2 and 8.8, respectively. Partial amino acid sequences, comprising approximately 30% of the total sequences from each enzyme, were determined for N-terminal, tryptic, and chymotryptic fragments of the enzymes. The AlaDH showed the highest similarity to AlaDHs from the psychrotroph Shewanella Ac10 and the mesophile Vibrio proteolyticus, whereas MDH was most similar to the MDHs from the mesophiles Escherichia coli and Haemophilus influenzae, with lower identity to the psychrophilic malate dehydrogenases from Vibrio 5710 and Photobacterium SS9.

A novel barophilic, hyperthermophilic, anaerobic sulfur-metabolizing archaeon, strain MPT (T = type strain), was isolated from a hydrothermal vent site (Snakepit) on the Mid-Atlantic Ridge (depth, 3550 m). Enrichments and isolation were done under 40 MPa hydrostatic pressure at 95 degrees C. Strain MPT was barophilic at 75, 80, 85, 90, 95 and 98 degrees C, and was an obligate barophile between 95 and 100 degrees C (Tmax). For growth above 95 degrees C, a pressure of 15.0-17.5 MPa was required. The strain grew at 48-95 degrees C under atmospheric pressure. The optimal temperature for growth was 85 degrees C at both high (40 MPa) and low (0.3 MPa) pressures. The growth rate was twofold higher at 85 degrees C under in situ hydrostatic pressure compared to at low pressure. Strain MPT cells were motile, coccoid, 0.8-2.0 microns in diameter and covered by a hexagonal S-layer lattice. The optimum pH and NaCl concentration for growth at low pressure were 7.0 and 20-30 g l-1, respectively. The new isolate was an obligate heterotroph and utilized yeast extract, beef extract and peptone for growth. Growth was optimal in the presence of elemental sulfur. Rifampicin and chloramphenicol inhibited growth. The core lipids consisted of a major archaeol and a complex lipid pattern consisting of a major phospholipid. The DNA G + C content was 37.1 mol%. Sequencing of the 16S rRNA gene revealed that strain MPT belonged to the genus Thermococcus and it is proposed that this isolate should be designated as a new species, Thermococcus barophilus.

An in situ sampler that encloses individual blocks of thermally stable solid medium made with Gelrite was successfully deployed at a deep-sea hydrothermal vent environment. A mixed population of microorganisms was enriched on the surface of the Gelrite blocks. Thermophilic Bacilli spp. and bacteria belonging to the Thermotogales were isolated.Key words: in situ enrichment, sampler, hydrothermal vent.

The physiology of the deep-sea hyperthermophilic, anaerobic vent archaeon Pyrococcus abyssi, originating from the Fiji Basin at a depth of 2,000 m, was studied under diverse conditions. The emphasis of these studies lay in the growth and survival of this archaeon under the different conditions present in the natural habitat. Incubation under in situ pressure (20 MPa) and at 40 MPa increased the maximal and minimal growth temperatures by 4(deg)C. In situ pressure enhanced survival at a lethal high temperature (106 to 112(deg)C) relative to that at low pressure (0.3 MPa). The whole-cell protein profile, analyzed by one-dimensional sodium dodecyl sulfate gel electrophoresis, did not change in cultures grown under low or high pressure at optimal and minimal growth temperatures, but several changes were observed at the maximal growth temperature under in situ pressure. The complex lipid pattern of P. abyssi grown under in situ and 0.1- to 0.5-MPa pressures at different temperatures was analyzed by thin-layer chromatography. The phospholipids became more complex at a low growth temperature at both pressures but their profiles were not superimposable; fewer differences were observed in the core lipids. The polar lipids were composed of only one phospholipid in cells grown under in situ pressure at high temperatures. Survival in the presence of oxygen and under starvation conditions was examined. Oxygen was toxic to P. abyssi at growth range temperature, but the strain survived for several weeks at 4(deg)C. The strain was not affected by starvation in a minimal medium for at least 1 month at 4(deg)C and only minimally affected at 95(deg)C for several days. Cells were more resistant to oxygen in starvation medium. A drastic change in protein profile, depending on incubation time, was observed in cells when starved at growth temperature.

