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A suite of lunar soils has been investigated by imaging and in-situ spectroscopy techniques. A suite of contaminant plastics and potential microbes has been found. Additional information is contained in the original extended abstract.
Microbial fossilization processes in the Dead Sea is primarily associated with the calcium cation. The putative fossilized microbes do not represent the reported living microbial population. Additional information is contained in the original extended abstract.
Mn-oxides are promising biomarkers because microbes greatly accelerate Mn-oxide formation rates and produce distinctive oxidation states. Mn minerals in terrestrial caves form subaerially and could conceivably be present in the subsurface of Mars. Additional information is contained in the original extended abstract.
A potential challenge for a human Mars mission is that while humans are by most measures the obvious best way to search for life on Mars, we may also be the most problematic in that we could unduly compromise the search for life by contaminating relevant environments and/or possibly adversely and irreversibly affecting indigenous life. Perhaps more problematic is the fundamental epistemic challenge of the "one data point" limitation which could decrease confidence in applying terrestrially based research to extraterrestrial life issues in general. An informal decision tree is presented as one way to begin thinking about contamination issues. There are many sub-questions and distinctions not shown such as biological vs. nonbiological (but biologically relevant) contamination, viable vs. dead organisms, masking indigenous organisms vs. merely making the search more difficult, and independent origin vs. panspermia distinctions. While it may be unlikely that terrestrial microbes could survive on Mars, let alone reproduce and unduly compromise the search for life, the unpredictable potential for microbial life to survive, grow exponentially, evolve and modify (and sometimes destroy) environments, warrants focusing carefully on biologically relevant contamination as we prepare to send humans to the first planet that may have indigenous life-forms.
Recently, strong evidence of liquid water under the surface of Mars and a meteorite that might contain ancient microbes have renewed interest in Mars exploration. With this renewed interest, NASA plans to send spacecraft to Mars approx. every 26 months. These future spacecraft will return higher-resolution images, make precision landings, engage in longer-ranging surface maneuvers, and even return Martian soil and rock samples to Earth. Future robotic missions and any human missions to Mars will require precise entries to ensure safe landings near science objective and pre-employed assets. Potential sources of water and other interesting geographic features are often located near hazards, such as within craters or along canyon walls. In order for more accurate landings to be made, spacecraft entering the Martian atmosphere need to use lift to actively control the entry. This active guidance results in much smaller landing footprints. Planning for these missions will depend heavily on Monte Carlo analysis. Monte Carlo trajectory simulations have been used with a high degree of success in recent planetary exploration missions. These analyses ascertain the impact of off-nominal conditions during a flight and account for uncertainty. Uncertainties generally stem from limitations in manufacturing tolerances, measurement capabilities, analysis accuracies, and environmental unknowns. Thousands of off-nominal trajectories are simulated by randomly dispersing uncertainty variables and collecting statistics on forecast variables. The dependability of Monte Carlo forecasts, however, is limited by the accuracy and completeness of the assumed uncertainties. This is because Monte Carlo analysis is a forward driven problem; beginning with the input uncertainties and proceeding to the forecasts outputs. It lacks a mechanism to affect or alter the uncertainties based on the forecast results. If the results are unacceptable, the current practice is to use an iterative, trial-and-error approach to reconcile discrepancies. Therefore, an improvement to the Monte Carlo analysis is needed that will allow the problem to be worked in reverse. In this way, the largest allowable dispersions that achieve the required mission objectives can be determined quantitatively.
Storr's Lake, an inland hypersaline lake on San Salvador Island, Bahamas, contains calcium carbonate-rich lithified mats of filamentous microorganisms, diatoms, associated photosynthetic and chemotrophic bacteria, and trapped sediment. In addition, 16S rRNA analysis indicates the presence of five sulfur-reducing genera of bacteria. These microbes are potential modern-day analogs to some ancient stromatolitic structures. The goals of this study are to identify unique compositional and biogenic features, possibly correlating some of these with some of the sulfate-reducing bacteria. Additional information is contained in the original extended abstract.
