EXECUTIVE SUMMARY

The Center for Subsurface Geomicrobiology at NUSL (NUSL-OSG) will be the only facility in the world where long-term, in situ geomicrobiology and biogeochemistry experiments explore the evolution, adaptation, and limits of microbial life, as we understand it, in the deep subsurface. The NUSL-OSG will provide a here-to-fore previously unattainable resource for multi-disciplinary and multi-institutional investigations for the international earth and biological scientific communities. The primary goal of NUSL-OSG is to provide an experimental and intellectual foundation for investigating the origin and bounds of life as well as to develop practical applications for the bioremediation, biotechnology and pharmaceutical industry. Integral to the scientific goals of the NUSL-OSG will be a very active program to foster education and training for future generations of scientists and teachers from K-12 to visiting researchers, focusing on those groups that have remained underrepresented throughout the 20th century. To accomplish its goals the activities of the NSUL-OSG must be closely linked and integrated with those of the physics community at NUSL and with other regional academic centers in order to take full advantage of shared technological infrastructure, intellectual prowess and educational outreach capabilities.

PART 1-RESEARCH THRUSTS AND EDUCATIONAL OPPORTUNITIES

The relatively recent discovery of deep subsurface microbial communities and what appears to be a subsurface biosphere has opened a new scientific frontier where earth sciences, chemistry, physics and biology merge to provide insights into how life on this planet and even extraterrestrial life, may have originated and evolved over billions of years (Fredrickson and Onstott, 1997). The geological isolation of these deep subsurface microbial communities offers the potential to answer questions on the origin of life and its diversity as well as constraining the possibilities for life beneath the surface of Mars and other planetary bodies. In addition to the role of microorganisms in shaping the life forms on earth, the importance of microorganisms in the dissolution and formation of minerals is only now becoming recognized as geomicrobiology comes to the fore. Advances in our understanding of the origins, diversity, distribution and function of microorganisms in deep, often extreme, subsurface environments will rapidly expand our knowledge of geomicrobiological and biogeochemical processes on Earth and beyond. The discovery of novel microorganisms from deep accessible subsurface habitats provides opportunities for discovering new pharmaceuticals, processes for biochemical and chiral-specific synthesis, environmental remediation and energy production. Finally, a fundamental knowledge of subsurface biogeochemical processes and elemental cycling is critical for predicting the impacts of subsurface contamination and underground waste isolation and for development of subsurface remediation strategies, including the storage of radioactive waste and CO2 sequestration. Indeed, this knowledge will be enabling for good environmental stewardship practices that will serve our posterity and us.

A major obstacle to understanding the subsurface biosphere has been our limited ability to access the deep terrestrial environment, acquire uncompromised samples and to place our knowledge of isolated microorganisms into the context of the geochemical and hydrogeological processes that control their existence, function and transport. In addition, opportunities to address biogeochemical and microbial transport in compromised but well-characterized deep fractured rock aquifers have been limited and have been restricted to geographically dispersed locations, markedly constraining the collaboration between disciplines critical to understanding complex interrelated phenomena.

The NUSL-OSG will provide a unique opportunity to overcome accessibility and intellectual obstacles by offering 1) three dimensional access to a large-scale subsurface environment that has been highly characterized for over 125 years; 2) a multidisciplinary, collaborative environment that ensures cutting-edge measurements essential to design and interpretation of microbial studies (e.g., high sensitivity isotopic tracer detection, laser and infrared imaging of chemical and physical properties); and 3) an international, multi-institutional research and development and educational environment. These attributes, along with the ability to conduct long-term experiments, the ability to replicate those experiments in similar subsurface environments and the ready access to on site instrumentation for real-time detailed biological and chemical interrogations would establish NUSL-OSG as the world leader in subsurface geomicrobiology research. A recent report on geobiology by Nealson and Ghiorse (2001) emphasized the need for establishing field laboratories for geomicrobiological research that are available for long-term studies; a need that NUSL-OSG can fulfill.

RESEARCH THRUSTS

The proposed research thrusts of the NUSL-OSG will focus on three major areas (and potential applications thereof); a) subsurface biological diversity, b) subsurface biogeochemical processes and c) subsurface microbial ecology.

A. Subsurface biological diversity
Subsurface biological diversity (the new Homestake gold) focuses on the discovery of new microorganisms, novel biological processes and microbial products with potential applications in pharmaceutical (e.g. antimicrobial agents), feedstock chemicals (e.g. chiral synthesis), bioremediation of industrial waste, industrial processing, nanotechnology, opportunities for comparative genomics/proteomics that will provide new insights into the mechanisms of prokaryotic and eukaryotic development and new technologies designed to detect and quantify these processes. Some of the hypotheses to be tested include those below.

A1. "The unique environments of the deep subsurface create microorganisms with unique antibiotic properties and other metabolic products that are important pharmaceuticals. (M. DeFlaun, Envirogen, Inc.; T. Hazen, Lawrence Berkeley National Laboratory)"

A natural adaptation to living in environments that are increasing recalcitrant is the incorporation and horizontal transfer of plasmids that code for enzymes capable of degrading more complex compounds, e.g. humics, that are normally recalcitrant. These plasmids produce the same enzymes that are responsible for many antibiotic activities. While an increasing plasmid load puts an obvious strain on bacteria in more dynamic near surface environments, it has the advantage of providing more carbon and energy sources in the deep subsurface. Indeed, we have also observed and increasing ability to use a broader range of carbon sources as we move deeper into the subsurface (Fredrickson et al; 1988; Jimenez, 1989). These phenomena were observed in 'aseptic' drilling studies down to 1700' done by the DOE Subsurface Science Program in South Carolina. More recent bioprospecting for drug discovery has turned its sights to the unique ecosystems populated by extremophiles (Adams et al., 1995). The diversity of habitats and uniqueness of these ecosystems yields microorganisms that can function under conditions previously thought uninhabitable. Several companies have based their business on bioprospecting (Diversa, Teragen, Chromosome), and consider these extreme environments to be fertile hunting grounds, because the microbes that have evolved in these environments would have developed unique or yet to be discovered metabolic properties.

A2. "Microbial communities that inhabit the mining-impacted environments of the subsurface have properties that would make them uniquely suited to act as agents for ore processing or contaminant remediation. (M. DeFlaun, Envirogen, Inc.)"