Existence of life in extreme environments has been known for a long time, and their habitants have been investigated by different scientific disciplines for decades. However, reports of multidisciplinary research are uncommon. In this paper, we report an interdisciplinary three-day field campaign conducted in the framework of the Coordination Action for Research Activities on Life in Extreme Environments (CAREX) FP7EU program, with participation of experts in the fields of life and earth sciences. In situ experiments and sampling were performed in a 20 m long hot springs system of different temperature (57 °C to 100 °C) and pH (2 to 4). Abiotic factors were measured to study their influence on the diversity. The CO2 and H2S concentration varied at different sampling locations in the system, but the SO2 remained the same. Four biofilms, mainly composed by four different algae and phototrophic protists, showed differences in photosynthetic activity. Varying temperature of the sampling location affects chlorophyll fluorescence, not only in the microbial mats, but plants (Juncus), indicating selective adaptation to the environmental conditions. Quantitative polymerase chain reaction (PCR), DNA microarray and denaturing gradient gel electrophoresis (DGGE)-based analysis in laboratory showed the presence of a diverse microbial population. Even a short duration (30 h) deployment of a micro colonizer in this hot spring system led to colonization of microorganisms based on ribosomal intergenic spacer (RISA) analysis. Polyphasic analysis of this hot spring system was possible due to the involvement of multidisciplinary approaches., The authors are grateful to the CAREX Project (Coordination Action for Research Activities on Life in Extreme Environments, FP7-ENV-2007-1 project No. 211700) for funding the fieldtrip to Iceland. A special thanks to N. Walter, ESF and to all other participants in the Icelandic CAREX fieldtrip for fieldwork assistance. The laboratory analyses of FlowCam were supported by a project of the Ministry of Education, Youth and Sports of the Czech Republic, no. LM2010009 CzechPolar²Czech polar stations: Construction and logistic expenses, by project Creating of the Working Team and Pedagogical Conditions for Teaching and Education in the Field of Polar Ecology and Life in Extreme Environment, reg. No. CZ.1.07/2.2.00/28.0190, co-financed by the European Social Fund and the state budget of the Czech Republic and as a long-term research development project no. RVO 67985939. Oxymetric analysis were supported by a Spanish Ministry of Economy and Competitive CGL2011/02254 Grant. F. Mapelli was supported by Università degli Studi di Milano.

We performed phenotypic and physiological studies with 20 hyperthermophilic microorganisms isolated from hydrothermal vents located in the North Fiji Basin (southwestern Pacific Ocean) at a depth of 2,000 m. These isolates were strict anaerobes that were regular to irregular coccoids and used elemental sulfur in their metabolism. Growth was observed at temperatures ranging from 50 to 101°C. The DNA base compositions varied from 43 to 60 mol%. All of these organisms were heterotrophs and fermented peptides to acetate, isovalerate, isobutyrate, and propionate. They contained both diether and tetraether lipids in their membranes, which indicates that they belong to the domain Archaea. DNA-DNA hybridization experiments revealed that there were two distinct homology groups, which correlated well with results obtained from sodium dodecyl sulfate-polyacrylamide gel electrophoresis of soluble whole-cell proteins, and these groups corresponded to the genera Pyrococcus and Thermococcus. Five isolates exhibited levels of DNA-DNA relatedness with Pyrococcus abyssi ranging from 71 to 100% and produced almost identical protein patterns. The remaining isolates formed a weakly homogeneous group based on DNA-DNA similarity data and protein patterns; the results of unweighted pair group cluster analyses suggested that these isolates were members of five new species of the genus Thermococcus.

Subglacial lakes beneath the Vatnajokull ice cap in Iceland host endemic communities of microorganisms adapted to cold, dark and nutrient-poor waters, but the mechanisms by which these microbes disseminate under the ice and colonize these lakes are unknown. We present new data on this subglacial microbiome generated from samples of two subglacial lakes, a subglacial flood and a lake that was formerly subglacial but now partly exposed to the atmosphere. These data include parallel 16S rRNA gene amplicon libraries constructed using novel primers that span the v3–v5 and v4–v6 hypervariable regions. Archaea were not detected in either subglacial lake, and the communities are dominated by only five bacterial taxa. Our paired libraries are highly concordant for the most abundant taxa, but estimates of diversity (abundance-based coverage estimator) in the v4–v6 libraries are 3–8 times higher than in corresponding v3–v5 libraries. The dominant taxa are closely related to cultivated anaerobes and microaerobes, and may occupy unique metabolic niches in a chemoautolithotrophic ecosystem. The populations of the major taxa in the subglacial lakes are indistinguishable (>99% sequence identity), despite separation by 6 km and an ice divide; one taxon is ubiquitous in our Vatnajokull samples. We propose that the glacial bed is connected through an aquifer in the underlying permeable basalt, and these subglacial lakes are colonized from a deeper, subterranean microbiome.

