C33B-01 INVITED
An In-Situ Deep-UV Optical Probe for Examining Biochemical Presence in Deep Glaciers and Sub-Glacial Lakes
The quest to study and understand extremophiles has led to many quite different research paths in the past 30 years. One of the more difficult directions has been the study of biochemical material in deep glacial ice and in subglacial lakes. Lake Vostok in Eastern Antarctica has been perhaps the most discussed subglacial lake because of its large size (~14,000 sq km), deep location under >3700 m of overlying ice, and thick sediment bed (~200m). Once the physical conditions of the Lake were assessed, questions immediately arose about the potential existence of biological material - either extinct or possibly extant under conditions of extremely limited energy and nutrients [1-2]. To investigate the biology of Vostok, via in-situ methods, is a major issue that awaits proven techniques that will not contaminate the Lake beyond what may have occurred to date. Lake Ellsworth, in West Antarctica, also discovered by ice penetrating radar, is of significantly smaller size, but is also >3500 m below the overlying ice. It represents a wonderful opportunity to design, engineer and build in-situ delivery systems that consider bio-cleanliness approaches to enable examination of its water, sediment bed and the "roof" area accretion ice for biochemicals [3]. Our laboratory has been developing deep UV fluorescence and UV Raman instrumentation to locate and classify organic material at a variety of extremophile locations. The confluence of the measurement techniques and the engineering for high external pressure instrument shells has enabled us to design and begin prototype fabrication of a biochemical sensing probe that can be inserted into a hot-water drilled ice borehole, functioning as a local area mapper in water environments as deep as 6000 m. Real-time command and control is conducted from a surface science station. We have been using the deep Vostok ice cores at the U.S. National Ice Core Lab to validate our science and data analysis approaches with an "inverted" system that has recently generated spatially resolved spectral images of material inside the Vostok cores without extraction or disturbance to the material in the ice. We will describe the instrumentation we will have available for the British Antarctica Survey Lake Ellsworth Exploration field campaign, provide a possible operational scenario and show examples of the kinds of possible measurement results that might be obtained, based upon our Lake Vostok core studies. [1] Siegert, M.J., Tranter, M., Ellis-Evans, C.J., Priscu, J.C. & Lyons, W.B. (2003) The hydrochemistry of Lake Vostok and the potential for life in Antarctic subglacial lakes. Hydrological Processes, 17, 795-814. [2] Priscu, J.C. and B.C. Christner (2004). Earth's icy biosphere, pp. 130-145, In "Microbial Diversity and Bioprospecting", A. Bull (editor). Chap 13. ASM Press, Washington, D.C. [3] Siegert M.J., Hindmarsh, R., Corr H., Smith, A., Woodward, J., King, E., Payne, A.J., and Joughin, I.(2004) Subglacial Lake Ellsworth: a candidate for in situ exploration in West Antarctica. Geophysical Research Letters, 31 (23), L23403, 10.1029/2004GL021477.
C33B-02
In Situ ATP Bioluminescent Measurements in Subglacial Environments – The Engabreen Glacier in the Norwegian Arctic
Engabreen is a northern outlet glacier from the western Svartisen ice cap on the Nordland coast of Norway just inside the Arctic Circle. A unique feature of the glacier is a man-made tunnel system within the bedrock beneath the glacier that offers scientists direct access to the glacier-bedrock interface. This unique facility - called the Engabreen Subglacial Laboratory - is ideal to test developments of new in situ analytical techniques. We have used the facility to perform the first in situ detection of microbial life in a subglacial environment using standard off-the-shelf ATP bioluminescence detection technology and therefore using ATP levels as a proxy of microbial life. Measurements were performed both in melt-waters in the tunnels and from melted ice samples directly from the glacier-bedrock interface. Levels of ATP above background were detected and appeared to be associated with suspended sediment particles rather than in the water or ice component. This indicated the presence of microbial life. Development of protocols for in situ sample processing and use of in situ ATP measurements in the directing and choice of sampling points for other techniques was explored. This study has shown that off-the-shelf portable ATP bioluminescence can be used to perform in situ measurements within sub-glacial environments but that further development work is required to optimize experimental protocols and to correlate findings with other life detection and enumeration techniques.
