The completion of Integrated Ocean Drilling Program (IODP) Exp 327 in Summer 2010 launched a series of multi-year, single-hole and cross-hole experiments in young oceanic crust (eastern flank of the Juan de Fuca Ridge (JFR)), to assess hydrogeologic, solute transport, and microbiological processes in multiple directions, depths and at numerous spatial and temporal scales (meters to kilometers, minutes to years)  The infrastructure to complete this work includes six boreholes (Holes 1026B, 1027C, 1301A, 1301B, 1362A, and 1362B) that are cased though the sediment section and open to the basaltic crust below. These boreholes are instrumented with sensors, loggers, and samplers located at depth in the crust and attached to wellheads at the seafloor. These systems are being used to monitor ambient subseafloor conditions in response to perturbations and quantify complex system responses within the crust [2, 2a]. Collectively, these experiments are elucidating fundamental questions related to linked properties and processes within the crust: (1) What is the nature (magnitude, distribution) of permeability in upper oceanic crust? (2) What are the magnitudes and directions of driving forces, fluid fluxes, and associated solute and heat transport? (3) What is the nature of fluid storage properties within the crust, including their pressure-, temporal- and spatial-dependence? (4) What are relationships between fluid flow, vertical and horizontal compartmentalization, distribution of microorganisms, alteration, structure, and primary crustal lithology? (5) How large are crustal fluid reservoirs, what are fluid and colloid flow velocities, and how do these respond to perturbations (pumping, tides, seismic events)?(6) What is spatial and temporal variability in phylogenetic and functional diversity of resident microbial communities, and what are the nature and magnitude of dominant metabolic processes?
We are collecting samples and data using crustal borehole observatories ("CORKs") to address these questions, taking two primary approaches. First, we have placed sensors, samplers, and experiments at the seafloor, using wellhead instrumentation to monitor conditions and collect fluids from depth. Earlier studies have focused on installing and servicing these seafloor systems to recover samples and data, and to collecting discrete samples of crustal fluids and microbial materials using tubing that extends from the seafloor to depth. Second, we have placed autonomous and self-contained instruments deep below the seafloor, within the volcanic ocean crust, in five of the borehole observatories (Holes 1026B, 1301A/B, 1362A/B). These instruments include temperature loggers, fluid samplers, and microbial incubation habitats and enrichment studies, which will have been operating for 4 to 5 years in 2014, collecting samples and data that record the response to drilling operations, observatory installation, subsequent perturbation and response to tracer injection. As described later in this proposal, a small number of seafloor samples collected in Summer 2011 show that a multi-tracer transport experiment, initiated during IODP Expedition 327 (with injection of metal salts, inert gas tracer, and fluorescent particles, as part of a 24-hour pumping test), has resulted in direct evidence for tracer transport hundreds of meters between basement boreholes. The seafloor samples are limited in temporal and spatial resolution, in contrast to the downhole systems deployed prior to tracer introduction to the crust, which will provide greater spatial coverage and a continuous (multi-year) record. This proposal seeks support to recover samples from deep within instrumented boreholes, completing the first cross-hole hydrogeologic and tracer experiments attempted in the ocean crust.
We request support for a 12-day on site ROV/submersible expedition to recover downhole instruments installed in subseafloor borehole observatories in 2009 and 2010. In addition, we propose to recover an autonomous flowmeter that is recording long-term discharge from one of the basement boreholes (as part of the cross-hole perturbation and sampling experiment), complete a final round of fluid and microbial sampling using seafloor valves and fittings to gather large-volume samples from depth, retrieve remaining wellhead samplers, and close valves to seal the CORKs so that they are available for future studies. After collection of samples and data, we will conduct an extensive analytical program to quantify concentrations and transport behavior of co-injected tracers (SF6, salts of Cs, Er, and Ho, two sizes of fluorescent microspheres and DAPI-stained cells, and chlorinity (for freshwater inputs); [1,2]), characterize microbial materials, and interpret the cross-hole and multi-depth pressure, thermal and chemical response to long-term discharge of fluids from two boreholes (1362A/B).
