Costantino Vetriani

Fingerprinting microbial assemblages from the oxic/anoxic chemocline of the Black Sea.

ABSTRACT Biomass samples from the Black Sea collected in 1988 were analyzed for SSU genes from Bacteria and Archaea after 10 years of storage at −80°C. Both clonal libraries and direct fingerprinting by terminal restriction fragment length polymorphism (T-RFLP) analyses were used to assess the microbial community. Uniform and discrete depth distributions of different SSU phylotypes were observed. However, most recombinant clones were not restricted to a specific depth in the water column, and many of the major T-RFLP peaks remain uncharacterized. Of the clones obtained, an ε-Proteobacteria and a Pseudoalteromonas-like clone accounted for major peaks in the fingerprint, while deeply branching lineages of α- and γ-Proteobacteria were associated with smaller peaks. Additionally, members were found among both the δ-Proteobacteria related to sulfate reducers and the Archaea related to phylotypes from the ANME groups that anaerobically oxidize methane.

Isolation and characterization of Erythrobacter sp. strains from the upper ocean

Seven strains of marine aerobic anoxygenic phototrophs belonging to the genus Erythrobacter were isolated. The strains were characterized regarding their physiological and biochemical properties, 16S rDNA and pufM gene sequences, morphological features, substrate preference, as well as pigment and lipid composition. All strains had functional type-2 reaction centers containing bacteriochlorophyll, served by small, light-harvesting complex 1, and were photosynthetically competent. In addition, large pools of carotenoids were found, but only some of the accessory pigments transfer energy to the reaction centers. All of the isolates were facultative photoheterotrophs. They required an organic carbon substrate for growth; however, they are able to supplement a significant fraction of their metabolic requirements with photosynthetically derived energy.

Mercury Adaptation among Bacteria from a Deep-Sea Hydrothermal Vent

ABSTRACT Since deep-sea hydrothermal vent fluids are enriched with toxic metals, it was hypothesized that (i) the biota in the vicinity of a vent is adapted to life in the presence of toxic metals and (ii) metal toxicity is modulated by the steep physical-chemical gradients that occur when anoxic, hot fluids are mixed with cold oxygenated seawater. We collected bacterial biomass at different distances from a diffuse flow vent at 9°N on the East Pacific Rise and tested these hypotheses by examining the effect of mercuric mercury [Hg(II)] on vent bacteria. Four of six moderate thermophiles, most of which were vent isolates belonging to the genus Alcanivorax, and six of eight mesophiles from the vent plume were resistant to >10 μM Hg(II) and reduced it to elemental mercury [Hg(0)]. However, four psychrophiles that were isolated from a nearby inactive sulfide structure were Hg(II) sensitive. A neighbor-joining tree constructed from the deduced amino acids of a PCR-amplified fragment of merA, the gene encoding the mercuric reductase (MR), showed that sequences obtained from the vent moderate thermophiles formed a unique cluster (bootstrap value, 100) in the MR phylogenetic tree, which expanded the known diversity of this locus. The temperature optimum for Hg(II) reduction by resting cells and MR activity in crude cell extracts of a vent moderate thermophile corresponded to its optimal growth temperature, 45°C. However, the optimal temperature for activity of the MR encoded by transposon Tn501 was found to be 55 to 65°C, suggesting that, in spite of its original isolation from a mesophile, this MR is a thermophilic enzyme that may represent a relic of early evolution in high-temperature environments. Results showing that there is enrichment of Hg(II) resistance among vent bacteria suggest that these bacteria have an ecological role in mercury detoxification in the vent environment and, together with the thermophilicity of MR, point to geothermal environments as a likely niche for the evolution of bacterial mercury resistance.

THE MESOZOIC RADIATION OF EUKARYOTIC ALGAE: THE PORTABLE PLASTID HYPOTHESIS1

Although all chloroplasts appear to have been derived from a common ancestor, a major schism occurred early in the evolution of eukaryotic algae that gave rise to red and green photoautotrophic lineages. In Paleozoic and earlier times, the fossil record suggests that oceanic eukaryotic phytoplankton were dominated by the green (chl b‐containing) algal line. However, following the end‐Permian extinction, a diverse group of eukaryotic phytoplankton evolved from secondary symbiotic associations in the red (chl c‐containing) line and subsequently rose to ecological prominence. In the contemporary oceans, red eukaryotic phytoplankton taxa continue to dominate marine pelagic food webs, whereas the green line is relegated to comparatively minor ecological and biogeochemical roles. To help elucidate why the oceans are not dominated by green taxa, we analyzed and compared whole plastid genomes in both the red and green lineages. Our results suggest that whereas all algal plastids retain a core set of genes, red plastids retain a complementary set of genes that potentially confer more capacity to autonomously express proteins regulating oxygenic photosynthetic and energy transduction pathways. We hypothesize that specific gene losses in the primary endosymbiotic green plastid reduced its portability for subsequent symbiotic associations. This corollary of the plastid “enslavement” hypothesis may have limited subsequent evolutionary advances in the green lineage while simultaneously providing a competitive advantage to the red lineage.

