Hydrothermal circulation extracts a significant fraction of lithospheric heat from the ocean crust, with most of this advective heat loss occurring on ridge flanks, far from mid-ocean ridges. Faults in ocean crust are common in many settings, and may serve as high-transmissivity structures that facilitate advective transport and focus discharge of fluid, heat, and solutes below and at the seafloor. Coupled flow along fault zones has been invoked in a variety of settings, but circulation patterns are not well constrained by observational data or earlier models. We present results from three-dimensional, fully coupled numerical simulations of fluid and heat flow in sediment-covered ridge-flank ocean crust cut by a fault. We explore a range of fault and surrounding crustal characteristics, including crust and fault permeability, fault dip angle, thickness, and depth. We are particularly interested in resolving relations between fault and crustal characteristics and seafloor heat flux patterns.
Simulation results show variability in patterns of fluid circulation and seafloor heat flux as a function of fault geometry and crustal properties. The seafloor heat flux pattern above fault traces tends to show variability along strike (in response to underlying regions of rapid upward and downward flow along the fault trace), and asymmetry in seafloor heat flux anomalies, with higher values above the fault trace and lower values in the immediately surrounding seafloor, especially above the hanging wall. The negative anomaly is generally greater when the fault dip angle is lower.
Higher permeability in the crustal rocks adjacent to the fault zone tend result in small-scale convection and small-amplitude variations in seafloor heat flux, and more diffuse convection cells in the fault zone itself. Convection in the surrounding crust decreases the importance of the fault zone in extracting lithospheric heat. Simulations also show that faults that penetrate deeper into the crust produce a significantly larger seafloor heat flux anomaly than do shallower faults, indicating that deeper faults extract lithospheric heat more efficiently. Patterns of seafloor heat flux from these simulations indicate that fault-zone hydrothermal circulation should produce thermal anomalies that are detectable in field measurements. Linking field observations directly to numerical simulations can provide better understanding of the geometry and properties of faults and fluid flow patterns in the volcanic ocean crust.
Water radiolysis is the dissociation of water molecules by ionizing radiation from the decay of radionuclides. Primary products of water radiolysis include reactive chemicals, such as H2 and H2O2. For this reason, radiolysis is studied in many domains, including nuclear waste, microbiology and planetary evolution. In order to understand the importance of radiolysis in many of these environments, accurate quantification of radiolytic production rates is vital. In this dissertation, I present a new quantitative model calculating radiolytic production rates at solid-water interfaces and apply it to understand the role that radiolysis plays in various environments. ^ This radiolytic model is the first to explicitly calculate radiolytic production due to α-, β- and γ-radiation near solid-water interfaces. We use this model to investigate the effects of radiolytic compounds on the dissolution rate of spent nuclear fuel. The production of rate of H2 and H2O2, which control the dissolution rate of the fuel, depends on the amount and type of radiation surrounding breached nuclear spent fuel rods. Understanding the distribution of radiolytic products is important in assessing different hazards associated with spent fuel storage and the potential release of radionuclides into the environment. We find that in old (1000-year-old) spent fuel α-radiation dominates radiolysis, while β- and γ-radiation control the production rates near young (20-year-old) spent fuel.^ Radiolysis also is an important process in understanding the extent of life on Earth, as well as possibly providing a means for life on Mars. We investigate the significance of water radiolysis in sustaining microbial communities in Earth’s oceanic crust and the potential extent of radiolysis in wet martian environments (such as the ancient martian surface and the present martian subsurface). These two studies focus specifically on the production of radiolytic H2 as an electron donor. H2 is an important source of energy in these two environments where there other resources for microbes are limited. In the oceanic basaltic aquifer of the South Pacific Gyre, we find that radiolytic H2 production yields depend largely on the width of fractures in basalt and on radionuclide concentrations. We show that in old seafloor (>10 Ma), where there are no other readily available electron donors, radiolytic H2 may dominate and is able to support up to 103 cells in the water adjacent to a square cm of basaltic fracture.^ The extent of water radiolysis on Mars can be determined for water-saturated martian environments, such as the ancient martian surface or the present martian subsurface. Using the fractured rock radiolytic model as well as a previously developed sediment radiolytic model, we calculate potential H2 production rates for eleven martian lithologies assuming contact with water. The highest rates on Mars are for water-saturated material with the radionuclide concentrations of Acidalia Planitia, a region with surface materials that are enriched in uranium and thorium. We also calculate production rates for the eight proposed Mars 2020 landing sites, assuming water-saturated porosity. Radiolytic H2 production rates calculated for wet martian sediment and water-filled microfractured rock are comparable to the range of rates calculated for Earth’s South Pacific basement basalt which is known to harbor low concentrations of microbial life.