This is a multi-investigator and multi-university proposal, with six co-PIs: Wheat (UAF), Fisher (UCSC), Cowen (UH), Clark (UCSB), Edwards (USC), and Becker (Miami). Four of the co-PIs (Wheat, Fisher, Cowen, and Edwards) also are co-PIs of the Center for Dark Energy Biosphere Investigations (C-DEBI), a Science Technology Center funded separately by NSF. These four co-PIs will draw upon existing C-DEBI support for partial support for students, postdocs, technicians, supplies, travel, and other expenses, significantly reducing budgets associated with the current proposal. In addition, although we are requesting UNOLS support for ship and ROV time as part of this proposal, we have submitted a complementary proposal to the Schmidt Ocean Institute (SOI), which would provide ship (R/V Falkor) and ROV (ROPOS) support should that proposal be approved. The SOI does NOT provide funds to conduct research, only ship and ROV time. This SOI proposal is currently in review (outcome anticipated in late August or early September), and if it is funded, we will not need NSF-funded ship or ROV support through UNOLS. The technical and scientific program we propose will be identical whether it is completed with SOI or UNOLS assets. The main difference would be the number of berths available, but we are planning for a lean shipboard party in any case, and a focused scientific plan comprising essential shipboard and land-based activities so that this suite of experiments can be completed.
II. SCIENTIFIC MOTIVATION
Most of the ocean floor is hydrogeologically active [e.g., 3, 4, 5] and the volcanic oceanic crust comprises a global-scale aquifer, containing as much fluid as that stored in ice caps and glaciers . Hydrothermal fluid flows through the oceanic crust are equal to or greater than those from rivers to the ocean, effectively recycling the ocean every 105 to 106 yr [7-9]. Global hydrothermal flows influence: alteration of the lithosphere and the composition of flowing fluids and the ocean; subseafloor microbial ecosystems; and diagenetic, seismic, and magmatic activity along plate-boundary faults [e.g., 10-14]. Scientific ocean drilling has long sought to quantify the dynamics and impacts of fluid flow through the oceanic crust . Although there have been notable successes in drilling seafloor spreading centers, particularly ore deposits and in sedimented environments, much work has focused on "ridge flanks" areas far from the magmatic influence of seafloor spreading, where overlying sediments stabilize the drillstring, and cooler basement conditions make sampling and experiments more tractable.
Hydrogeologic testing of the ocean crust , has traditionally involved very short (20-30 min), single-hole experiments [17-19]. The recent development of pressure-tight, subseafloor observatories (CORKs) comprises a major advance in marine hydrogeologic studies [20, 21]. These systems allow thermal, pressure, chemical and microbial disturbances associated with drilling to dissipate, allowing monitoring, sampling and in situ experimentation. CORK systems have been used mainly for passive monitoring and sampling in single holes. Initial tracer tests with these systems have been limited to single holes and limited in scope [Gieskes et al., unpublished work in 504B, 22, 23]. Our project uses a three dimensional network of CORKed boreholes to run the first controlled, cross-hole tests, the first in-situ microbial enrichment experiments, and the most sophisticated microbial colonization experiments attempted in volcanic crust. These tests and experiments are possible only now, with the recent development of novel seafloor sampling systems (GeoMICROBE Sled; ), an autonomous flow meter , downhole osmotic systems , in-situ colonization and enrichment experimental systems , and high-resolution pressure recorders .With such instrumentation and novel wellhead innovations, we are able to conduct the first hydrogeologic studies of the crust to assess vertical compartmentalization, and to quantify azimuthal (directional) fluid flow properties in oceanic crust. These experiments also are designed to provide the most pristine and most continuous multi-year record of ridge-flank crustal (borehole) fluids anywhere on the planet.
The eastern flank of the JFR has become a de-facto "type setting" for ridge flank hydrothermal processes, in part because there is a long history of multidisciplinary research in this area. There are other locations where complementary studies are underway, most notably the North Pond field site, on young crust (8 Ma) west of the Mid-Atlantic Ridge, where conditions are cooler  but crustal fluid flow paths are less well defined. The eastern flank of the JFR has the most extensive network of observatories and instrumentation, arranged with a geometry that allows both single and cross-hole experimentation, including evaluation of lateral and vertical variations in scalar and vector (flow, transport) properties. There is nowhere else on the planet where these kinds of experiments can be completed at present, not without considerably greater investments in drilling time, infrastructure, and scientific instrumentation. Considerable work and resources have been focused on the JFR eastern flank in the last decade, and success has not come easily. But having made the long-term commitment and taken significant technical risks, we are poised to reap considerable scientific rewards with this final phase of operations.