Thermococcus barophilus is a hyperthermophilic, anaerobic, mixed heterotrophic, and carboxydotrophic euryarchaeon isolated from the deep sea hydrothermal vent Snakepit site on the mid-Atlantic ridge at a depth of 3,550 m. T. barophilus is the first true piezophilic, hyperthermophilic archaeon isolated, having an optimal growth at 40 MPa. Here we report the complete genome sequence of strain MP, the type strain of T. barophilus . The genome data reveal a close proximity with T hermococcus sibiricus , another Thermococcus isolated from the deep biosphere and a possible connection to life in the depths.

International audience; Bacteria inhabiting crystalline rocks from two terrestrial Icelandic volcanic lava flows of similar age and from the same geographical region, but differing in porosity and mineralogy, were characterised. Microarray (PhyloChip) and clone library analysis of 16S rRNA genes revealed the presence of a diverse assemblage of bacteria in each lava flow. Both methods suggested a more diverse community at the Dómadalshraun site (rhyolitic/andesitic lava flow) than that present at the Hnausahraun site (basaltic lava flow). Proteobacteria dominated the clone library at the Dómadalshraun site, while Acidobacteria was the most abundant phylum in the Hnausahraun site. Although analysis of similarities of denaturing gradient gel electrophoresis profiles suggested a strong correlation of community structure with mineralogy, rock porosity may also play an important role in shaping the bacterial community in crystalline volcanic rocks. Clone sequences were most similar to uncultured microorganisms, mainly from soil environments. Of these, Antarctic soils and temperate rhizosphere soils were prominent, as were clones retrieved from Hawaiian and Andean volcanic soils. The novel diversity of these Icelandic microbial communities was supported by the finding that up to 46% of clones displayed

Background: International fish trade reached an import value of 62.8 billion Euro in 2006, of which 44.6% are covered by the European Union. Species identification is a key problem throughout the life cycle of fishes: from eggs and larvae to adults in fisheries research and control, as well as processed fish products in consumer protection. Methodology/Principal Findings: This study aims to evaluate the applicability of the three mitochondrial genes 16S rRNA (16S), cytochrome b (cyt b), and cytochrome oxidase subunit I (COI) for the identification of 50 European marine fish species by combining techniques of "DNA barcoding" and microarrays. In a DNA barcoding approach, neighbour Joining (NJ) phylogenetic trees of 369 16S, 212 cyt b, and 447 COI sequences indicated that cyt b and COI are suitable for unambiguous identification, whereas 16S failed to discriminate closely related flatfish and gurnard species. In course of probe design for DNA microarray development, each of the markers yielded a high number of potentially species-specific probes in silico, although many of them were rejected based on microarray hybridisation experiments. None of the markers provided probes to discriminate the sibling flatfish and gurnard species. However, since 16S-probes were less negatively influenced by the "position of label" effect and showed the lowest rejection rate and the highest mean signal intensity, 16S is more suitable for DNA microarray probe design than cty b and COI. The large portion of rejected COI-probes after hybridization experiments (, SCOPUS: ar.j, info:eu-repo/semantics/published

In 2006, we sampled the anoxic bottom waters of a volcanic lake beneath the Vatnajokull ice cap (Iceland). The sample contained 5 x 10(5) cells per ml, and whole-cell fluorescent in situ hybridization (FISH) and PCR with domain-specific probes showed these to be essentially all bacteria, with no detectable archaea. Pyrosequencing of the V6 hypervariable region of the 16S ribosomal RNA gene, Sanger sequencing of a clone library and FISH-based enumeration of four major phylotypes revealed that the assemblage was dominated by a few groups of putative chemotrophic bacteria whose closest cultivated relatives use sulfide, sulfur or hydrogen as electron donors, and oxygen, sulfate or CO(2) as electron acceptors. Hundreds of other phylotypes are present at lower abundance in our V6 tag libraries and a rarefaction analysis indicates that sampling did not reach saturation, but FISH data limit the remaining biome to