Reduced, sulfidic ores when exposed to oxygenated, wet conditions produce a leachate that is typically acidic and contains high concentrations of toxic metals. Aerobic microorganisms derive energy from this environment, become adapted to its lethal conditions and even promote its development. In the presence of organic matter and suboxic conditions, the same leachate provides a source of energy for anaerobic microorganisms to flourish. Within old stopes at Homestake mine where these two processes are occurring, aerobic microorganisms may exist that can be adapted to large-scale industrial processing of metallic ores. Just such a strategy has proven quite profitable for Biox of South Africa. Metal resistant anaerobic microbial communities could also be present that can be utilized to remediate toxic metal contaminated groundwater or mine wastes.

A3. "High temperature, highly saline groundwater harbors microorganisms with unique metabolic properties that are useful in biotechnological applications (M. DeFlaun, Envirogen, Inc.)"

Microorganisms that are capable of surviving at high temperatures in highly saline aqueous solution and extracting energy from recalcitrant organic or inorganic compounds have the ability to maintain their enzymatic proteins under energy deficient conditions. This environment may promote the selection of microorganisms with proteins that are temperature and salinity tolerant. Such proteins have enormous potential for industrial processes.

A4. "The physiological and genetic diversity of subsurface microbial community members will be a function of habitat characteristics. The more diverse the available energy resources and electron acceptors in a given habitat, the higher will be the level of biological diversity (J.K. Fredrickson, Pacific Northwest National Laboratory)."

A4.1. "The highest degree of community complexity and microbial diversity will occur in the mine-altered, subsurface environments. The influx of air as a result of ventilation will promote development of complex microbial communities based on lithoautotrophic metabolism involving energy generation and growth based on oxidation of reduced metals and S associated with local rock and ground waters coupled to O2 respiration."

A4.2. "The diversity of microorganisms associated with groundwater will be low and dominated by slow-growing oligotrophic chemoheterotrophic prokaryotes that can utilize very low concentrations of dissolved organic compounds associated with surface-derived waters. The diversity of prokaryotes associated with older, potentially geothermal water, will also be low but metabolisms will be distinct and will include organisms that can utilize dissolved gases such as H2, CH4, or CO as energy sources."

A5. "The overall genetic (and phylogenetic) diversity of indigenous microbes associated with ancient (tens to thousands of Kyrs.) ground water will be similar to that found in surface environments where the primary energy source is sunlight. The genes and pathways for energy metabolism and biosynthesis will also be similar. A core set of genes should also be present, however, that will allow for growth and survival under low nutrient conditions utilizing reduced gases for energy (J.K. Fredrickson, Pacific Northwest National Laboratory)."

Based on previous, although somewhat limited, knowledge the overall phylogenetic diversity of subsurface microbial communities is no greater or less than in other environments. However, adaptations that allow for function and survival in the deep subsurface should be reflected in abundant unique genes that will initially be characterized as unique hypothetical genes based on comparative DNA sequence analysis. Comparative genomic approaches can be used at the organism or community levels to test these hypotheses. Functional genomics, however, will ultimately be required to unravel the role of these unique, "deep subsurface" genes. For example, because the concentrations of carbon and energy sources will be low, the microbiota would be expected to have highly adapted systems for assimilating nutrients at such concentrations, i.e., enzymes with low Km values.

A6. "The milling process for Au with the addition of cyanide and a 'litany of other chemicals', followed by treatment and subsequent filling of this waste into excavated stopes in various parts of the mine have stimulated the extant and allocthonous microbial community to degrade organic components found in the mine wastes over the long time periods the mine has operated. These activities and the small spills and leaks of fuel, transformer fluids, solvents, and other toxic chemicals make the mine an excellent source for biodegraders that have unique abilities to bioremediate a variety of toxic organics. (T. Hazen, Lawrence Berkeley National Laboratory)"

It is well known amongst microbial ecologists that the best place to find microorganisms that can degrade toxic compounds is in environments where the microbial community has been exposed to them for long periods of time (decades). Indeed, the original patent on life was a bacterium capable of degrading crude oil that was isolated from soil around an oil well that had been in operation for many years. The Homestake mine operations over more than 125 years has resulted in a very large number of small toxic chemical introduction events and a very large number of large scale low concentration chemical introduction events into the deep subsurface. The age of the mine and the fact that many of these point sources have been isolated from air for various lengths of time suggest that there could be a plethora of different organisms capable of degrading toxic chemicals that could be used in other operating mines and in other types of deep and shallow subsurface contaminated areas, especially fractured rock environments. By examining these stopes and spills of different ages we could also determine the feasibility of natural attenuation of these contaminants in mines and fractured rock environments. Since the stopes, the milling process, and handling have been quite similar through the years we could determine rates of biodegradation and the natural succession of microbial communities in contaminated deep subsurface environments. This has implications for clean-up strategies for mines and a variety of fractured rock environments. Homestake mine has already patented a cyanide degrading bacteria that they isolated from their mine wastes and has been using it in their mill treatment process for a number of years. These studies may also have implications for the environmental impact of future activities, including spills and unintentional releases of chemicals used by NUSL physics experiments and detectors, e.g. tetrachloroethylene.

B. Subsurface biogeochemical processes
Subsurface biogeochemical processes emphasizes the role of microorganisms in the dissolution and precipitation of mineral phases, the transport /transformation of chemical species, and the alteration of hydrological properties (i.e. storage capacity and permeability) of aquifers. This information has vital applications to deep carbon sequestration and nuclear and non-nuclear waste disposal and stabilization. Experimentation can provide direct measurement of the release rates of CO2 from the injection zones or of entrapment in the formation, of the chemical form and mobility of waste, and the survival of microorganisms in different thermal and chemical regimes that may be imposed by borehole injection or repository conditions. Some of the hypotheses to be investigated include those below.