C33B-03
Detection of Microbial Life in Glacial Samples – Laboratories Studies and Development for Field use
Adenosine triphosphate (ATP) is frequently used as a proxy for bulk microbial biomass in environmental sciences and, in the food and health industries. Despite successful ATP detection in a variety of ecosystems, very little data are available on ATP levels in the glacial system. In this study, protocols for ATP detection on glacial ice and sediment samples are investigated, in order to aid in the development of a single-use device for in-field life detection, and also to increase the available data on biomass estimates in the cryosphere. ATP detection in two glacial samples reveals concentrations indistinguishable from internal blanks. Therefore, the samples were centrifuged and their particulate loads were subjected to four different extraction processes. Applying these extraction methods resulted in higher ATP concentration than samples with no extraction process; the different techniques increase the ATP detected between 5 and 15 times (also relative to an internal standard). Concurrent with the laboratory based development of extraction protocols is the development of a single-use device for the detection of ATP at the sampling site, in icy environments. The device is microfluidic-based, using commercially available reagents for the detection of ATP by bioluminescence. In order to produce a robust measure of biomass, both laboratory and field based analyses need to be carried out. This work shows the potential of ATP detection in glacial samples and the early development of a device for in situ life detection. The quantification of ATP in microfluidic format is being developed as the preliminary target for an integrated life detection and characterisation device.
C33B-04 INVITED
Ecological Investigations of the Subglacial Biome: Challenges and Potential
More than 80% of the Earth's biosphere is cold (<4°C) and over 10% of the Earth's surface is covered by ice. Cold icy environments are now known to support a surprising diversity of active microbial life. This talk will provide a brief background on the current understanding of life in icy environments and discuss methods being used by the subglacial community for life detection and in situ measurements as well as a "wish list" of desired biogeochemical observations. Despite new discoveries, there are considerable challenges to the exploration of icy environments. Research on subglacial ecosystems offer significant potential for insight into natural systems; they provide a unique "model" to study microbial communities in relative geological isolation. The long time-scale of entrapment relative to average lifetimes of a microbial cell provides an opportunity to explore questions ranging from the linkages between metabolic pathways, thermodynamics and substrate availability; to the potential for insight about fundamental rates of evolution and constraints on biodiversity. The subglacial environment is one of the most difficult portions of the cryosphere to access and only recently, in collaboration with large-scale drilling projects, are ecologists and biogeochemists beginning to explore the more remote reaches of the subglacial biome. The small sample sizes often retrieved from cold systems, which contain low cell densities and low metabolic activity, present significant challenges to measurements of phylogenetic and metabolic diversity and rates of metabolic activity. While the physical nature of most frozen environments makes in situ observations of the interaction between the natural biological assemblage and the surrounding ecosystem difficult at best. There is a growing urgency to understand the role of icy environments in the global system as well as a steady interest in exobiology, thus life detection in cold environments will continue to be an important scientific and engineering challenge with the potential for high reward.
C33B-05
Spaceflight Engineering For The Remote Study Of The Terrestrial Cryosphere: The CryoEgg Platform
We describe a remote sensing platform for deployment at considerable depth within the cryosphere, targeted particularly at glacial and polar ice environments. The design exploits our team's heritage in space instrumentation and engineering to meet the requirements of autonomous operation under conditions of extreme pressure and temperature, in a compact and rugged package. The modular design permits reconfiguration of the device to meet the specific demands and scientific requirements of each deployment site. The platform is a core element of the CryoEgg programme, the aims of which are to produce an autonomous sensor system capable of monitoring basic chemical/biological and physical variables over the full spectrum of conditions in subglacial environments, and we describe the performance of the platform in verification tests and preliminary field trials at the Engabreen glacial laboratory in Norway.
C33B-06
A Practical Method to Autonomously Obtain Vertical Profiles of Biomarkers in Ice Sheets on Earth and Mars
We describe experiments using a low-power thermal drill capable of autonomously retrieving and analyzing meltwater samples from an ice sheet with a vertical resolution of a centimeter or less. The drill operates by passively creating a melt front at the nose, then pumping the melt water to an analytical instrument on the surface. This "open hole" strategy minimizes thermal contact between the drill and the ice, limiting the power consumption to little more than is necessary to melt the ice. Even in the cold martian ice sheet (<175K) a 7.5 cm diameter drill can descend at speeds of 25 cm per hr using only about 250W. In relatively warm terrestrial ice the technique is useful only to a few hundred meters, below which the hole would collapse. The drill has successfully been tested in Greenland to a depth of 50 m. In cold martian ice, given enough time, such a drill could penetrate two or more kilometers to the base of the northern ice sheet. For Mars, the primary objective of such a drill is to explore the climate record through visual inspection of layering and analysis of isotopic ratios in the meltwater. A secondary objective would be to seek biomarkers through detection of relevant chemical signatures such as methane or fluorescent molecules. To this end, studies were performed to determine the limits of vertical resolution imposed by tube flow of the meltwater from the drill to the surface. Mixing in the tube could dilute a localized biomarker to the point of undetectability, or could degrade the ability to associate such a signal with a specific historical climate marker. Chemical markers (salts, detected by conductivity changes) and fluorescent markers (including quantum dots) were introduced abruptly into the meltwater stream, and the effluent at the end of the tube was then analyzed to determine the persistence of the signature. Theoretically, the transfer function is strongly dependent on factors that are difficult to quantify such as turbulence and wall interactions. In the configuration studied, the detected signal was significantly more abrupt than might have been expected from a purely laminar tube flow model.