The microbial diversity of intertidal hot springs on the seashore of northwest Iceland was examined by combining directed in situ enrichments, artificial support colonization, and mat sampling. Analysis of 16S rRNA genes revealed the presence of clones related to both marine and terrestrial, thermophilic, mesophilic, and psychrophilic microorganisms scattered among 11 bacterial divisions. No archaea were found. The species composition of the enrichments was affected by the length of the hot periods experienced at low tide and was very different from those found in the biomass. A total of 36 chitinase genes were detected by molecular screening of the samples with degenerate primers for glycoside hydrolase family 18. The chitinase gene diversity was at least twofold higher in the enrichment samples than in the controls, indicating that a much higher diversity of hydrolytic genes can be accessed with this approach.

A family 18 chitinase gene chiA from the thermophile Rhodothermus marinus was cloned and expressed in Escherichia coli. The gene consisted of an open reading frame of 1,131 nucleotides encoding a protein of 377 amino acids with a calculated molecular weight of 42,341 Da. The deduced ChiA was a non-modular enzyme with one unique glycoside hydrolase family 18 catalytic domain. The catalytic domain exhibited 43% amino acid identity with Bacillus circulans chitinase C. Due to poor expression of ChiA, a signal peptide-lacking mutant, chiADeltasp, was designed and used subsequently. The optimal temperature and pH for chitinase activity of both ChiA and ChiADeltasp were 70 degrees C and 4.5-5, respectively. The enzyme maintained 100% activity after 16 h incubation at 70 degrees C, with half-lives of 3 h at 90 degrees C and 45 min at 95 degrees C. Results of activity measurements with chromogenic substrates, thin-layer chromatography, and viscosity measurements demonstrated that the chitinase is an endoacting enzyme releasing chitobiose as a major end product, although it acted as an exochitobiohydrolase with chitin oligomers shorter than five residues. The enzyme was fully inhibited by 5 mM HgCl2, but excess ethylenediamine tetraacetic acid relieved completely the inhibition. The enzyme hydrolyzed 73% deacetylated chitosan, offering an attractive alternative for enzymatic production of chitooligosaccharides at high temperature and low pH. Our results show that the R. marinus chitinase is the most thermostable family 18 chitinase isolated from Bacteria so far.

Geothermal energy has been harnessed and used for domestic heating in Iceland. In wells that are typically drilled to a depth of 1,500 to 2,000 m, the temperature of the source water is 50 to 130°C. The bottoms of the boreholes can therefore be regarded as subterranean hot springs and provide a unique opportunity to study the subterranean biosphere. Large volumes of geothermal fluid from five wells and a mixture of geothermal water from 50 geothermal wells (hot tap water) were sampled and concentrated through a 0.2-μm-pore-size filter. Cells were observed in wells RG-39 (91.4°C) and MG-18 (71.8°C) and in hot tap water (76°C), but no cells were detected in wells SN-4, SN-5 (95 to 117°C), and RV-5 (130°C). Archaea and Bacteria were detected by whole-cell fluorescent in situ hybridization. DNAs were extracted from the biomass, and small-subunit rRNA genes (16S rDNAs) were amplified by PCR using primers specific for the Archaea and Bacteria domains. The PCR products were cloned and sequenced. The sequence analysis showed 11 new operational taxonomic units (OTUs) out of 14, 3 of which were affiliated with known surface OTUs. Samples from RG-39 and hot tap water were inoculated into enrichment media and incubated at 65 and 85°C. Growth was observed only in media based on geothermal water. 16S rDNA analysis showed enrichments dominated with Desulfurococcales relatives. Two strains belonging to Desulfurococcus mobilis and to the Thermus / Deinococcus group were isolated from borehole RG-39. The results indicate that subsurface volcanic zones are an environment that provides a rich subsurface for novel thermophiles.

