Methane-carbon flow into the benthic food web at cold seeps – a case study from the Costa Rica subduction zone

Cold seep ecosystems can support enormous biomasses of free-living and symbiotic chemoautotrophic organisms that get their energy from the oxidation of methane or sulfide. Most of this biomass derives from animals that are associated with bacterial symbionts, which are able to metabolize the chemical resources provided by the seeping fluids. Often these systems also harbor dense accumulations of non-symbiotic megafauna, which can be relevant in exporting chemosynthetically fixed carbon from seeps to the surrounding deep sea. Here we investigated the carbon sources of lithodid crabs (Paralomis sp.) feeding on thiotrophic bacterial mats at an active mud volcano at the Costa Rica subduction zone. To evaluate the dietary carbon source of the crabs, we compared the microbial community in stomach contents with surface sediments covered by microbial mats. The stomach content analyses revealed a dominance of epsilonproteobacterial 16S rRNA gene sequences related to the free-living and epibiotic sulfur oxidiser Sulfurovum sp. We also found Sulfurovum sp. as well as members of the genera Arcobacter and Sulfurimonas in mat-covered surface sediments where Epsilonproteobacteria were highly abundant constituting 10% of total cells. Furthermore, we detected substantial amounts of bacterial fatty acids such as i-C15:0 and C17:1ω6c with stable carbon isotope compositions as low as −53‰ in the stomach and muscle tissue. These results indicate that the white microbial mats at Mound 12 are comprised of Epsilonproteobacteria and that microbial mat-derived carbon provides an important contribution to the crab's nutrition. In addition, our lipid analyses also suggest that the crabs feed on other 13C-depleted organic matter sources, possibly symbiotic megafauna as well as on photosynthetic carbon sources such as sedimentary detritus

First Investigation of the Microbiology of the Deepest Layer of Ocean Crust

We would like to thank Frederick (Rick) Colwell for input on molecular analyses in low biomass environments, Donna Blackman, Benoît Ildefonse, Adélie Delacour, and Gretchen Früh-Green for discussions regarding geological and geochemical aspects of this manuscript, and the Integrated Ocean Drilling Program Expeditions 304/305 Science Party. We would also like to thank Captain Alex Simpson and the entire crew of the JOIDES Resolution.Conceived and designed the experiments: OUM MRF SJG. Performed the experiments: OUM TN MR JDVN AM. Analyzed the data: OUM TN MR JDVN AM. Contributed reagents/materials/analysis tools: TN MR JZ MRF SJG. Wrote the paper: OUM.The gabbroic layer comprises the majority of ocean crust. Opportunities to sample this expansive crustal environment are rare because of the technological demands of deep ocean drilling; thus, gabbroic microbial communities have not yet been studied. During the Integrated Ocean Drilling Program Expeditions 304 and 305, igneous rock samples were collected from 0.45-1391.01 meters below seafloor at Hole 1309D, located on the Atlantis Massif (30 °N, 42 °W). Microbial diversity in the rocks was analyzed by denaturing gradient gel electrophoresis and sequencing (Expedition 304), and terminal restriction fragment length polymorphism, cloning and sequencing, and functional gene microarray analysis (Expedition 305). The gabbroic microbial community was relatively depauperate, consisting of a low diversity of proteobacterial lineages closely related to Bacteria from hydrocarbon-dominated environments and to known hydrocarbon degraders, and there was little evidence of Archaea. Functional gene diversity in the gabbroic samples was analyzed with a microarray for metabolic genes (“GeoChip”), producing further evidence of genomic potential for hydrocarbon degradation - genes for aerobic methane and toluene oxidation. Genes coding for anaerobic respirations, such as nitrate reduction, sulfate reduction, and metal reduction, as well as genes for carbon fixation, nitrogen fixation, and ammonium-oxidation, were also present. Our results suggest that the gabbroic layer hosts a microbial community that can degrade hydrocarbons and fix carbon and nitrogen, and has the potential to employ a diversity of non-oxygen electron acceptors. This rare glimpse of the gabbroic ecosystem provides further support for the recent finding of hydrocarbons in deep ocean gabbro from Hole 1309D. It has been hypothesized that these hydrocarbons might originate abiotically from serpentinization reactions that are occurring deep in the Earth's crust, raising the possibility that the lithic microbial community reported here might utilize carbon sources produced independently of the surface biosphere.Yeshttp://www.plosone.org/static/editorial#pee

Optimization of PCR amplification for sensitive capture of Methanopyrus isoleucyl-tRNA synthetase gene in environmental samples

Microbes are abundant in marine and terrestrial ecosystems. However, the available evidence indicates that most microbial species in the environment cannot be cultivated. Accordingly, methodologies for characterizing genomes in environmental samples directly without cultivation have become foremost in importance. In this study, experimental conditions using PCR amplification to characterize such genomes were investigated with respect to the detection of target sequences present at low frequencies. Amplification of the isoleucyl-tRNA synthetase (IleRS) gene in Methanopyrus isolates was optimized regarding the choice of DNA polymerase, target sequence size, PCR primer size and the use of degenerate primers. Detection of IleRS was most sensitive when the target sequence size was about 1 kb. Among KOD, Taq and Pfu DNA polymerases, Pfu displayed the lowest efficiency, while KOD and Taq showed similar efficiency. Primers no shorter than 18mers gave satisfactory results. In addition, PCR bias caused by primer degeneracy could be alleviated by varying the balance between different added primers. Employing a combination of Taq DNA polymerase, a target DNA sequence approximately 1 kb in size, and primers of 20mer in length, PCR amplification of the IleRS gene could be achieved with as little as 2 pg pure isolated DNA template

Dynamics and diversity of newly produced virioplankton in the North Sea

Viral diversity has been studied in a variety of marine habitats and spatial and seasonal changes have been documented. Most of the bacteriophages are considered host specific and are thought to affect fast growing prokaryotic phylotypes more than slow growing ones. We hypothesized that viral infection and consequently, lysis occurs in pulses with only a few prokaryotic phylotypes lysed at any given time. Thus, we propose that the newly produced viruses represent only a fraction of the viral diversity present at any given time. Virioplankton diversity was assessed by pulsed-field gel electrophoresis in the surface waters of three distinct areas of the North Sea during the spring and summer. Bulk virioplankton diversity was fairly stable in these waters. Viral diversity produced by the indigenous bacterioplankton, however, exhibited day-to-day variability with only a few bands produced at any given time. These bands frequently matched bands of the in situ virioplankton; however, bands not present in the band pattern of the in situ virioplankton community were also found. These new bands probably indicate infection and subsequent release of viruses from bacterioplankton phylotypes previously not infected by these specific viruses. Overall, our results demonstrate that viral infection and lysis are rather dynamic processes. The main targets of viral infection are changing apparently on time scales of hours to days indicating that viral infection might effectively regulate and maintain bacterioplankton diversity