Organic carbon (OC) preserved in marine sediments acts as a reduced carbon sink that balances the global carbon cycle. Understanding the biogeochemical mechanisms underpinning the balance between OC preservation and degradation is thus critical both to quantifying this carbon reservoir and to estimating the extent of life in the deep subsurface biosphere. This work utilizes bulk and spatially-resolved X-ray absorption spectroscopy to characterize the OC content and composition of various environmental systems in order to identify the role of minerals and surrounding geochemistry in organic carbon preservation in sediments. Biogenic manganese (Mn) oxides formed either in pure cultures of Mn-oxidizing microorganisms, in incubations of brackish estuarine waters, or as ferromanganese deposits in karstic cave systems rapidly associate with OC following precipitation. This association is stable despite Mn oxide structural ripening, suggesting that mineral-associated OC could persist during early diagenetic reactions. OC associated with bacteriogenic Mn oxides is primarily proteinaceous, including intact proteins involved in Mn oxidation and likely oxide nucleation and aggregation. Pelagic sediments from 16 sites underlying the South Pacific and North Atlantic gyres and spanning a gradient of sediment age and redox state were analyzed in order to contrast the roles of oxygen exposure, OC recalcitrance, and mineral-based protection of OC as preservation mechanisms. OC and nitrogen concentrations measured at these sites are among the lowest globally (<0.1%) and, to a first order, scale with sediment oxygenation. In the deep subsurface, however, molecular recalcitrance becomes more important than oxygen exposure time in protecting OC against remineralization. Deep OC consists of primarily amide and carboxylic carbon in a scaffolding of aliphatic and O-alkyl moieties, corroborating the extremely low C/N values observed. These findings suggest that microbes in oxic pelagic sediments are carbon-limited and may preferentially remove carbon relative to nitrogen from the organic matter pool. As a whole, this work documents how interactions with mineral surfaces and exposure to oxygen generate a reservoir of OC stabilized in sediments on at least 25-million year time scales.
Methane seeps are globally distributed geologic features in which reduced fluid from below the seafloor is advected upward and meets the oxidized bottom waters of Earth’s oceans. This redox gradient fuels chemosynthetic communities anchored by the microbially-mediated anaerobic oxidation of methane (AOM). Both today and in Earth’s past, methane seeps have supported diverse biological communities extending from microorgansisms to macrofauna and adding to the diversity of life on Earth. Simultaneously, the carbon cycling associated with methane seeps may have played a significant role in modulating ancient Earth’s climate, particularly by acting as a control on methane emissions.
The AOM metabolism generates alkalinity and dissolved inorganic carbon (DIC) and at a 2:1 ratio, promoting the abiogenic, or authigenic, precipitation of carbonate minerals. Over time, these precipitates can grow into pavements covering hundreds of square meters on the seafloor and dominating the volumetric habitat space available in seep ecosystems. Importantly, carbonates are incorporated into the geologic record and therefore preserve an inorganic (i.e., d13C) and organic (i.e., lipid biomarker) history of methane seepage. However, the extent to which preserved biomarkers represent a snapshot of microorganisms present at the time of primary precipitation, a time-integrated history of microbial assemblages across the life cycle of a methane seep, or a view of the final microorganisms inhabiting a carbonate prior to incorporation in the sedimentary record is unresolved.