With the submersible JAGO and by scuba diving we discovered three remarkable geothermal cones, rising 33, 25, and 45 m from the seafloor at a depth of 65 m in Eyjafjordur, northern Iceland. The greatest geothermal activity was on the highest cone, which discharged up to 50 liters of freshwater per s at 72°C and pH 10.0. The cones were built up from precipitated smectite, formed by mixing of the hot SiO2-rich geothermal fluid with the cold Mg-rich seawater. By connecting a rubber hose to one outflow, about 240 liters of pure geothermal fluids was concentrated through a 0.2-μm-pore-size filter. Among 50 thermophilic isolates, we found members ofBacillusandThermonemaand a new unidentified low-G+C gram-positive member of theBacteriaas well as one member of theArchaea,Desulfurococcus mobilis. Analysis of small-subunit rRNA genes PCR amplified and cloned directly from environmental DNA showed that 41 out of 45Bacteriasequences belonged to members of theAquificales, whereas all of the 10Archaeasequences belonged to theKorarchaeota. The physiological characteristics of isolates from different parts of the cones indicate a completely freshwater habitat, supporting the possibility of subterranean transmittance of terrestrial organisms.

In solfataric fields in southwestern Iceland, neutral and sulfide-rich hot springs are characterized by thick bacterial mats at 60 to 80°C that are white or yellow from precipitated sulfur (sulfur mats). In low-sulfide hot springs in the same area, grey or pink streamers are formed at 80 to 90°C, and a Chloroflexus mat is formed at 65 to 70°C. We have studied the microbial diversity of one sulfur mat (high-sulfide) hot spring and one Chloroflexus mat (low-sulfide) hot spring by cloning and sequencing of small-subunit rRNA genes obtained by PCR amplification from mat DNA. Using 98% sequence identity as a cutoff value, a total of 14 bacterial operational taxonomic units (OTUs) and 5 archaeal OTUs were detected in the sulfur mat; 18 bacterial OTUs were detected in the Chloroflexus mat. Although representatives of novel divisions were found, the majority of the sequences were >95% related to currently known sequences. The molecular diversity analysis showed that Chloroflexus was the dominant mat organism in the low-sulfide spring (1 mg liter −1 ) below 70°C, whereas Aquificales were dominant in the high-sulfide spring (12 mg liter −1 ) at the same temperature. Comparison of the present data to published data indicated that there is a relationship between mat type and composition of Aquificales on the one hand and temperature and sulfide concentration on the other hand.

The whole-cell protein inventory of the deep-sea barophilic hyperthermophile Thermococcus barophilus was examined by one-dimensional SDS gradient gel electrophoresis when grown under different pressure conditions at 85 degrees C (Topt). One protein (P60) with a molecular mass of approximately 60 kDa was prominent at low pressures (0.3 MPa hydrostatic pressure and 0.1 MPa atmospheric pressure) but not at deep-sea pressures (10, 30, and 40 MPa). About 17 amino acids were sequenced from the N-terminal end of the protein. Sequence homology analysis in the GenBank database showed that P60 most closely resembled heat-shock proteins in some sulfur-metabolizing Archaea. A high degree of amino acid identity (81%-93%) to thermosome subunits in Thermococcales strains was found. Another protein (P35) with molecular mass of approximately 35.5 kDa was induced at 40 MPa hydrostatic pressure but not under low-pressure conditions. No amino acid sequence homology was found for this protein when the 40 amino acids from the N-terminal end were compared with homologous regions of proteins from databases. A PTk diagram was generated for T. barophilus. The results suggest that Phabitat is about 35 MPa, which corresponds to the in situ pressure where the strain was obtained.

We examined a single, non-spore-forming, aerobic, thermophilic strain that was isolated from a deep-sea hydrothermal vent in the Guaymas Basin at a depth of 2000 m and initially placed in a phenetic group with Thermus scotoductus (X-1). We identified this deep-sea isolate as a new strain belonging to Thermus thermophilus using several parameters. DNA-DNA hybridization under stringent conditions showed 74% similarity between the deep-sea isolate and T. thermophilus HB-8T (T = type strain). Phenotypic characteristics, such as the utilization of carbon sources, hydrolysis of different compounds, and antibiotic sensitivity were identical in the two strains. The polar lipids composition showed that strain Gy1211 belonged to the genus Thermus. The fatty acids composition indicated that this strain was related to the marine T. thermophilus strain isolated from the Azores. The new isolate T. thermophilus strain Gy1211 grew optimally at 75 degrees C, pH 8.0; and 2% NaCl. A hydrostatic pressure of 20 MPa, similar to the in situ hydrostatic pressure of the deep-sea vent from which the strain was isolated, had no effect on growth. Strain HB-8T, however, showed slower growth under these conditions.