The direct detection of organic biomarkers for living or fossil microbes on Mars by an in situ instrument is a worthy goal for future lander missions. Several new and innovative biotechnology approaches are being explored. Firstly we have proposed an instrument based on immunological reactions to specific antibodies to cause activation of fluorescent stains. Antibodies are raised or acquired to a variety of general and specific substances that might be in Mars soil. These antibodies are then combined with various fluorescent stains and applied to micron sized numbered spots on a small (2-3 cm) test plate where they become firmly attached after freeze drying. Using technology that has been developed for gene mining in DNA technology up to 10,000 tests per square inch can now be applied to a test plate. On Mars or the planet/moon of interest, a sample of soil from a trench or drill core is extracted with water and/or an organic solvent and ultrasonication and then applied to the test plate. Any substance, which has an antibody on the test plate, will react with its antibody and activate its fluorescent stain. At the moment a small UV light source will illuminate the test plate, which is observed with a small CCD camera, although other detection systems will be applied. The numbered spots that fluoresce indicate the presence of the tested-for substance, and the intensity indicates relative amounts. Furthermore with up to a thousand test plates available false positives and several variations of antibody can also be screened for. The entire instrument can be quite small and light, on the order of 10 cm in each dimension. A possible choice for light source may be small UV lasers at several wavelengths. Some of the wells or spots can contain simply standard fluorescent stains used to detect live cells, dead cells, DNA, etc. The stains in these spots may be directly activated, with no antibodies being necessary. The proposed system will look for three classes of biomarkers: those from extant life, such as DNA, those from extinct life such as hopanes, and those from organic compounds not necessarily associated with life such as PAHs, rocket exhaust contamination and other a/pre-biotic chemicals. Both monoclonal and polyclonal antibodies can be used. Monoclonal antibodies react with a very specific compound, but polyclonal antibodies may react to any of a whole family of compounds. Furthermore the technique of phage display to raise antibodies against classically non-antigenic molecules is also being considered. Additional information is contained in the original extended abstract.
Perhaps one of the most fundamentally difficult challenges facing those who would search for life is that of scale determination. Spatial scales of life on Earth range over more than 15 orders of magnitude in mass and volume, and more than 8 orders of magnitude in 2 dimensional space. If the distribution of life is sparse in comparison to the background on which it is found, then the choice of the right scale is critical to finding that life. But how does one identify the proper scale? To put this in other words, how does one recognize the "haystacks" in which the needles (biosignatures and evidence of life) might be most profitably searched for? The problem is further exacerbated when conditions get extreme because much of the life moves from the clement surface environment into the pores and more clement environments inside of rocks, minerals and soils. Once encased in their lithic homes, these microbes become nearly impossible to study by standard techniques because of the opacity of the rocks. It is this problem that we propose to address in the work proposed here. Computer Tomography (CT) has been a very valuable tool in medicine, where the best resolution available has typically been of the order of about 0.5 mm. However, to adapt the approach for life detection of microbial endoliths, the resolution needs to be moved to the micrometer and even submicrometer levels. Thus for the studies proposed here, we begin with a commercially available instrument that can yield resolution of approximately 10 micrometers. The rational for this is twofold: first, this is the "state of the art" in laboratory instruments; and second, that while the usual size of a microbial cell is about 1 micron, microorganisms tend to live in communities that usually exceed the 10 micrometer size range. The resolution also depends on the sample size itself, so having a small lab instrument into which small samples can be placed will be beneficial to the resolution. We have now used several different CT systems, beginning with the medical scanners (Arcadia CT group) for the detection of layered communities in sandstone rocks from Antarctica. Even this crude instrument was able to point to the areas of the rock that were dominated by microbial populations - this provides the critical first information that says, "Go back and look at these sites with other methods." We showed that without sample preparation or destruction it was possible to gain knowledge as to the presence of density differences suggestive of life.