This thesis addresses the ecology of carbonate-associated seep microorganisms. Chapters One and Two contextualize the extant microbial diversity on seep carbonates versus within seep sediments, as determined through 16S rRNA gene biomarkers. Small, protolithic carbonate “nodules” recovered from within seep sediments are observed to be capable of capturing surrounding sediment-hosted microbial diversity, but in some cases also diverge from sediments. Meanwhile, lithified carbonate blocks recovered from the seafloor host microbial assemblages demonstrably distinct from seep sediments (and seep nodules). Microbial 16S rRNA gene diversity within carbonate samples is well-differentiated by the extent of contemporary seepage. In situ seafloor transplantation experiments further demonstrated the microbial assemblages associated with seep carbonates to be sensitive to seep quiescence and activation on short (13-month) timescales. This was particularly true for organisms whose 16S rRNA genes imply physiologies dependent on methane or sulfur oxidation. With an improved understanding of the modern ecology of carbonate-associated microorganisms, Chapter Three applies intact polar lipid (IPL) and core lipid analyses to begin describing whether, and to what extent, geologically relevant biomarkers mimic short-term dynamics observed in 16S rRNA gene profiles versus archive a record of historic microbial diversity. Biomarker longevity is determined to increase from 16S rRNA genes to IPLs to core lipids, with IPLs preserving microbial diversity history on timescales more similar to 16S rRNA genes than core lipids. Ultimately, individual IPL biomarkers are identified which may be robust proxies for determining whether the biomarker profile recorded in a seep carbonate represents vestiges of active seepage processes, or the profile of a microbial community persisting after seep quiescence.
At the broadest scale, this thesis is an investigation of how life modulates the movement of essential elements (carbon, sulfur, nitrogen, and silicon) on modern and geologic timescales. Chapters 1 and 2 explore carbon and sulfur cycling microbial communities found centimeters below the seafloor in hydrocarbon-rich methane seep ecosystems. At the Hydrate Ridge methane seep, we investigated how microbial partnerships direct the flow of methane and sulfide in these benthic oases by using identity-based physical separation methods developed in our lab (Magneto-FISH) in conjunction with community profiling and metagenomic sequencing. This method explores the middle ground between single cell and bulk sediment analysis by separating target microbes and their physically associated community for downstream sequencing applications. Magneto-FISH captures were done at a range of microbial taxonomic group specificities and sequenced with both clone library and next-gen iTag 16S rRNA gene methods. Chapter 1 provides a demonstration of how FISH probe taxonomic specificity correlates to resultant Archaeal taxonomic diversity in Magneto-FISHed seep sediments, with specific attention to preparation of Archaea-enriched samples for downstream metagenomic sequencing. In Chapter 2, a Bacteria-focused parallel environmental isolation and sequencing effort was subjected to co-occurrence analyses which suggested there may be far more microbial associations in methane seep systems than are currently appreciated, including partnerships that do not involve the canonical anaerobic methane oxidizing archaea and sulfate reducing bacteria. With samples from IODP Expedition 337 Shimokita coalbed biosphere, Chapter 3 provides evidence for an active microbial assemblage kilometers below the sea floor in the deepest samples ever collected by marine scientific ocean drilling. Using in situ temperature Stable Isotope Probing (SIP) incubations and NanoSIMS, we investigated whole community activity (with the passive tracer D2O) and substrate specific activity with C1-carbon compounds methylamine and methanol. We found deuterium-based turnover times to be faster (years) than previous deep biosphere estimates (hundreds to thousands of years), but methylotrophy rates to be slower than previous carbon metabolic rates.
Our knowledge of microbial life residing in the oceans- photic and aphoticand within the subseafloor sediment has grown exponentially within the last few decades. This is partly because of advances in next-generation sequencing technology, which has provided an opportunity to address previously unanswerable questions regarding microbial diversity and biogeography (Hamady & Knight, 2009; Petrosino, Highlander, Luna, Gibbs, & Versalovic, 2009). By utilizing a next-generation sequencing approach, I determined microbial community compositions and assessed their response to environmental and geographic variation within and between five different oceanic regimes (i) the South Pacific Gyre (SPG), (ii) the Eastern and Central Equatorial Pacific (EQP), (iii) the North Pacific Gyre (NPG), (iv) the Bering Sea (U1343), and (v) the Indian Ocean (NGHP-1-14). My first manuscript, “ The bacterial and archaeal biogeography of the deep chlorophyll maximum of the South Pacific Gyre”, examines the prokaryotic community composition at a continuous and biologically significant horizon, the deep chlorophyll maximum (DCM), across Earth’s largest oceanographic province, the SPG. Our results demonstrate that bacterial and archaeal tag-sequence assemblages of the DCM are strikingly similar throughout the SPG, in terms of the presence and abundance of the most dominant bacterial taxa. Comparison of our SPG bacterial results to samples from the NPG and the relatively nutrient- and chlorophyll-rich EQP shows that DCM assemblages of the SPG, NPG and EQP are statistically distinct from each other, although they have many abundant tags in common. This distinctness is influenced by environmental conditions, as the communities of the two gyres (SPG and NPG) resemble each other more closely than either resembles the EQP community (which lives geographically between them).