Proposed work will provide samples and data that are unique; are essential for understanding ocean crustal hydrogeology, biogeochemistry and microbiology; and cannot be obtained in any other way. Earlier short-term hydrologic tests in ocean crustal boreholes [18. 19, 29], and tests from open holes resulting from natural or induced differential pressures [30,31], have provided no information on formation compressibility, which is essential for understanding crustal response to transient pressure events. The use of natural formation overpressure to run long-term free flow experiments, as we are doing, quantifies formation transmissive and storage properties across a continuum of crustal scales (meters to tens of kilometers). The bulk formation properties indicated by earlier cross-hole responses in this area suggest permeability that is lower than determined with single hole tests , and several orders of magnitude less than estimated by numerical modeling and analyses of formation responses to tidal and tectonic perturbations. This discrepancy (which is contrary to expected scaling of permeability with the lateral scale of testing) may be reconciled if the upper crust in this area is azimuthally anisotropic with respect to basement permeability, with higher permeability in the "along-strike" direction (trending N20E). Testing this hypothesis is a fundamental goal of the complete experimental program, along with evaluating whether distinct physical and chemical zones in the crust are hydrogeologically connected or share common microbial ecosystems. Applying a suite of techniques using a single network of boreholes is the only way to accurately determine the nature of crustal permeability and storage properties, resolve scaling influences, and test the validity of idealized crustal representations commonly used in models [e.g., 32-37]. The JFR is an ideal place to address these issues, because individual tests can be run for years, delineating the scale-dependence of hydrologic properties using one experimental method.
Similarly, there is no other way to assess actual fluid, solute, and particle flow directions and rates except through a tracer experiment. This kind of experiment has never before been attempted in the ocean crust. The extent of water mixing and water-rock interaction within an aquifer depends on effective porosity (fraction of open space involved in fluid flow) and hydrodynamic dispersivity (spreading of solutes by mechanical dispersion and diffusion) [38-43]. Understanding these properties is critical to successful reactive-transport modeling and interpreting the age distributions of subseafloor fluids. Simply observing tracer arrival across meters to kilometers is a significant accomplishment, but the most valuable records of tracer transport during these experiments remain deep below the seafloor. Samplers and loggers currently deployed at depth in existing CORKs contain long-term records of temperature, geochemical conditions, and microbial populations, from before, during, and after the initiation of cross-hole experiments. Recovery of this instrumentation will provide critical information contributing to the success of the multidisciplinary experiments.
III. FIELD SITE AND EARLIER RESULTS
Eastern Flank of the Juan de Fuca Ridge (JFR). Numerous studies summarize geology, geophysics, basement-fluid composition, hydrogeology, and microbiology within young seafloor on the eastern flank of the Endeavour segment of the JFR [e.g., 11, 23, 24-32]. Topographic relief associated with the JFR axis and abyssal hill bathymetry on the ridge flank has helped to trap turbidites flowing from the continental margin to the east (Fig. 1) . This has resulted in burial of young oceanic basement rocks under thick (200 -700 m) sediments. Sediment cover is regionally thicker and more continuous to the east, but there are seamounts and smaller basement outcrops located up to 100 km east of the spreading center on 3.5-3.6 Ma crust. Regional basement relief is dominated by linear ridges and troughs oriented parallel to the spreading center and produced mainly by faulting, variations in ridge magmatism, and off-axis volcanism [78, 79]. Low-permeability sediment limits advective heat loss regionally, and leads to strong thermal and chemical differences between bottom seawater and basaltic formation fluids [e.g., 22].
ODP Exp. 168 (1996) and IODP Exp. 301 (2004).Ocean Drilling Program (ODP) Leg 168 drilled a transect of eight sites on 0.9 to 3.6 Ma crust east of the JFR . These operations included installation of four CORKs that extended into uppermost basaltic crust, two of which are located on 3.5–3.6 Ma seafloor near the eastern end of the drilling transect (Holes 1026B and 1027C, Fig. 1) [51, 52]. Prior to ODP Leg 168, there was a largely two-dimensional view of the dominant fluid circulation pathways across the eastern flank of the JFR, with recharge occurring across large areas of basement exposure close to the ridge (near the western end of the Leg 168 transect), then flowing towards the east. However, results from ODP Leg 168 were inconsistent with this view and consistent with flow dominantly parallel to the spreading system to the west with seamounts to the south and north providing conduits for seawater inflow and egress [53, 54]. Numerical models of outcrop-to-outcrop hydrothermal circulation between recharging and discharging seamounts show that rapid flow is sustained (as a "hydrothermal siphon") and temperatures matched to observations if basement permeability along the flow path is about 10–11 m2 .
Figure 1. Maps of the field sites. A. Regional map with primary field area (yellow box), located between the continental margin and JFR. B. Detail contour chart of individual CORK sites (circles) and nearby basaltic outcrops (yellow contours). Labeled depth contours are in meters. Most CORK sites are aligned along the direction of primary structural strike in basement, N20E, with Site 1027C located to the east (~70° oblique to strike).