B1. "Microbiologically-induced calcite precipitation contributes to mineralization in subsurface environments. Common soil microbes (e.g., Bacillus pasteurii) participate in CO2 sequestration in alkaline environments. It is likely that subsurface microbial communities will contribute to CO2 sequestration in deep subsurface environments (S. Bang, South Dakota School of Mining and Technology)"

Many anaerobic microbial processes lead to substantial increases in the pH of the environment, especially NH3 production (Bachmeier et al., 2002) and Fe(III) and sulfate reduction, respectively. This elevation of the pH is essential in order to transform dissolved CO2 pumped into a subsurface aquifer into more soluble HCO3- and CO32- species that lead to the permanent sequestration of CO2 as solid phase carbonate (e.g. calcite or siderite). The microbial activities that occur at the interface of supercritical CO2 and saline groundwater can be directly examined at NUSL-OSG using instrumented fracture zones maintained. Supercritical CO2 can be leaked into the fracture system at ambient formation pressure and its concentration, the speciation and the resulting chemical reactions monitored downgradient of the injection under a range of microbial redox conditions. This approach will provide insight into the storage capacity of a subsurface environment for CO2 and whether any environmentally hazardous outcomes result from such a mitigation approach. These experiments will also have enormous ramifications for any postulated subsurface microbial environments on Mars where liquid to supercritical CO2 has recently been postulated to exist (Hoffman, 2001) and on the importance of dissolved inorganic carbon to the sustenance of autotrophic communities (Stevens and McKinley, 1995). Our preliminary investigations indicate that the groundwater at Homestake is carbonate buffered, but other groundwater composition may be found, in which case microbial precipitation of noncarbonate phases can also be explored.

B2. "In deep subsurface environments where gas and water occur as separate phases, microbial communities may be concentrated at the interface between gas and water where the flux of dissolved gases into the water is the greatest. (T. Onstott, Princeton University)"

Investigations into subsurface communities to date have focused upon the relationship of microbial communities and the mineral-liquid interface. In natural gas reservoirs and in some of the Au mines of South Africa, both gas and liquid are present as distinct phases. The gas constituents, typically composed of methane, light hydrocarbons and H2, are all excellent electron donors when dissolved in water, which in turn contains electron acceptors. As microorganisms consume these electron donors, they are replenished by diffusion from the gas phase resulting in a concentration gradient. Although such an environment is easy to conceive, observing such a system is virtually impossible without having access to a fluid filled fracture that is or can be partially dewatered. Homestake may provide an opportunity to study the geochemistry and microbiology of such fracture systems. To test this hypothesis requires either detailed spatial and temporal sampling of a gas-water fracture zone or experimental simulation of a gas-water system utilizing a high-pressure manifold.

B3. "The composition of the indigenous microbial community will reflect the geochemical properties, in particular the dominant electron acceptor, of the hydrogeological regime. (T. Onstott, Princeton University)"

In previously published investigations of the subsurface communities in hydrocarbon reservoirs and in reports from deep subsurface communities in the Au mines of South Africa and Aspo in Sweden, the environments are typically rich in electron donors, particularly hydrogen, but limited by the bioavailability of electron acceptors, e.g. nitrate, Fe(III), sulfate or CO2. The Fe-rich Homestake Formation ranges from 0-125m in thickness because of plastic deformation. It is generally a silicate-carbonate-type iron formation, typically containing 25-30 wt% FeO+Fe2O3 with an average FeO/(FeO+ Fe2O3) of 0.79-0.88. Fe-carbonates (ankerite, siderite), Fe-chlorite and stilpnomelane occur in the low metamorphic grade rocks. In higher grade rocks fewer Fe-carbonates and more Fe-silicates (grunerite, almandine, Fe-biotite) exist. In the ore zones, on the other hand, the Fe sulfides are abundant (arsenopyrite+pyrrhotite+/-pyrite). Magnetite and ilmenite are quite minor or absent. Almost all iron mines are surface excavations, so Homestake provides a unique window into the role of Fe in subsurface microbial communities. In the Fe sulfide rich portions of the formation, Fe(II) and S can act as a potent electron donors leading to the in situ formation of Fe(III) weathering products. Where these oxidized products are abundant, Fe(III) reducing bacteria should be the dominant component of the microbial community. Where the reduced Fe(II) carbonate strata exist, CO2 reducers or autotrophic microorganisms may dominate.

C. Subsurface microbial ecology
Subsurface microbial ecology explores the evolution and adaptation of life in the deep subsurface; the relationships between microbial community structure and function and the sources of nutrients/energy that allow them to survive and reproduce, in some cases, independently of photosynthesis; identification of the subsurface chemical, geological and hydrodynamic properties that contribute to the sustenance, migration, adaptation and evolution of life in the deep subsurface and beyond. Importantly, these studies will provide an understanding with which to approach investigations of life on other planets and perhaps extra-terrestrial environments. Some of the hypotheses to be investigated include those below.

C1. "The rate of fluid flux through subsurface fracture networks should dictate the quantity and type of microorganisms present. High fluid fluxes should lead to greater biomass. Variable fluid fluxes should create greater diversity. (F. Colwell, Idaho National Engineering and Environmental Laboratory)"

Fluid flux is believed to be a key component of whether microbial cells flourish or perish in the subsurface. Electron donors and acceptors are required for energy conservation. Without some movement through the fracture system, however, life will only exist proximal to those sources of energy for a brief moment in geological time, before life reduces the chemical free energy to zero. This hypothesis could be tested at the NUSL-OSG using two approaches. The first approach is to combine measures of microbial biomass and diversity on samples of groundwater and rock cores adjacent to fractures with noble gas isotopic analyses. In fractures where fluid migration rates are rapid (~1 m/yr), the noble gas isotopic composition of the fracture fluid will be distinct from that of the rock's pore water. In fractures where fluid migration rates are extremely slow (~1 mm/yr) the noble gas isotopic composition of the fracture fluid will be similar to that of the rock's pore water. The second approach is to install into a borehole-intersected fracture zone a device that allows: 1) regulation of the rate of flow through the fracture zone and thus the flux of groundwater substrates and 2) insertion of some colonizable and retrievable substrates that can be evaluated over time to determine the relationship between flux rate and biomass and diversity.

C1.1. "Different genes should be expressed by subsurface microbial communities in fractures depending upon the rate at which fluids pass over the communities. (F. Colwell, Idaho National Engineering and Environmental Laboratory)"

Because the fluid flux will control the relative concentrations of energy and growth substrates and toxic species in the fracture environment, existing subsurface microbial communities must have metabolically adapted to this nutrient flux, otherwise they would have expired. One approach to observing these adaptations under forced flow conditions is monitor the expression of genes responsible for specific electron transport pathways.