My second manuscript, “Vertical changes in bacterial diversity and community composition from seasurface to subseafloor”, investigates the degree of connectivity between bacterial communities that reside in deep-sea sediment to those that reside throughout the water column at three Pacific Ocean (EQP and NPG) stations. In this study, my collaborators and I investigate a series of ecological gradients through examination into the vertical structure and richness of marine microbes and the how they are influenced by geographic location, light, oxygen concentration and depth. We provide the first pyrosequencing results to (i) address a possible mechanism by which deeply buried sedimentary communities develop deep beneath the seafloor and (ii) assess the degree to which the organisms in those communities are related to communities in the overlying ocean.
My third manuscript, “Bacterial diversity, sediment age and organic respiration in the marine sedimentary environment”, investigates the drivers of microbial diversity and taxonomic richness in deep subseafloor sediment of four geographically distant sites in the Pacific and Indian oceans (U1343, NGHP-1-14, EQP1 and EQP8). To accomplish this goal, my collaborators and I took samples for molecular analysis and interstitial water from a wide range of sediment depths (up to 404 meters below seafloor) and sediment age (up to 5.5 Ma). Our study of these samples demonstrates that abundance-weighted bacterial community composition shifts in response to availability of dissolved metabolic reactants (e.g. oxygen, sulfate, methane). Our study also demonstrates taxonomic richness declines exponentially with sediment age and generally matches the canonical expectation for changing rates of organic oxidation in subseafloor sediment over time (Jorgensen, 1978; Middelburg, 1989; Westrich & Berner, 1984).
Most of the hydrothermal circulation through the ocean crust, in terms of mass, heat, and many solute fluxes, occurs on ridge-flanks. Far from the magmatic influence of mid-ocean ridges, fluid flow is driven by lithospheric heating from below and channeled through volcanic rock outcrops that serve as high-permeability conduits between the ocean and the underlying volcanic crust. Field data in this setting is sparse due to difficulties associated with accessing these remote locations, making geologically accurate modeling particularly valuable to assessing the nature of ridge-flank hydrothermal circulation. Each study in this thesis applies a combination of modeling and field observations to constrain the hydrogeologic properties and behaviors of ridge-flank hydrothermal systems, including: (1) deriving permeability estimates from flowing subsea boreholes, (2) investigating the sustainability of outcrop-to-outcrop hydrothermal flow, and (3) constraining the properties and behaviors on a well-studied outcrop-to-outcrop system. In the first study, thermal records from flowing boreholes in young oceanic crust are used to assess borehole and formation properties, including permeability, using analytic equations and a Markov chain Monte Carlo analysis to quantify uncertainty. We find the median bulk permeability at all sites to be between 0.4 to 1.5 x 10^-11 m2, with a standard deviation of 0.2 to 0.3 log-cycles at each borehole. These results are remarkably homogenous, given the much larger variability in permeability measurements in the oceanic crust. Results from the second study illuminate the controls on hydrogeologic sustainability, flow rate, and preferred flow direction in outcrop-to-outcrop hydrothermal systems. We find that sustained flow between outcrops over tens of kilometers depends on a contrast in transmittance (the product of outcrop permeability and the area of outcrop exposure) between recharging and discharging sites, and that discharge is favored through less transmissive outcrops. These systems require aquifer permeability values ranging from 10^-12 to 10^-11 m2, consistent with field measurements and values inferred from the first chapter. In the third study, a suite of three-dimensional numerical simulations are used to characterize and constrain the permeability and thickness of the upper crustal aquifer, the permeability of outcrops, and the potential for multiple discharging outcrops and azimuthal permeability anisotropy to influence hydrothermal processes at a field site on the eastern flank of the Juan de Fuca Ridge.