Recently, evidence has begun to grow supporting the possibility that the Viking GC-MS would not have detected certain carboxylate salts that could have been present as metastable oxidation products of high molecular weight organic species. Additionally, despite the instrument's high sensitivity, the possibility had remained that very low levels of organic matter, below the instrument's detection limit, could have been present. In fact, a recent study indicates that the degradation products of several million microorganisms per gram of soil on Mars would not have been detected by the Viking GC-MS. Since the strength of the GC-MS findings was considered enough to dismiss the biology packet, particularly the LR results, any subsequent evidence suggesting that organic molecules may in fact be present on the Martian surface necessitates a re-evaluation of the Viking LR data. In addition to an advanced mass spectrometer to look for isotopic signatures of biogenic processes, future lander missions will include the ability to detect methane produced by methanogenic bacteria, as well as techniques based on biotechnology. Meanwhile, the identification of Mars samples already present on Earth in the form of the SNC meteorites has provided us with the ability to study samples of the Martian upper crust a decade or more in advance of any planned sample return missions. While contamination issues are of serious concern, the presence of indigenous organic matter in the form of polycyclic aromatic hydrocarbons has been detected in the Martian meteorites ALH84001 and Nakhla, while there is circumstantial evidence for carbonaceous material in Chassigny. The radiochronological ages of these meteorites are 4.5 Ga, 1.3 Ga, and 165 Ma respectively representing a span of time in Earth history from the earliest single-celled organisms to the present day. Given this perspective on organic material, a biological interpretation to the Viking LR results can no longer be ruled out. In the LR experiment, a solution containing C-14 labeled organic compounds was injected into soil samples. The detection of radioactivity in the overhead space would indicate that one or more of the substrates had been chemically converted into a carbon-containing gas. To serve as a control, some samples were heated enough to destroy most known terrestrial microbes so that an indication for life would be a positive response from unheated samples and a negative response from heated samples. On Mars, the LR results had met minimum criteria for a biological interpretation but due to the GC-MS results, the LR responses were later attributed to putative soil inorganic oxidants. Since the time of Viking, studies have been carried out with the objective of determining an oxidant or combination of oxidants that might exist on Mars and have produced the observed kinetics of the LR response. To date, no such agent has been found that produces all aspects of the LR results on Mars. While the above considerations in no way imply the existence of life forms at the two Viking landing sites, inorganic and biological explanations for the Viking LR data should now be considered equally plausible until more complete studies of the Martian surface are carried out. Therefore, in light of the SNC meteorites data and their implications for the possibility of organic matter near or on the Martian surface the Viking biology experiments should thus be seen, not as failures for their inability to provide unambiguous evidence for or against Martian life, but as a foundation for the development of future life-detection instruments. Additional information is contained in the original extended abstract.
Microorganisms preserved within the permafrost, glaciers, and polar ice sheets of planet Earth provide analogs for microbial life forms that may be encountered in ice or permafrost of Mars, Europa, Callisto, Ganymede, asteroids, comets or other frozen worlds in the Cosmos. The psychrophilic and psychrotolerant microbes of the terrestrial cryosphere help establish the thermal and temporal limitations of life on Earth and provide clues to where and how we should search for evidence of life elsewhere in the Universe. For this reason, the cold-loving microorganisms are directly relevant to Astrobiology. Cryo-preserved microorganisms can remain viable (in deep anabiosis) in permafrost and ice for millions of years. Permafrost, ice wedges, pingos, glaciers, and polar ice sheets may contain intact ancient DNA, lipids, enzymes, proteins, genes, and even frozen and yet viable ancient microbiota. Some microorganisms carry out metabolic processes in water films and brine, acidic, or alkaline channels in permafrost or ice at temperatures far below 0 T. Complex microbial communities live in snow, ice-bubbles, cryoconite holes on glaciers and ancient microbial ecosystems are cryopreserved within the permafrost, glaciers, and polar caps. In the Astrobiology group of the NASA Marshall Space Flight Center and the University of Alabama at Huntsville, we have employed advanced techniques for the isolation, culture, and phylogenetic analysis of many types of microbial extremophiles. We have also used the Environmental Scanning Electron Microscope to study the morphology, ultra-microstructure and chemical composition of microorganisms in ancient permafrost and ice. We discuss several interesting and novel anaerobic microorganisms that we have isolated and cultured from the Pleistocene ice of the Fox Tunnel of Alaska, guano of the Magellanic Penguin, deep sea sediments from the vicinity of the Rainbow Hydrothermal Vent and enrichment cultures from ice of the Patriot Hills of Antarctica. The microbial extremophiles recovered from permafrost, ice, cold pools and deep sea sediments may provide information relevant to the question of how and where we should search for evidence of extant or extinct microbial life elsewhere in the Cosmos.