C2. "Spontaneous mutagenesis will be less in subsurface environments shielded from cosmic rays and low in radiogenically generated gamma radiation than in soil zones. (P. Zimmerman, South Dakota School of Mining and Technology)"

The rate of microbial evolution in the subsurface is affected by growth rates, spontaneous mutagenesis, the rate of genetic exchange and the rate at which the environment changes. Growth rates in the subsurface are much less than those in surface marine and terrestrial environments and hence the rate of DNA repair or propagation of altered DNA is also much lower. Depending upon the rate of spontaneous mutagenesis by radiation, mutagenesis by DNA damage and growth may not be the principal mechanism for evolution of subsurface microorganisms. NUSL-OSG offers an opportunity to study the effects of ambient radiation on mutagenesis in an environment in which the radiation flux and spectra are extremely well established. Mutation rates could quantified by utilizing the; 1) Ames test, which uses an auxotrophic mutant strain of Samonella typhimurium and text for back mutations in a single gene, and 2) bacteria for which the entire genome has been sequenced. These bacterial dosimeters can be situated at different depths within NUSL and their spontaneous mutagenic rate compared as a function of ambient radiation.

C3. "In ancient, deep subsurface groundwater environments where electron donors are abundant, but electron acceptors are limited microorganisms will either possess metabolic plasticity to utilize several electron acceptors and or live synergistically with other microorganisms that consume or cycle their waste products. (T. Kieft, New Mexico Institute of Mining and Technology)"

Stevens and McKinley (1997) have observed that in the H2 rich subsurface environment of the Columbia River Basaltic Aquifer, that acetogens (utilizing H2 and CO2) generate acetate and postulate that this acetate my be utilized by heterotrophic microorganisms, such as acetate oxidizing sulfate reducers, to regenerate the CO2. The syntrophic relationship ensures that the acetate concentrations never approach the level that shut down the autotrophic activity of the acetogens. If ancient, geohydrologically isolated fluid filled fractures exist at Homestake, then the microorganisms they contain are likely to have been isolated from the surface for millions of years. By comparing the microbial communities and geochemistry of these environments with those found in fractures where the fluid is being more directly recharged from the surface will determine to what extent deep subsurface microbial communities are dependent upon transport of life-sustaining substrates from the surface. Such a comparison will also reveal what adaptations these microorganisms have made for extremely long-term survival.

C4. "The large scale transport of shallow subsurface or soil microorganisms to the terrigenous deep subsurface through fracture zones will be more limited by their ability to survive and grow in the deep-subsurface rather than by their transport properties. (T. Kieft, New Mexico Institute of Mining and Technology)"

The concentration of microorganisms in soils are many orders of magnitude greater than that of solid rocks in the deep subsurface; whereas, the concentration of microorganisms in shallow groundwater is only two orders of magnitude greater (Whitman et al. 1998; Onstott et al., 1998). Are soil microbes ultimately the feedstock for subsurface microbial communities? In the absence of a dynamic, hydrothermal fluid system, e.g. deep-sea vents, fluid velocities in the terrigenous crust are typically very slow. At such velocities, the adhesion of microorganisms to solid mineral surfaces are so high that the planktonic cell density of downward migrating soil microbes would be reduced by three orders of magnitude long before they have reached the deep subsurface. Consequently, only those surface microorganisms that can readily grow in the anaerobic, nutrient limited, higher temperature environments will ultimately appear as a member in the subsurface community. Those that do not grow will be quickly winnowed from the water mass by adsorption to the rock matrix. Two approaches can be followed to examine this hypothesis. The first is to document the microbial community structure as a function of depth along a fracture system that is demonstrably connected to the surface. If soil microorganisms are penetrating over a kilometer into the crust, their characteristic 16SrDNA signature should be found in the subsurface communities. A second approach is to instrument several well-characterized fracture zones, NUSL-OSG will be the only facility in the world to be able to perform microbial transport experiments in a fractured rock system over a depth range of kilometers. Using the same instrumentation described above for fluid flux experiments, surface organisms collected at Homestake can be injected into the fracture zones at depth and their transport and growth monitored relative to that of subsurface microorganisms adapted to the fracture environment.

C5. "The base of the subsurface biosphere is currently pinned at 120oC, the higher survival temperature known for a microorganism in the lab. The upper temperature limit of life in the subsurface however is controlled by a balance between the energy available for hyperthermophiles to repair their cell membranes, enzymes and DNA and the thermal stability of their membranes, enzymes and DNA. This may mean that unlike the laboratory experiments and the deep-sea vents where nutrient fluxes are very high, the upper temperature limit for life in the terrigenous subsurface is much less than 120°C. (T. Onstott, Princeton University)"

The geothermal gradient at Homestake Mine is 22.5oC/km (Ashworth, 1983) corresponding to a formation temperature of approximately 55oC at the 7000 ft. (2.1 km) level. From a small drilling platform located at the 8000' level vertical coring to depths of 16,500' (or an additional 2.5 kilometers) should reach depths for which the formation temperature is 120oC, the highest temperature limit for known life forms. By undertaking a small coring program, NUSL-OSG could provide the first unequivocal evidence for the upper temperature limit of life in the deep subsurface. Once completed the packered borehole could be utilized to test the survivability of known hyperthermophiles in this oligotrophic environment by submerging them into the borehole on filter enclosed, rock chip coupons.

C6. "As surface microbes enter a deep-subsurface microbial community the potential for genetic exchange between the two populations exists. Do genetic elements from surface communities survive and become stably incorporated into the deep-subsurface microbial community? (T. Kieft, New Mexico Institute of Mining and Technology and (T. Hazen, Lawrence Berkeley National Laboratory)".

The Homestake Mine has been in continuous operation for more than 125 years. The mine has more than 300 miles of drifts at more than 50 levels down to 8000 ft. For decades during the early days of it operations horses and mules were used in the mine and even raised there. Until a few years ago human waste was disposed of directly in the mine. These activities combined with the moving of rock and materials in and out of the mine resulted in the inoculation of surface microbes into the deep subsurface community. NUSL-OSG would have a unique opportunity to study plasmid and nucleic acids from microbial communities that were exposed to surface microbes over a century ago or months ago and determine resiliency of both the genetic elements that may have been transferred and the resiliency of the surface microbes plasmids and genetic material in deep-subsurface environments. Homestake has stopes and drifts that have been isolated, after their initial exposure, for various periods of times that allow a unique time and dispersal perspective on these types of studies.