When exposed at Earth’s surface, rocks are out of thermodynamic equilibrium with respect to their environment. This disequilibrium drives the chemical transformation, or weathering, of these rocks into soils and governs the chemical composition of natural waters and the atmosphere. Over geologic timescales, complex feedbacks associated with weathering processes are presumed to regulate the concentrations of CO₂ and O₂ in the atmosphere with profound implications for the habitability of the planet. However, a mechanistic understanding of how biologic, tectonic, and climatic conditions interact to control weathering fluxes has remained elusive. In part, our understanding of weathering processes is hindered by the fact that they operate continuously over an enormous range of spatial (atomic to global) and temporal (microseconds to millions of years) scales, but we can only make measurements over discrete ranges of these values. While each scale of observation available offers unique insights, it is often difficult to link observations made at different scales. For my Ph.D., I focused on three distinct projects that span the range of observable scales in order to better understand the links between chemical weathering and long-term biogeochemical cycles. ❧ Chapter 2: Laboratory insights into microbial mineral dissolution. Rocks and minerals represent a major reservoir of bio-essential nutrients. While abundant, some of these lithogenic nutrients, like iron, are not readily bio-available. As a result, many organisms produce metal-binding ligands to scavenge these trace nutrients from the environment. Using targeted laboratory experiments with live microbial cultures and purified microbial ligands, I explored efficacy by which microbes can access trace nutrients from common silicate minerals (Torres et al., in prep). In addition to providing insight into biological nutrient scavenging strategies, this work also provides the basic research necessary to develop microbe-based CO₂ sequestration techniques since the dissolution of silicate minerals for nutrient acquisition also sequesters CO₂. ❧ Chapters 3 & 4: Geomorphic control on the hydrology and carbon budget of weathering. Erosional processes and hydrology are known to influence chemical weathering rates by controlling the timescales over which minerals react. Accurately describing the complex linkages between weathering, erosion, and hydrology observed in natural environments remains a major research challenge. To help address this problem, a major part of my Ph.D. was focused on characterizing how chemical weathering and hydrology are coupled in distinct erosional environments. This work combines hydrologic monitoring, solute chemistry, and water isotope analyses in order to robustly document how water is stored in catchments and link this to measured solute fluxes from chemical weathering (Clark et al., 2014; Torres et al., 2015). The results of this study are intriguing in that the hydrological control of weathering was found to vary predictably with the erosional regime, which has important implications for how changes in tectonic activity affect global weathering fluxes. ❧ By affecting the timescales over which weathering reactions occur, erosional processes also influence which minerals react due to the intrinsic variability in the reaction rates of different minerals. Rapid erosion rates favor the oxidation of sulfide minerals relative to the dissolution of silicate minerals, which leads to the release of CO₂ into the ocean-atmosphere system. To trace sulfide oxidation and its effect on the carbon budget, I combined multiple isotopic systems (e.g., S, C, and Sr) with major and trace element analyses (Torres et al., in prep). By making observations in catchments with diverse erosional regimes, it was possible to interrogate how sulfide oxidation fluxes relate to erosional processes. My results showed that sulfide oxidation dominates in rapidly eroding environments and leads to the significant release of CO₂. This is in contrast to more slowly eroding environments, where CO₂ consumption during silicate weathering dominates. ❧ Chapter 5: The evolution of the Cenozoic carbon cycle. My research on sulfide oxidation in modern systems suggested a link between tectonic uplift and the carbon budget of weathering processes with important implications for the long-term carbon cycle. To test this hypothesis, I incorporated the effects of sulfide-oxidation driven CO₂ release into a model of the Cenozoic carbon cycle. The Cenozoic carbon cycle has long plagued geochemists as isotopic records suggest changes in weathering fluxes that appear to be inconsistent with the requirement of mass balance in the long-term carbon cycle. By incorporating sulfide oxidation as a CO₂ source, I was able to provide a novel solution to the "Cenozoic isotope-weathering paradox" (Torres et al., 2014).