Vertical distribution and diversity of bacteria and archaea in sulfide and methane-rich cold seep sediments located at the base of the Florida Escarpment

The bacterial and archaeal communities of the sediments at the base of the Florida Escarpment (Gulf of Mexico, USA) were investigated using molecular phylogenetic analysis. The total microbial community DNA of each of three vertical zones (top, middle and bottom) of a sediment core was extracted and the 16S rRNA genes were amplified by PCR, cloned and sequenced. Shannon–Weaver Diversity measures of bacteria were high in all three zones. For the archaea, diversity was generally low, but increased with depth. The archaeal clonal libraries were dominated by representatives of four groups of organisms involved in the anaerobic oxidation of methane (ANME groups). Phylogenetic analysis of bacteria suggests the dominance of ε-proteobacteria in the top zone, the ε-, δ- and γ-proteobacteria in the middle zone and the δ-proteobacteria in the bottom zone of the core. Members of the Cytophaga–Flexibacter–Bacteroidetes group, the Chloroflexi/green non-sulfur bacteria, the Gram+ (Firmicutes), the Planctomyces, candidate division WS3 and Fusobacterium were also detected. Our data suggest that the community structure and diversity of microorganisms can shift greatly within small vertical distances, possibly in response to changes in the physical and chemical conditions.

Culture dependent and independent analyses of 16S rRNA and ATP citrate lyase genes: a comparison of microbial communities from different black smoker chimneys on the Mid-Atlantic Ridge

The bacterial and archaeal communities of three deep-sea hydrothermal vent systems located on the Mid-Atlantic Ridge (MAR; Rainbow, Logatchev and Broken Spur) were investigated using an integrated culture-dependent and independent approach. Comparative molecular phylogenetic analyses, using the 16S rRNA gene and the deduced amino acid sequences of the alpha and beta subunits of the ATP citrate lyase encoding genes were carried out on natural microbial communities, on an enrichment culture obtained from the Broken Spur chimney, and on novel chemolithoautotrophic bacteria and reference strains originally isolated from several different deep-sea vents. Our data showed that the three MAR hydrothermal vent chimneys investigated in this study host very different microbial assemblages. The microbial community of the Rainbow chimney was dominated by thermophilic, autotrophic, hydrogen-oxidizing, sulfur- and nitrate-reducing Epsilonproteobacteria related to the genus Caminibacter. The detection of sequences related to sulfur-reducing bacteria and archaea (Archaeoglobus) indicated that thermophilic sulfate reduction might also be occurring at this site. The Logatchev bacterial community included several sequences related to mesophilic sulfur-oxidizing bacteria, while the archaeal component of this chimney was dominated by sequences related to the ANME-2 lineage, suggesting that anaerobic oxidation of methane may be occurring at this site. Comparative analyses of the ATP citrate lyase encoding genes from natural microbial communities suggested that Epsilonproteobacteria were the dominant primary producers using the reverse TCA cycle (rTCA) at Rainbow, while Aquificales of the genera Desulfurobacterium and Persephonella were prevalent in the Broken Spur chimney.

Interrelationships Between Vent Fluid Chemistry, Temperature, Seismic Activity, and Biological Community Structure at a Mussel-Dominated, Deep-Sea Hydrothermal Vent Along the East Pacific Rise