C7. "Pathogens and their indicators were introduced into the mine at all levels through the disposal of human and animal waste in the mine drifts. These pathogens and indicators may have been able to survive for extended periods of time due to the thermal nature of some parts of the mine, especially if water and organics were abundant in the areas they were introduced. Since many of these human and animal source microbes may have harbored unique plasmids for antibiotic resistance, and pathogenicity the may have been able to horizontally transfer these genetic elements via conjugation or transformation to indigenous species. The survival of pathogens and the transfer of human/animal genetic elements to subsurface or mine microbes have tremendous implications for human and environmental impact. (T. Hazen, Lawrence Berkeley National Laboratory)"

Only in the last several years of mine operation have human and animal wastes (early on horses and mules were used in the mine) been removed and disposed of at the surface. Since both human and animal waste contains potential pathogens, and benign organisms with antibiotic resistance plasmids or virulence plasmids, the mine represents a unique place to study survival and transfer of unique genetic elements in the deep subsurface. Pits used for disposal of urine and feces could be sampled for unique organisms found only in man or animals and for indigenous species that contain genetic elements that are unique to man or animals. A number of studies have shown that tropical environments, with higher soil temperatures, higher moisture, and higher organics can support pathogen/indicators like Escherichia coli indefinitely (Hazen and Toranzos, 1990) and facilitate the transfer of genetic elements to indigenous species (Cruz-Cruz et al., 1988). The higher temperatures, presence of water, and high organic inputs in the stopes and in spill and disposal areas suggest that increased survival and transfer of genetic elements to indigenous organisms may also be possible. These studies will also directly aid environmental threat analyses from both accidental and deliberate releases of human and animal pathogens.

EDUCATIONAL OBJECTIVES

NUSL-OSG will be involved in international, national, regional and local educational outreach at all levels from casual site visitors to K-12 to graduate and post graduate and through to senior scientists. Out reach programs will include the following:

1. NUSL-OSG would be responsible for the biological component to a scientific visitors' center at NUSL.
2. Secondary school workshops focusing on local schools, and real time instruction aides over the Internet for regional schools with lesson plans for K-12.
3. Secondary school teacher training workshops focusing on local schools and supplied with educational aides and video tours of excursions.
4. Deployment of experiments designed by local and regional high school students and that can be monitored real time through the Internet. The International/High Plains Regional Science and Engineering Fair provides an ideal venue for organizing the student participation and the mentoring experience by their teachers. Experiments could include the insertion of media-bearing cartridges designed by the students into boreholes along with pH, Eh, O2 sensors connected to short-term memory storage at the borehole head that can be periodically downloaded through the Internet. The media cartridges can be removed for analyses at the high school or electron microscopic observations at EMES with images displayed real time on the Internet.
5. Summer field research institute-training-local, national, and international college students and senior scientists-workshop modeled after the summer program of the Marine Biological Lab (MBL) at Woods Hole and the recent minority, educational workshop recently held in South Africa http://web.utk.edu/~kdavis.
6. NUSL-OSG will host an REU site for undergraduates nationwide and targeting minority groups. This could be coordinated with the pending REU for South Africa where students have an opportunity to attend both and including excursions and experiments examining biogeochemical processes of samples.
7. NUSL-OSG will target the Native American community in South Dakota at the secondary school and college level and internships, undergrad, grad, postmasters, postdoctoral programs.
8. Undergraduate and graduate students funded through NSF's IGERT Program at Oregon State University and Portland State University (Subsurface Biosphere Interdisciplinary Doctoral Program) could conduct research at NUSL-OSG.
9. NUSL-OSG could host visiting scientists intent on performing underground experiments and conferences for such societies as the ISSM (International Society for Subsurface Microbiology) and the ISE (International Society for Extremophiles).

IMPLEMENTATION
During the initial 3 to 5 years, the primary focus of the geomicrobiological effort at NUSL will be on examining the biological diversity of the deep subsurface microbial at Homestake and establishing an educational and outreach program. This requires gathering observational and experimental data over a multi-year time scale from a variety of subsurface settings. The early establishment of a NUSL-OSG that coordinates this multidisciplinary research effort with educational objectives is essential to its successful implementation. NUSL-OSG should be comprised of a surface facility with laboratories and support personnel and subsurface experimental facilities at several localities where the migration, diversity and metabolic activity of microbial communities can be monitored over extended time periods. More sophisticated laboratories and supervising faculty can be cited at the local universities (see Appendix 1 as an example of facilities available), complemented by affiliated national and international institutions where highly specialized measurements (e.g. ion probe and accelerator mass spectrometer analyses) can be performed.

The NUSL-OSG will be the only facility of its kind where long-term experiments can be performed by the international community in fractured, heterogeneous, metamorphic rock with manned infrastructure at depths up to 2.7 kilometers and with deeper flow fields, boreholes and screened wells extending beyond 4 km beneath the surface. Microbiological observations and experiments conducted at NUSL will build upon 15 years of research investment in subsurface biogeochemistry sponsored by the Department of Energy's Subsurface Science Program and the NSF's Life in Extreme Environments (LExEn) program, now both defunct. The data gathered from this lab will complement observations that have been reported over the last 10 years from the shallower 400-meter deep Äspo Underground Laboratory in Sweden, where experiments have just recently begun in underground facilities hosted by a fractured, homogenous, granite aquifer. Long term observations made at NUSL can also be compared to shorter-term data gathered in active South African mines, where samples collected from depths as great as 3.5 kilometers have been obtained. Long term experiments performed at NUSL-OSG could address many questions raised by the observations made in South African mines and in boreholes or mines around the world and could be designed to test the microbial impact on subsurface CO2 sequestration and underground radioactive waste disposal. The NUSL-OSG could also provide an invaluable test bed and training ground for and opportunity for technology transfer to NASA engineers attempting to develop methods for detecting life beneath the surface of Mars. Finally, NUSL-OSG could tap into a new gene pool with potentially valuable biotechnological applications by fostering interactions and CRADA's with biotechnology and pharmaceutical firms.

PART 2-PROPOSED SHORT TERM RESEARCH AND INFRASTRUCTURE ACTIVITIES

As stated previously the research thrusts of the NUSL-OSG are; 1) the analysis of subsurface biological diversity, 2) documenting subsurface biogeochemical processes, and 3) the examination of subsurface microbial ecology and interactions between subsurface and surface microbial communities. Some of the hypotheses (e.g. C6) associated with these research thrusts require a multi-institutional, multidisciplinary research team to acquire, process and analyze water, gas/air and rock samples during the early phase of NUSL construction as drift advancement at the 7400' level and the downward extension of the Yates Shaft enters virgin rock formations (Fig. 1). Other hypotheses (e.g. C3) can take advantage of the various environments offered by the present, extensive mine workings and can be phased in over time. For all objectives, the development of the NUSL-OSG should be closely coordinated with other research activities and with NUSL construction in order to make maximum use of resources and take advantage of complimentary scientific capability. Below is an outline of proposed activities, required facilities and how these should relate to NUSL plans.