Members of the Marinobacter genus play an important role in hydrocarbon degradation in the ocean - a topic of special significance in light of the recent Deepwater Horizon oil spill of 2010. The Marinobacter group has thus far lacked a genus level phylogenetic probe that would allow in situ identification of representative members. Here, two new 16S rRNA-targeted oligonucleotide probes (Mrb-0625-a and Mrb-0625-b) were developed to enumerate Marinobacter species by fluorescence in situ hybridization (FISH). In silico analysis of this probe set demonstrated 80% coverage of the Marinobacter genus. A competitor probe was developed to block hybridization by Mrb-0625-a to six Halomonas species with which it shared a one base pair mismatch. The probe set was optimized using pure cultures, and then used in an enrichment experiment with a deep sea oil plume water sample collected from the Deepwater Horizon oil spill. Marinobacter cells rapidly increased as a significant fraction of total microbial abundance in all incubations of original contaminated seawater as well as those amended with n-hexadecane, suggesting this group may be among the first microbial responders to oil pollution in the marine environment. The new probe set will provide a reliable tool for quantifying Marinobacter in the marine environment, particularly at contaminated sites where these organisms can play an important role in the biodegradation of oil pollutants. The next sections of this dissertation focus on the hydrothermally active sediments at Guaymas Basin, which show a wide range of shallow subsurface temperatures: from 3°C to 200°C in the first 45 cm depth. A combination of extreme thermal gradients and compressed geochemical and metabolic zones limits the depth range of microbial colonization in Guaymas sediments. Using stable carbon isotopic values for methane and dissolved inorganic carbon compared to associated temperatures the upper thermal limits for the anaerobic oxidation of methane and organic carbon remineralization in Guaymas sediments are suggested to be 80oC and 100oC, respectively. At higher temperatures the isotopic imprints of these microbially mediated processes cannot be detected. Additionally, 16S rRNA gene clone libraries demonstrate differential biogeographical zonation patterns for archaea versus bacteria, with archaeal community structure being more heavily influenced by hydrothermal regimes. Chloroflexi and Deltaproteobacteria dominated the bacterial clone libraries, and anaerobic methane-oxidizing (ANME) archaea represented nearly half of the total archaeal clone library. Thermal zonation of ANME archaeal subgroups is strong: ANME-2c is restricted to low temperature sediments (<25oC), ANME-1 is dominant at warmer temperatures, and the ANME-1 Guaymas archaea appear to have access to the deepest and hottest sediment horizons up to approximately 80oC. In the last chapter of this dissertation, microbial life at extreme temperatures was investigated further by RNA-based methodologies. Using push core samples collected by the Alvin submarine at four high temperature sites with 40-cmsbf thermal maxima ranging from 100°C to 185°C, the composition of the active microbial community and its possible influence on carbon and sulfur cycling was investigated. Here, evidence is presented indicating that hydrothermal fluctuations are frequent enough to restrict hyperthermophilic life to sediments with average in situ temperatures between 70°C and 95°C, where temperatures may vary by 25°C in as little as a day. Strong microbially mediated sulfate reduction is implicated by sharp decreases in porewater sulfate within the upper 15 cm of all four high temperature cores, while stable isotopic evidence of methane oxidation is only expressed in a single core. Archaeal sequence recovery was greater than bacterial sequence recovery in six out of eight samples from the four cores, but bacterial sequence recovery was particularly strong for a single core, yielding 35% of the total archaeal and bacterial recovery from all samples. Although putative anaerobic methane oxidizing (ANME) archaea were very common, distinct cores hosted diverse and distinct sequence assemblages, including ANME-1 Guaymas, ANME-2c, and ANME-2d/GoM Arc-1/Methanoperedenaceae. Dominant bacterial groups fell within the Thermodesulfobacteriaceae family in the Thermodesulfobacteria phylum, the Helicobacteriaceae family in the subphylum Epsilonproteobacteria, or were close relatives of Desulfocapsa exigens in the subphylum Deltaproteobacteria. The most probable thermo- or hyperthermophilic groups were investigated by co-occurrence of OTUs across the four hottest samples within the sediment cores and appear to be ANME-1 Guaymas and an uncultured representative of the Miscellaneous Crenarchaeotal Group (MCG)-15 for archaea, and members of the Thermodesulfobacteriaceae family for bacteria.