Abstract In April 1991, submarine volcanic eruptions initiated the formation of numerous hydrothermal vents between 9°45′ and 9°52′N along the crest of the East Pacific Rise (EPR). Dramatic changes in biological community structure and vent fluid chemistry have been documented throughout this region since the eruptive event. By April 2004, mussels (Bathymodiolus thermophilus) dominated the faunal assemblages at several of the vent sites formed during of after the 1991 eruptions, whereas other habitats within the region were dominated by the vestimentiferan Riftia pachyptila. In the present paper, we build upon the extensive data sets obtained at these sites over the past decade and describe a manipulative experiment (conducted at 9°49.94′N; 104°14.43′W on the EPR) designed to assess interrelationships between vent fluid chemistry, temperature, biological community structure, and seismic activity. To this end, in situ voltammetric systems and thermal probes were used to measure H2S/HS− and temperature over time in a denuded region of an extensive mussel bed in which an exclusion cage was placed to inhibit the subsequent migration of mussels into the denuded area. Fluid samples were taken from the same locations to characterize the associated microbial constituents. Basalt blocks, which were placed in the cage in April 2004 and subsequently recovered in April 2005, were colonized by more than 25 different species of invertebrates, including numerous vestimentiferans and remarkably few mussels. Recorded temporal changes in vent fluid chemistry and temperature regimes, when coupled with microbiological characterization of the vent fluids and seismic activity data obtained from ocean bottom seismometers, shed considerable light on factors controlling biological community structure in these hydrothermal ecosystems.

Hydrothermal Vent Mussel Habitat Chemistry, Pre- and Post-Eruption at 9°50′North on the East Pacific Rise

Abstract Between October 2005 and March 2006, a seafloor volcanic eruption occurred at 9°50′N East Pacific Rise (EPR), establishing a “time zero” for characterizing newly-formed hydrothermal vent habitats and comparing them to pre-eruption habitats. Before the eruption, mussels (Bathymodiolus thermophilus) formed large aggregates between 9°49.6′ and 9°50.3′N. After the eruption, the few mussels remaining were in sparsely-distributed individuals and clumps, seemingly transported via lava flows or from mass wasting of the walls of the axial trough. In situ voltammetry with solid state gold-amalgam microelectrodes was used to characterize the chemistry of vent fluids in mussel habitats from 2004 to 2007, providing data sets for comparison of oxygen, sulfide, and temperature. Posteruption fluids contained higher sulfide-to-temperature ratios (i.e., slopes of linear regressions) (10.86 μM °C−1) compared with pre-eruption values in 2004 and 2005 (2.79 μM °C−1 and −0.063 μM °C−1, respectively). These chemical differences can be attributed to the difference in geographic location in which mussels were living and physical factors arising from posteruptive fluid emissions.

Ammonificins A and B, Hydroxyethylamine Chroman Derivatives from a Cultured Marine Hydrothermal Vent Bacterium, Thermovibrio ammonificans

Two hydroxyethylamine chroman derivatives, ammonificins A (1) and B (2), were isolated from the marine hydrothermal vent bacterium Thermovibrio ammonificans. The molecular structures of these compounds were determined using a combination of NMR, mass spectrometry, and CD analyses. Biological activities were determined using an antimicrobial assay and the patented ApopScreen cell-based screen for apoptosis induction and potential anticancer activity. To our knowledge, this is the first report of secondary metabolites from the marine hydrothermal vent bacterium T. ammonificans.

Deep-sea hydrothermal vent Epsilonproteobacteria encode a conserved and widespread nitrate reduction pathway (Nap)

Despite the frequent isolation of nitrate-respiring Epsilonproteobacteria from deep-sea hydrothermal vents, the genes coding for the nitrate reduction pathway in these organisms have not been investigated in depth. In this study we have shown that the gene cluster coding for the periplasmic nitrate reductase complex (nap) is highly conserved in chemolithoautotrophic, nitrate-reducing Epsilonproteobacteria from deep-sea hydrothermal vents. Furthermore, we have shown that the napA gene is expressed in pure cultures of vent Epsilonproteobacteria and it is highly conserved in microbial communities collected from deep-sea vents characterized by different temperature and redox regimes. The diversity of nitrate-reducing Epsilonproteobacteria was found to be higher in moderate temperature, diffuse flow vents than in high temperature black smokers or in low temperatures, substrate-associated communities. As NapA has a high affinity for nitrate compared with the membrane-bound enzyme, its occurrence in vent Epsilonproteobacteria may represent an adaptation of these organisms to the low nitrate concentrations typically found in vent fluids. Taken together, our findings indicate that nitrate reduction is widespread in vent Epsilonproteobacteria and provide insight on alternative energy metabolism in vent microorganisms. The occurrence of the nap cluster in vent, commensal and pathogenic Epsilonproteobacteria suggests that the ability of these bacteria to respire nitrate is important in habitats as different as the deep-sea vents and the human body.