C33B-07
Fluorescence Spectroscopy as a Rapid, High-Resolution Tool for Detecting Biomolecules in Glacial Ice
We have developed new instruments utilizing the intrinsic fluorescence of specific biomolecules as a sensitive, non-destructive tool for detecting microorganisms. Using a 224-nm excitation, we detect protein-bound tryptophan (an amino acid present in all cells) at a detection threshold of approximately 1 cell per laser excitation volume and a duty cycle of 100 ms per measurement. Tryptophan is easily distinguished from inorganic backgrounds due to its characteristic spectral shape and ~300 times higher intensity per unit volume than typical inorganic compounds. A different excitation was also used to detect coenzyme F420, a characteristic marker for viable methanogenic cells. At the National Ice Core Laboratory, systematic scans of a 1 meter core sections took about 15 minutes and generated ~5000 measurements per meter. The high-resolution of this work revealed strong variability of microbial content on a scale of cm within individual cores, which suggests that microbial deposition at polar sites is strongly influenced by meteorological events (e.g. storms) on subannual and interannual scales. In addition, high levels of microbes are found to correlate with anomalously high concentrations of metabolic gases (e.g. methane, nitrous oxide, and 18O/16O of O2), suggesting that many of the isolated "gas artifacts" identified in deep ice cores are the accumulated waste products of in situ metabolism. This means that fluorescence spectroscopy may be a useful tool for identifying regions where high microbial concentrations have contaminated gas records. The existing instrumentation is suitcase portable and could be easily deployed in a variety of environments. Future versions of these instruments may be practical for continuous, rapid scans of entire cores, as an on-site deployable technique for characterizing microbial abundances in ice, and for searching for as few as 1 microbe per cm3 in ice-bound planets. This work was supported by NSF grant ANT-0440609.
C33B-08
Searches for Microbial Cells with Fluorescence Loggers with Single-cell Sensitivity
Two known habitats for microbial metabolism in ice are surfaces of mineral grains and liquid veins along three- grain boundaries. Several problems suggest the need for a third habitat: veins usually contain toxic liquid; some microorganisms are too large to fit into a vein; veins may not be present at all depths; and the oxygen concentration in veins does not permit the coexistence of both strict anaerobes and aerobes in the same region. We show that a more general habitat avoids these problems. Isolated microbes frozen in ice and not in contact with a vein or grain can metabolize by redox reactions with dissolved small molecules diffusing through the ice lattice. The two requirements are that the gaseous reactants have sufficiently high equilibrium concentrations and diffusion coefficients to provide enough metabolic energy to repair macromolecular damage as it occurs. Molecules with less than ~6 atoms (e.g., H2, O2, N2¬, CO, CO2, CH4, H2S, NH3, HNO3, HCHO, and HCOOH) have values of diffusion coefficient D(T) that exceed ~10- 15 m2 s-1, which is sufficient to sustain microbial life in ice. For terrestrial environments, we show that there is an adequate supply of such molecules diffusing throughout deep glacial ice to sustain metabolism for millions of years. Our recent noninvasive observations of ice cores from GISP2 and WAIS Divide provide evidence for this habitat. Using scanning fluorimetry to map proteins (a proxy for cells) and F420 (a proxy for methanogens) in ice cores, we find isolated spikes of fluorescence consistent with as few as one microbial cell in a volume 0.16 microliter with the protein mapper and in 1.9 microliter with the methanogen mapper. With such precise localization one could use a nanomanipulator to extract single cells for molecular identification. Low- power, miniaturized versions of these instruments could search for single cells in subglacial lakes, Martian ice- rich permafrost, and Europan ice.