The deep subseafloor biosphere represents a frontier for the discovery of new microbial life and for investigations of the extent, versatility, and perseverance of life on earth. However, there are many challenges in studying this community of microorganisms, and the past 20 years of study have only begun to shed light on this vast and complex ecosystem. With each chapter herein I have taken on some of those challenges and have made progress in overcoming them, while also contributing to the knowledge of an environment that—despite its potential significance—remains relatively unexplored. In particular I have focused on the application of molecular methods to the study of the subseafloor biosphere, which is complicated by difficulties such as low biomass, extracellular and fossil DNA, potential for drilling-induced contamination, and method biases. In chapter 2, I examined the potential sources of molecular signals suggestive of phototrophic organisms in the subseafloor via cultivation, DNA sequencing, and PCR-based inquiries. Although I found that most likely the molecular signals of phototrophic organisms found in the deep biosphere do not represent viable cells, factors such as the uncertainty of DNA survival time and the paucity of information on many subseafloor taxonomic lineages made it difficult to furnish an explanation for these molecular signals. Additionally, while I was unable to succeed in demonstrating phototrophy in cultivations, my results suggested that I was able to stimulate other microbial growth, and that in most cases the organisms that became dominant in the cultivations had been only a minor proportion of the original uncultivated sediment. This is a good demonstration of the potential importance of even the “minor” components of a microbial community under changing environmental conditions. In chapter 3, I took on the challenges of carrying out molecular work on very low biomass sediment samples by developing and testing a novel method of whole genome amplification that overcame some of the limitations of previous methods for subseafloor samples. While the method solved some problems specific to low biomass samples and seemed a viable alternative to previous methods of whole genome amplification for these samples, my work reaffirmed previous studies in showing that there are still dangers in interpreting community data based on DNA that has been subjected to whole genome amplification with any method. Further, I identified problems with comparing data from different sequencing technologies and with different data analysis and classification methods. In chapter 4, I carried out a follow-up to the study of whole genome amplification utilizing samples from 2 previously uncharacterized subseafloor locations in the Eastern Equatorial Pacific. While the limitations of utilizing amplified DNA were again reinforced, my results showed that at some levels community analysis on amplified DNA was relatively accurate. Here I was able to show a robust taxonomic distinction between these 2 new sites from the pelagic abyss and metagenomes from 3 previously available coastal margin subseafloor locations, while also demonstrating the potential validity of predicting microbial community composition in a subseafloor location based on results from a nearby and very similar subseafloor location. In the final chapter I took on the problem of drilling-induced contamination by carrying out the first study of its kind to extensively characterize the microbial community from both the sediments and the corresponding drilling fluid used during sample acquisition, on a range of sample depths including several samples taken with the more aggressive extended core barrel coring (XCB) method. I found the drilling fluid to have a very minor influence in the molecular analysis of all samples except for one, which lends confidence to the study of deep cores while at the same time reinforcing the importance of making drilling fluid controls a standard part of every molecular study of subseafloor sediment samples. Additionally, I characterized the previously unexplored microbial communities of the Costa Rica Margin subseafloor and discussed some potential linkages between subseafloor microbial taxa and pore-water geochemistry variables. Considering the current state of knowledge in this environment due to its challenging nature, the work herein contributes greatly to our understanding of microbial biogeography and relationships with environmental conditions, as well as to the many complexities in performing and interpreting molecular analyses in the subseafloor. It also provides a wealth of new 16S rRNA and metagenomic datasets that can continue to be used for further investigations.