1. Hypotheses A3, A4, A5, B2, B3, C1, C3 and C5. Ancient, thermophilic, indigenous, subsurface microbial communities. The geothermal gradient at Homestake Mine is 22.5oC/km (Ashworth, 1983) corresponding to a formation temperature of approximately 55oC at the 7000 ft. (2.1 km) level. This temperature is high enough to select for moderately thermophilic microorganisms. The available paleothermal history information suggests that these rock formations experienced much higher temperatures approximately 50 million years ago during a hydrothermal episode associated with intrusive igneous rocks. Subsurface microbial communities adjacent to these intrusive units would have been eliminated, but others may be preserved in hydrologically isolated pockets of fluid and gas-filled fractures and pore water of rock far removed from the Tertiary, hydrothermal impact. These environments could harbor ancient microbial communities, possibly representing a relatively small number of generations, surviving for hundreds of thousands of years or more under nutrient-limited conditions. The goal of this research is to characterize these environments using microbiological, molecular, geochemical, and isotopic techniques designed to establish community size, diversity, physiological characteristics, energy resources, age and origin.

A. Proposed Activities
1) Coring and borehole logging should occur concurrently with the construction of new underground excavations. It requires the implementation of specialized drilling, tracer, core handling and core preservation procedures for recovery, isolation and evaluation of (perhaps previously unknown) microorganisms from the rock cores and from water emanating from fractures intercepted by the borehole. Because drilling has to precede the kilometer of tunneling that will take place during the NUSL construction phase and as a precaution against water incursions into the workings, we propose to coordinate or piggyback this sampling activity with the construction phase of NUSL. This approach offers several advantages including increasing the assets of the geomicrobiology center during the first year (i.e. unique microbial isolates), identifying critical locations for future experiments, avoiding additional drilling costs, supplying information to the NUSL construction team on the types of water and gas and their origin, whether they represent a potential threat to NUSL and the best ways to avoid or to seal the ingression. As a result, the involvement of academic hydrologists and geochemists during the construction phase will reduce the construction costs.
2) Microbial, molecular, geochemical, isotopic, petrographic and petrophysical characterization of fluids, gases and cores from drilling activity will focus on evaluation of fractures, because significant fluid flux and microbial communities within consolidated rock are mainly confined to these structures. Characterization of the fractures encountered is a high priority, because uncontaminated fractures can only be obtained by subsurface coring at depth as opposed to coring downwards from the surface. The petrophysical information on the cores and fractures will be useful to engineers responsible for assessing the structural integrity of the underground workings. Samples of the air and drilling water will also be analyzed and the microbial assays will be particularly valuable to engineers responsible for ventilation and cooling of the NUSL.
3) Preservation of boreholes of scientific interest will be important (i.e., cementation and casing would be avoided provided this does not pose an environmental threat to the NUSL operations. These boreholes can be capped for future experiments with their effluent channeled to sump pumps. At a later stage, multilevel samplers can be installed so that individual fractures can be isolated and sampled separately. The types of experiments that can be performed include radiotracer and stable isotope experiments to delineate carbon, nitrogen, phosphorous and sulfur cycling. The detectors utilized by the physics experiments at NUSL may provide a means for quantifying extremely low-level microbial activity rates. The high ambient pressures in the boreholes can be used to isolate barophilic microorganisms and to test the effects of super critical CO2 injections.
4) From a small drilling platform located at the 7400' level at the base of the extended Yates Shaft or at the 8000' level (Fig. 1) vertical coring to depths of 16,500' (or an additional 2.5 kilometers) should reach depths for which the formation temperature is 120oC, the upper temperature limit for known life forms. The properties of microbial communities discovered under these conditions will be compared to those found in Yellowstone hot springs to the west and to deep-sea hydrothermal environments. Coring at shallower depths should be used to develop a vertical or depth profile of the indigenous microbial communities to delineate changes in the energy sources and growth substrates as a function of depth. Specific comparisons of microbial communities from unsaturated depths to those from saturated strata will provide insight into how microorganisms stratify with regard to water activity. Detailed coring of a gas/water-bearing fracture could determine whether microbial community structure and total biomass is concentrated at the water/rock or gas/water interface.

B. Facilities and Footprint
1) Laboratory facilities at the surface should be established for this initial phase of fieldwork. These facilities would house equipment for the measurement of tracers, sterilization of sampling equipment, preparation of media for microbial culturing, chemical stockroom, incubators (20-120°C, at 10°C intervals), a simple DNA extraction and amplification laboratory, a filtration laboratory, a wet chemistry laboratory, and freezers for preservation of cores and enrichments, Access to a machine shop for preparation of field sampling tools will be required as well as microscope facilities for examination of physical tracers or cells from enrichments. The above ground facilities

Fig. 1. Cross section of Homestake mine indicating location of NUSL excavation and focus of geomicrobiological research activity in the first few years.

should be a few thousand square feet (2-5) that could potentially be dispersed among a few nearby buildings. In out years, should the resident research staff exceed 25 FTE's then expansion beyond 15,0000 square feet may be justified. Augmentation of this laboratory or local academic institutions with more sophisticated equipment, such as DNA sequencers or GC mass spectrometers or teaching laboratories could occur later.
2) Drilling of boreholes either as part of underground construction or specifically for the installation of scientific experiments will require the use of drilling rigs. The footprint for the rig would typically be less than 20x20x20 ft. Most are typically smaller, with small drills requiring floor space of only 8 x 10 ft with a 5 ft high clearance for periods of days to a few weeks. For the drilling/stabilization of the largest 2-3 km deep holes, operations may extend over a period of a several months. The monitoring and sampling equipment for long term observations would typically require footprints of 2 ft deep by 4-8 ft long by 5-8 ft high, and only during scheduled 2 hour sampling visits. Instrumented boreholes in the newly excavated portion of NUSL would contain multilevel samplers from which fluid and gas emanating from the fractures can be piped into a sterile, anaerobic laboratory for collection and preservation of samples. The lab could be the size of a very small house trailer or large meat-cooler room of 500-800 sq feet that can be easily be assembled from weatherproof panels. The facility would accommodate an anaerobic glove bag, sample processing equipment, supplies, and would include digital sensor/loggers that monitor the temperature, pH, Eh, salinity, etc. of each fracture. The multilevel sampler would also enable injection into fracture zones. Access to these boreholes on a daily basis would be required.