Table 1. Primary work sites for proposed Summer 2014 Expedition. Locations are shown in Fig. 1.
Leg 168/Exp. 301
IODP Exp. 301 returned to the eastern end of the Leg 168 transect and drilled deeper into basement; sampled sediment, basalt, and microbiological materials; replaced the borehole observatory in Hole 1026B; and established two additional CORK observatories at Site 1301 for use in long-term, multi-dimensional hydrogeologic experiments . Results based on Expedition 301 and site surveys demonstrate that: (A) perturbations in crustal boreholes in this setting are transmitted across distances of at least 2.4 km, and natural circulation paths extend tens of kilometers, ideally suited for hydrologic tests with the geometery of the boreholes and observatories; (B) downhole temperature and osmotic sampling systems provide a quantitative measure of chemical and thermal changes in borehole conditions; (C) colonization substrates provide a useful means for characterizing microbial community composition; and (D) computer simulations coupled with a diversity of local and regional and data provide key constraints to hydrologic conditions.
(A) Pressure data recovered in 2005 indicated that CORKs at Site 1301 were not sealed, because casing seals could not be installed during Exp. 301, and cold seawater was flowing down both holes and into basement. A large-scale assessment of basement hydrogeologic properties was made from the long-term pressure perturbation observed in the Hole 1027B CORK, resulting from leakage of cold bottom seawater into the crust at Site 1301 during the initial 13 months post-drilling . Basement operations in Hole 1301B caused the greatest pressure response in Hole 1027C, 2.4 km away (Figs. 1 and 2) whereas operations in shallower Holes 1301A and 1026B had little or no influence, indicating that the shallowest crustal intervals are not the best connected laterally in this area. The dynamic cross-hole pressure response was modeled analytically, indicating a bulk permeability within the upper 300 m of crust of k = 0.7 to 2 x 10-12 m2, and aquifer compressibility of α = 3 to 9 x 10-10 Pa-1. This crustal permeability is at the lower end of estimates based on single-hole experiments , even through cross-hole tests investigate a much larger rock volume, extending ~10–30 km from the borehole. This difference can be reconciled by azimuthal anisotropy in basement permeability, with the direction between Sites 1301 and 1027 being 50° oblique to structural strike of basement (N20E) (Fig. 2B). The flow experiments currently underway, and to be completed as part of the current proposal, will assess permeability along strike and up to 65° oblique to strike, allowing the first direct test of permeability anisotropy in the ocean crust.
Figure 2. Results from an inadvertent cross-hole experiment between Holes 1301B and 1027C. A. Pressure response in Hole 1027C CORK to long-term flow into Hole 1301B, 2.4 km away. Pressure record from Hole 1027C (blue line), corrected for tidal loading and other instantaneous response, is fit with an analytical model of crustal fluid flow, indicating basement permeability that is lower than indicated by packer tests or larger-scale methods. B. This pressure response could be explained by permeability anisotropy in basement, based on calculations shown. Calculated effective (apparent) permeability in an anisotropic medium depending on the angle of testing relative to the primary flow direction. Because the direction from Hole 1027C to 1301B is oriented 50° from strike, permeability is dominated by the "low permeability" direction. Tests to be completed as part of the current proposal will test and even more oblique direction, simultaneously with along-strike testing, resolving anisotropy.
(B) During CORK servicing in Summer 2008, we learned that Hole 1301A had "turned around" and was discharging shimmering (~64°C) crustal fluids. Recovery of downhole OsmoSamplers and temperature loggers revealed that the abrupt change in borehole conditions occurred over just a few days in September 2007, following a long period of irregular downflow (Fig. 3). The composition of the fluid collected from Hole 1301A after Fall 2007 is similar in some ways to fluids from Baby Bare springs (to the south) and fluids that vented from Hole 1026B (to the north) . The rapid change in fluid composition when borehole flow reversed illustrates both a lack of mixing of inflowing and (subsequently) outflowing fluids, and the extent of subseafloor inorganic and microbial reactions. The evolution of these fluids is analogous, in several respects, to the larger scale circulation pathway from Grizzly Bare to Baby Bare, between recharge and discharge sites [53,54, 57]. Furthermore, the record of Cs (introduced during sampling) documents exchange between the borehole and formation, illustrating the potential for hydrologic characterization using introduced tracers (albeit in a single hole).