The first detection of an SF6 patch south of the injection source apparently contradicts the prior hypothesis that crustal fluid is flowing northward from Grizzly Bare to Baby and Mama Bares, but there are extensive leaks in Hole 1301A, which may have inadvertently pulled SF6 southwards. CORKs 1301B and 1026B are also leaking hydrothermal fluid, and any data collected from these holes should be interpreted carefully. Both 1362A and injection Hole 1362B are operating as intended with no known leaks, although a discharge valve was opened at 1362B in 2011 and closed in 2013. Following this perturbation to the flow field, the 1362A valve was opened in 2013 and then closed in 2014. Tracer was detected in both 1362A and 1362B from the beginning of sampling in 2013, one year after injection. The mean tracer concentration for Holes 1362B and 1362A was 4.6 nM and 4.5 nM, respectively. The sustained signal at these CORKs suggests a hovering of an SF6 patch spanning at least 311 meters in the region. The detection of a considerable amount of tracer to the north at 1362A is consistent with the hypothesized SW-NE fluid flow direction. The hydrogeological fabric at our site appears to be heterogeneous, with fluid transport occurring through small, isolated permeable zones found in the upper volcanic portion of the ocean crust., A tracer injection experiment was performed in 3.5 Myr old seafloor comprised of sediment-buried abyssal hills oriented N20°E, and located 100 km east of the Endeavor Segment of the Juan de Fuca spreading ridge in the northeastern Pacific. In the summer of 2010, a mixture of tracers (metal salts, dissolved sulfur hexafluoride [SF6], microspheres) was injected into the crust via borehole 1362B as part of a 24-hour injection experiment during IODP Expedition 327. Fluid samples were subsequently collected from 1362B, and from four additional holes (1026B, 1362A, 1301A, and 1301B) located 300 to 550 m away from the injection hole. The borehole array penetrates a hydrothermal fluid flow system thought to be flowing from SW to NE along a buried abyssal hill often referred to as Second Ridge (SR). Hydrothermal flow is thought to be controlled by a series of exposed volcanic outcrops located along the same buried hill. According to the hypothesis, recharge of bottom seawater occurs through Grizzly Bare, an exposed outcrop 52 km south of the borehole array, and discharge seeps through Baby Bare and possibly also Mama Bare outcrops, which are both located within 5 km to the existing boreholes. The goal of this study is to test hypothesized fluid flow direction, flow velocity and crustal permeability using the conservative gas tracer SF6., and Two small cylinders of SF6 were injected at a fluid pumping rate of 6.7 L/s for 20.2 hours, resulting in a mean concentration of 47.6 microM or 47,600 nM (total of ~23 mol of SF6 was injected). Borehole fluid was continuously sampled in 1.8 mm ID copper tubes using osmosamplers (OS) from each of the long-term, subseafloor observatories (known as CORKs) that had previously been installed in the boreholes. The OS spools were recovered using the ROV Jason in 2011 and 2013. Following recovery, fluid samples were transferred into evacuated vials for measuring SF6 via gas chromatography in a shore-based laboratory. Results of samplers recovered in 2011 indicate the first arrival of injected SF6 ~305 days after injection at Hole 1301A, located 550 m south of the injection Hole 1362B. This suggests that the mean lateral transport of tracer is ~1.8 m/day (660 m/yr), a rate at the upper end of previous upper crustal fluid flow velocity estimates.
Porewater inorganic carbon concentration and total alkalinity from deeply buried marine sediment reflect biological activity, mineral diagenesis, sedimentary processes and past bottom ocean water composition. Reliable interpretation of these data is often complicated and/or limited due to (i) major physical environment changes taking place during sediment core retrieval, and (ii) the resulting precipitation of calcium carbonate (CaCO3) in the course of sample collection, processing and storage. Here we describe a robust method for quantifying the in-situ porewater carbonate system chemistry in deepsea sediment cores. The method relies on the over-determination of the dissolved carbonate system by measuring three of its parameters, and explicitly assumes CaCO3 saturation in the sediment and equilibrium conditions in-situ. The principles of the method are presented.
We experimentally test the proposed approach using concentration profiles of dissolved carbonate system components collected from the Integrated Ocean Drilling Program (IODP) Site U1368 in the Southern Pacific. Our results show that this method can be used to accurately reproduce the in-situ aqueous carbonate system chemistry if dissolved inorganic carbon, total alkalinity and calcium concentration are measured simultaneously. The method is well suited for use over a broad range of porewater chemistry and applicable for sediment over ca. 50% of the seafloor.