2. Hypotheses A1, A2, A6, B1, C1, C2, C4, C6 and C7. Adaptation and transport of surface microbial communities to deep subsurface environments and environmental changes. The present mine workings contain well-developed biofilms wherever water weeps from boreholes or fractures. The mining activities undoubtedly carried microbial contaminants from the surface and provided opportunities for colonization by rare subsurface microbes to take advantage of the energy afforded by coupling the oxidation of reduced chemicals in the rock and water to O2 respiration (provided by ventilation). Mining activities have also dewatered the rock formations and replaced a hot, reducing environment with a cool, oxygenated one. Many stopes also exist that have been isolated from the mine environment at different times in the past and are potentially returning to their original state. The old mining galleries, therefore, provide a unique opportunity to document the penetration and adaptation of surface microbial communities to subsurface environments over the multideccenary time scale. Such experiments are impossible to conduct in the laboratory or with present day field experiments performed in fractured rock environments where the average hydraulic conductivity is 10-8 cm/sec as is the case for the formations at Homestake. Such information is not only valuable to understanding the mechanisms and rates at which the subsurface biosphere interacts with the surface, but it is relevant to developing subsurface bioremediation strategies in fractured rock aquifers and assessing the microbial impact on the subsurface storage of high level radioactive wastes. The lattermost application is strongly supports research underway at the Äspo underground laboratory in Sweden. Despite the importance of examining, microbial transport or colonization in fractured rocks in zones of high fluid flux, field sites do not exist for this purpose, even at Äspo.

A. Proposed Activities
1) Utilizing the same techniques as in the first category, the microbial communities present in open tunnels and isolated stopes of varying ages would be characterized. The rate of colonization of new tunnels is highly dependent upon the location with respect to the direction and rate of ventilation. For example, corrosion rates of metals are much higher down gradient of hot and humid return airflow towards the fan drifts than dry and cool air entering from the elevator shafts. These studies would provide valuable guidelines to engineers responsible for ventilation of NUSL and maintenance of the scientific equipment. In isolated stopes, the air should become more anaerobic with time and the water, which initially may be acidic, would become more alkaline with time. The temporal trends in microbial communities and aqueous geochemistry can be utilized to estimate long-term corrosion rates, which can be compared to rates derived from in situ activities derived from radiotracer experiments. Microbial samples from isolated stopes may also provide information on the rate of microbial mutation or genetic exchange in the subsurface environment. The extensive biofilms present at various levels can be examined in terms of the types of minerals being precipitated and the ongoing biomineralization processes. This information can be compared to the surface mineral composition of the rock as mapped by laser and infrared imaging of the tunnel face.
2) Bacterial transport through well-characterized fractures in the old galleries will be examined using both indigenous and nonindigenous microbial species (including spores) with select adhesion properties. Microbial transport rates will be compared to that of conservative and nonconservative tracers and studied as a function of water tension in the dewatered portions of the formation and salinity and pH in the deeper fractures and rock types, i.e. Ellison versus Homestake versus Poorman formations. Locations for these experiments can probably be selected from among those galleries where mining water is known to seep downwards from higher levels. Natural tracer experiments could further refine the flow paths and rates. Sidewall coring could be used to establish a nest of boreholes containing multilevel samplers designed to sample individual fractures identified in the cores. Characterization of the fracture geometry across two or more levels could be performed using forced gradient tracer tests and cross borehole tomography. Instrumentation of the multilevel samplers with autosamplers would complete the subsurface fracture flow field. Such a facility would be truly one-of-the-kind. Maximum transport distance will probably be 100 meters, but could be longer depending upon the survival of the microbial tracer. Again, once the flow fields are established the long-term footprint is quite small, i.e. less than 3 x 8 ft with 6 ft high clearance at each well-head. During periods of intense activity, access to hundreds of square feet of local tunnel space may be required periods of days to weeks. Timing could be arranged to minimize impacts on other uses of the tunnel tracks

B. Facilities and Footprint
1) The laboratory located at the surface should be equipped for measurement of some tracers and preparation of inocula for microbial injection. Local academic institutes would facilitate preparation of injected microorganisms and analyses of breakthrough results.
2) Two or three flow cells distributed in different portions of the mine should be enough to sample the variety of environments that the old galleries offer. One would be situated in the dewatered zone to examine microbial transport under vadose conditions. Each borehole nest on a level would also contain a small, trailer-size laboratory where effluent from the multilevel samplers would be collected with autosamplers.

IMMEDIATE RECOMMENDATIONS

1. Characterization of existing fluid emanating from boreholes and addition of this information and the mine records of water intersections to the 3-D spatial map for Homestake. This level of characterization will identify potential water-rich fracture zones and provide a basis from which to coordinate geomicrobiological sampling/experimentation with NUSL excavations in more detail. Larry Stetler and Heidi Sieverding of South Dakota School of Mines have volunteered to coordinate this effort.

2. Close coordination of construction activities with research activity1 above. Dr Tom Langworthy of the Univ. of South Dakota will serve as principal liaison with the planning committee for phased construction of the laboratory. A geomicrobial advisory committee should be formed to provide feedback to Dr. Langworthy on requirements.

3. Planning for a NUSL-OSG center: infrastructure, outreach to other sciences and scientific societies, the path forward and NSF and South Dakota agencies. Workshops, booths at national meetings, ASM, AHU, AAAS, to identify collaborators, affiliates, , international interest.

4. Development of a budget for supporting activities and for performing sampling and analyses during NUSL construction (to Oct. 2002?). This budget should include support for geomicrobiologists (PIs and students) to travel to Homestake, to conduct appropriate characterization studies at Homestake, and in their respective laboratories, workshops, planning outreach, service to NSF and to the state of South Dakota.
REFERENCES

Adams, M.W.W., Perler, F.B. and Kelly, R.M. (1995) Extremozymes: Expanding the Limits of Biocatalysis. BIO/TECHNOLOGY 13: 662-668.

Ashworth, E. (1983) The application of finite element analysis to thermal conductivity measurements. MSc. Thesis, South Dakota School of Mines and Technology, Rapid City, South Dakota, pp.

Bachmeier, K., Williams, A.E., Warmington, J., and Bang, S.S. (2002)Urease Activity in Microbiologically Induced Calcite Precipitation, J. Biotech. 93:171-181.

Cruz-Cruz, N. E., G. A. Toranzos, D. G. Ahearn and T. C. Hazen. (1988) In situ survival of plasmid and plasmidless Pseudomonas aeruginosa in pristine tropical waters. Appl. Environ. Microbiol. 54:2574-2577.