Figure 3. Four-year record of temperature and geochemistry of fluids from Hole 1301A illustrates hydrologic, geochemical, and microbial dynamics . A. Mg (red and blue) and temperature (orange) data, relative to Mg data concentrations in seawater (black) and in formation fluids (green), from downhole sensors and OsmoSamplers (OS) in Hole 1301A. These data clearly illustrate the timing of the event and the path towards the event. B. Flow reversal is apparent from thermal record (purple), and is quantified by single-hole tracer experiment using Cs (blue data, idealized with dashed orange line). Rapid downflow slowed after ~1 year, then abruptly reversed during one week in 9/07. Rapid upflow of formation fluid shifted tracer baseline to match natural conditions (red triangles), as in Hole 1026B (dotted green line).
(C) Subseafloor microbialsubstrate and enrichment experiments were recovered from the Hole 1301A CORK during servicing in Summer 2008, revealing a direct record of microbial adaptation to the changing hydrologic and biogeochemical conditions  (Fig. 4). Organisms that colonized the growth media initially were adapted to cold, oxidative conditions, but were subsequently replaced by microbes that are more representative of natural crustal conditions (warm, reducing), dominated by the Firmicutes phylum. Some sequences grouped near Ammonifex genera, thermophilic, chemolithoautotrophs capable of growth with hydrogen, formate, or pyruvate coupled to nitrate, sulfate, or S° reduction. The highest cell counts within flow-through experiments from Hole 1301A were found on samples of olivine, and culturable organisms from these samples were capable of nitrate reduction and iron oxidation . In addition, repeat sampling from Hole 1301A has revealed significant differences in microbial community structure, with changes in both bacterial and archaeal lineages and absolute abundances (44).
Figure 4. Microbiological growth substrate deployed and recovered after four years from depth in Holes 1026B and 1301A . A. Polished chips of basalt (bas), peridotite (per), pyrite (pyr) and biotite (bio) as deployed in CORKs on Exp. 301. Top image is pristine substrate prior to deployment, center image is substrate recovered from Hole 1301A, bottom is substrate recovered from Hole 1026B. B. Cells observed on rock chip surfaces from Hole 1301A, as imaged by epifluorescent confocal microscopy, showing chains of bulbous cocci.
(D) Analytical and numerical models simulated outcrop-to-outcrop hydrothermal circulation between Grizzly Bare and Baby Bare outcrops, separated by 52 km along-strike, to estimate the nature of basement properties required to allow inferred patterns and rates of fluid circulation [54, 57]. Outcrop-to-outcrop hydrothermal circulation can be sustained when basement permeability is ≥10-12 m2. At lower permeabilities, too much energy is lost during lateral fluid transport for circulation to continue without forcing, given the limited driving pressure difference at the base of recharging and discharging fluid columns. In addition, fluid temperatures in upper basement are highly sensitive to modeled permeability. When basement permeability is too high (10-10 to 10-9 m2), fluid circulation is so rapid that basement is chilled to temperatures below those seen regionally (modeled values of 20–50°C). A good match is achieved to observed upper basement temperatures of 60–65°C when lateral basement permeability is 10-11 m2, overlapping with values determined with single-hole packer experiments .
IODP Exp. 327 (2010).Two new subseafloor borehole observatory systems were installed in Holes U1362A and U1362B during IODP Exp. 327, a CORK instrument string previously deployed in Hole U1301B was recovered and replaced, and a long-term tracer experiment was initiated. Injected tracers included SF6, salts of Cs, Er, Ho, and several sizes of fluorescent microspheres and fluorescent-stained microbes filtered from surface seawater (particles). Tracers were injected as part of a 24-hour pumping experiment intended to test a large volume of basement rock around Hole 1362B, using Hole 1362B for perturbation and other CORKs for monitoring formation response. Instruments deployed at depth in nearby CORKs were designed and deployed specifically to sample tracers in the crust over time.
During Atlantis/Jason Expedition AT18-07 in 2011, we: (1) recovered wellhead OsmoSampling systems deployed on pre-Exp. 327 CORKs, and deployed new OsmoSampling systems on the new CORKs (for geochemical sampling and microbial growth experiments); (2) downloaded long-term pressure data; (3) retrofited the old CORK in Hole 1027C for pressure monitoring with the newest generation of data loggers; (4) recovered and deployed a long-term GeoMICROBE sled used for autonomous, large-volume fluid sampling and electrochemistry ; (5) collected large volumes of fluids from depth using a variety of pumping, sampling and filtering systems; and (6) deployed an autonomous, electromagnetic flowmeter on one of the Site U1362 CORKs, then opened a large-diameter ball valve to initiate a long-term, cross-hole flow experiment [2 and 2a].