Prokaryotic life is controlled by many parameters that play a role in every colonized habitat. However, only a few biotopes are exposed to elevated pressures. That is the case for most of the oceans whose average depth is 3800 m, which corresponds to a hydrostatic pressure of 38 MPa. However, recently discovered subsurface biotopes are also exposed to high pressures. These biotopes are colonized by procaryotic organisms that display different responses to high pressures, mostly depending on their origins. Most of the studies dealing with pressure on microorganisms living in high pressure habitats concern psychrophiles (usual inhabitants of the deep-sea) and some hyperthermophiles isolated from deep-sea hydrothermal vents. The organisms so far isolated and described, their physiology and adaptations to high pressure conditions will be presented.

A novel, hyperthermophilic, anaerobic, sulfurmetabolizing archaeon was isolated from a fluid sample from recently discovered hydrothermal vents in the North Fiji basin (SW Pacific), at 2000 m depth. The new organism, strain GE5, is a gram-negative, highly motile coccus. It grows between 67° and 102°C under atmospheric pressure, with an optimum at 96°C (doubling time 33 min). The upper growth temperature is extended by at least 3°C when cells are cultivated under in situ hydrostatic pressures (20 MPa). Strain GE5 is an obligate heterotroph, fermenting peptides, or mixtures of amino acids to acetate, isovalerate, isobutyrate, propionate, H2 and CO2. Hydrogen inhibits growth unless sulfur is present. In the presence of sulfur, H2S is then produced. Phylogenetic analyses of the 16 S rRNA sequence of strain GE5 places the new isolate within the Thermococcales. By its high growth temperature and physiological features the new isolate ressembles Pyrococcus sp. However it deffers by a 7% mol upper G+C-content and shows low level of DNA similarity with the two previously described species. Based on these differences the description of strain GE5 as a new species Pyrococcus abyssi (CNCM I-1302) is proposed.

Background The bacteriology during storage of the North-Atlantic cod has been investigated for the past decades using conventional cultivation strategies which have generated large amount of information. This paper presents a study where both conventional cultivation and cultivation independent approaches were used to investigate the bacterial succession during storage of cod loins at chilled and superchilled temperatures. Results Unbrined (0.4% NaCl) and brined (2.5% NaCl) cod loins were stored at chilled (0°C) and superchilled (-2 and -3.6°C) temperatures in air or modified atmosphere (MA, % CO2/O2/N2: 49.0 ± 0.6/7.4 ± 0.2/43.7 ± 0.4). Discrepancy was observed between cultivation enumeration and culture independent methods where the former showed a general dominance of Pseudomonas spp. (up to 59%) while the latter showed a dominance of Photobacterium phosphoreum (up to 100%). Gas chromatography-mass spectrophotometry (GC-MC) showed that trimethylamine was the most abundant volatile in mid- and late storage periods. Terminal restriction polymorphism (t-RFLP) analysis showed that the relative abundance of P. phosphoreum increased with storage time. Conclusion The present study shows the bacteriological developments on lightly salted or non-salted cod loins during storage at superchilled temperatures. It furthermore confirms the importance of P. phosphoreum as a spoilage organism during storage of cod loins at low temperatures using molecular techniques. The methods used compensate each other, giving more detailed data on bacterial population developments during spoilage.

Pseudomonas spp. is a group of microorganisms commonly found in fish and other fresh foods and is involved in their spoilage process. The aim of this study was to develop a rapid and accurate quantitative assay for Pseudomonas spp. in fish using real-time PCR. The assay targets the carbamoyl phosphate synthase gene (carA) with SYBR green based real-time PCR. The selectivity of the assay was confirmed using 24 Pseudomonas strains and 55 non-pseudomonad strains. A linear quantification was established over seven orders of magnitude, from 40 - 4(7) copies reaction(-1). The assay was validated on cod samples collected during two shelf life trials and showed a high degree of correlation to the plate count method (rP = 0.891) where the difference between the methods was 0.04 log(10) CFU g(-1) on average. The study shows that it is possible to quantify accurately the specific spoilage organisms belonging to the genus Pseudomonas in fish using real-time PCR. The method takes less than 5 h from sampling to results. The short detection time of the method can provide the fish industry with an important tool for quality control and processing management.