Fredrickson, J. K., R. J. Hicks, S. W. Li and F. J. Brockman (1988) Plasmid incidence in bacteria from deep subsurface sediments. Appl. Environ. Microbiol. 54:2916-2923.

Fredrickson, J.K. and Onstott, T.C. (1996) Microbes Deep inside the Earth, Scientific American, v. 275, No. 4, p. 68-73.

Hazen, T. C., and G. A. Toranzos. (1990) Microbiology of tropical source water. In: Advances in Drinking Water Microbiology research (Ed.) G. A. McFeters, p 30-51. Springer-Verlag, New York.

Hoffman, N. (2000) White Mars: A new model for Mars' surface and atmosphere based on CO2, Icarus, 146:326-342.

Jimenez, L. (1990) Molecular analysis of deep-subsurface bacteria. Appl. Environ. Microbiol. 56:2108-2113.

Nealson, K. and Ghiorse, W.A. (2001) Geobiology-A report from the American Academy of Microbiology. American Academy of Microbiology, Washington, D.C. 16 pp.

Onstott, T. C., Phelps, T. J., Kieft, T., Colwell, F. S., Balkwill, D. L., Fredrickson, J. K., Brockman, F. J. (1998) A global perspective on the microbial abundance and activity in the deep subsurface, In: Seckbach, J. (Eds.), Enigmatic microorganisms and life in extreme environments, Kluwer Publications, 489-499.

Stevens, T. O., McKinley, J. P. (1995) Lithoautotrophic microbial ecosystems in deep basalt aquifers. Science 270:450-454.

Whitman, W. B., Coleman, D. C., Wiebe, W. J. (1998) Prokaryotes: The unseen majority. Proceedings of the National Academy of Sciences of the United States of America 95:6578-6583.

AUTHORS

T.C. Onstott-Princeton University; Jim Fredrickson and Duane Moser-Pacific Northwest National Laboratory, Ray Wildung- Pacific Northwest National Laboratory, Terry Hazen-Lawrence Berkeley National Laboratory, Tommy Phelps-Oak Ridge National Laboratory, Susan Pfiffner-University of Tennessee, Tom Kieft-New Mexico Institute of Mining and Technology, Tom Langworthy-Univ. of South Dakota, Vermillion, Mary DeFlaun-Envirogen, Inc., Rick Colwell-Idaho National Engineering and Environmental Laboratory, Sookie Bang-South Dakota School of Mines and Technology and contributions from Bill Roggenthiem- South Dakota School of Mines and Technology and Joe Wang- Lawrence Berkeley National Laboratory.

Appendix 1.

Analytical Facilities - Engineering and Mining Experiment Station (EMES)

Voice: (605)-394-2496
Facsimile: (605)-394-5360
Internet: eduke@silver.sdsmt.edu
http://www.sdsmt.edu/es/emes/

Offices located in the Mineral Industries Building, Room 234

X-Ray Diffraction (XRD)
The X-ray diffraction facility maintains two fully automated instruments: a Siemens-Nicolet transmission diffractometer, and a Philips Bragg-Brentano diffractometer. Qualitative identification of a broad variety of solid phases is possible, in addition to quantitative determination of the weight percents of the various components.

Electron Microscopy
Scanning Electron Microscope (SEM)
The SEM laboratory features a JEOL JSM-840A scanning electron microscope equipped with an Oxford Instruments energy-dispersive X-ray analyzer for elemental analysis. This combination provides simultaneous morphological and chemical characterization of specimens at magnifications of 10X to 100,000X. Output options include secondary electron and backscattered electron images, X-ray point analyses, X-ray element distribution maps, X-ray line profiles, and digital image processing. Gold sputter coating and carbon-evaporation coating are available for sample preparation. An Oxford Instruments liquid-nitrogen freezing/heating stage is available for operations at temperatures in the range -185°C to +200°C.
Transmission Electron Microscope (TEM) and
Scanning Transmission Electron Microscope (STEM)
The TEM/STEM laboratory is equipped with a Hitachi H-7000 FA transmission electron microscope that includes a H-7110 scanning transmission (STEM) module and a Kevex energy-dispersive X-ray spectrometer. The instrument has demonstrated resolution of 1.44Å in TEM mode, 10Å in STEM mode, and 20Å in SEM mode. Single and double-tilt specimen holders are available, and sample preparation facilities include dimple grinding, jet electropolishing, and ion milling.

Scanning Probe Microscopy (SPM)
The scanning probe microscopy laboratory features a Digital Instruments MultiMode AFM/STM. In addition to scanning tunneling and contact atomic force microscopy (AFM/STM) the instrument features several other operation modes, including non-contact AFM, TappingMode™ AFM, force modulation and magnetic force modes. Also, the system is equipped with a Hysitron Inc. nanoindentation device capable of sample imaging and measurement of material properties such as hardness and elastic modulus at the nanometer level.

Optical Microscopy
Facilities for optical microscopy include an Olympus BX60 polarizing microscope, color video camera, Sony 27-in monitor, and digital image analysis. A specialized heating and cooling stage is also available for the study of fluid inclusions in minerals or for studies of other high- or low-temperature phase transitions. The temperature range for this stage is from -196°C to +700°C. The microscope is also equipped with a UV fluorescence system for the analysis of hydrocarbon fluid inclusions and studies of petroleum migration.

Analytical Chemistry
The Analytical Chemistry Facility uses atomic absorption spectrophotometry (AA) and inductively coupled argon plasma mass spectrometry (ICP-MS) to provide a wide range of elemental analyses. The laboratory also conducts metal assaying including classical gold fire assay methods, and performs many other types of analysis following standard gravimetric, colorometric, spectrophotometric, and wet-chemical methods.

Other Services Available on Campus
Portable Visible and Near Infrared Spectroradiometer (350-2500 nm)
Petrographic Services
Thin Section Preparation
Concrete Petrography
Fourier-Transform Infrared Spectroscopy
Micro-Raman Spectroscopy
UV-Visible Spectroscopy
Nuclear Magnetic Resonance Spectroscopy (solid and liquid state)
Gas Chromatography
High Pressure Liquid Chromatography
Fiber and Polymer Analysis
Metallographic Analysis
Mechanical Testing Instrument
Hardness Testing
Concrete Testing
Remote Sensing and Image Processing Services
Geographic Information Systems Laboratory