Terrestrial life appears to have arisen very quickly during late accretion, sometime between approximately 3.5 and 4.2 Ga. During this same time, liquid water appears to have been abundant at the surface of Mars and it is quite plausable that life originated there as well. We now believe that the last common ancestor of terrestrial life was a sulfur-metabolizing microbe that lived at high temperatures. Rooting of the RNA tree in thermophily probably reflects high temperature "bottle-necking" of the biosphere by giant impacts during late accretion, sometime after life had originated. If high temperature bottle-necking is a general property of early biosphere development, Martian life may have also developed in close association with hydrothermal systems. Several independent lines of evidence suggest that hydrothermal processes have played an important role during the geological history of Mars. Because hydrothermal deposits on Earth are known to capture and retain abundant microbial fossil information, they are considered prime targets in the search for an ancient Martian biosphere. An important step in planning for future landed missions to Mars is the selection of priority targets for high resolution orbital mapping. Geotectonic terranes on Mars that provide a present focus for ongoing site selection studies include channels located along the margins of impact crater melt sheets, or on the slopes of ancient Martian volcanoes, chaotic and fretted terranes where shallow subsurface heat sources are thought to have interacted with ground ice, and the floors of calderas and rifted basins. Orbital missions in 1996, 1998 and 2001 will provide opportunities for high resolution geological mapping at key sites in such terranes, as a basis for selecting targets for future landed missions for exopaleontology.
New spectral measurements of bacteria taken at cryogenic temperatures provide insights on the surface composition of Europa as well as the detectability of microbes on the surface. Additional information is contained in the original extended abstract.
Microorganisms preserved within the permafrost, glaciers, and polar ice sheets of planet Earth provide analogs for microbial life forms that may be encountered in ice or permafrost of Mars, Europa, Callisto, Ganymede, asteroids, comets or other frozen worlds in the Cosmos. The psychrophilic and psychrotolerant microbes of the terrestrial cryosphere help establish the thermal and temporal limitations of life on Earth and provide clues to where and how we should search for evidence of life elsewhere in the Universe. For this reason, the cold-loving microorganisms are directly relevant to Astrobiology. Cryopreserved microorganisms can remain viable (in deep anabiosis) in permafrost and ice for millions of years. Permafrost, ice wedges, pingos, glaciers, and polar ice sheets may contain intact ancient DNA, lipids, enzymes, proteins, genes, and even frozen and yet viable ancient microbiota. Some microorganisms carry out metabolic processes in water films and brine, acidic, or alkaline channels in permafrost or ice at temperatures far below 0 C. Complex microbial communities live in snow, ice-bubbles, cryoconite holes on glaciers and ancient microbial ecosystems are cryopreserved within the permafrost, glaciers, and polar caps. In the Astrobiology group of the NASA Marshall Space Flight Center and the University of Alabama at Huntsville, we have employed advanced techniques for the isolation, culture, and phylogenetic analysis of many types of microbial extremophiles. We have also used the Environmental Scanning Electron Microscope to study the morphology, ultra-microstructure and chemical composition of microorganisms in ancient permafrost and ice. We discuss several interesting and novel anaerobic microorganisms that we have isolated and cultured from the Pleistocene ice of the Fox Tunnel of Alaska, guano of the Magellanic Penguin, deep-sea sediments from the vicinity of the Rainbow Hydrothermal Vent and enrichment cultures from ice of the Patriot Hills of Antarctica. The microbial extremophiles recovered from permafrost, ice, cold pools and deep-sea sediments may provide information relevant to the question of how and where we should search for evidence of extant or extinct microbial life elsewhere in the Cosmos.
Most naturally occurring microbes live in complex microbial communities consisting of thousands of phylotypes of microorganisms living in close proximity. Each of these draws nutrients from the environment and releases metabolic waste products, which may in turn serve as substrates for other microbial groups. Gross environmental changes, such as irradiance level, hydrodynamic flow regime, temperature or water chemistry can directly affect the productivity of some community members, which in turn will affect other dependent microbial populations and rate processes. As a first step towards the development of "standard" natural communities of microorganisms for a variety of potential NASA applications, we are measuring biogeochemical cycling in artificially structured communities of microorganisms, created using natural microbial mat communities as inoculum. The responses of these artificially assembled communities of microorganisms to controlled shifts in ecosystem incubation conditions is being determined. This research requires close linking of environmental monitoring, with community composition in a closed and controlled incubation setting. We are developing new incubation chamber designs to allow for this integrated approach to examine the interplay between environmental conditions, microbial community composition and biogeochemical processes.
Geomicrobiological phenomena are among the most fundamental of interactions between Earth and its biosphere. Actively growing and lithifying stromatolites at Highborne Cay Bahamas, have recently been documented and allow for detailed examination of the roles microbes play in the mineralization process. These stromatolites contain a variety of complex microbial communities with distinct distribution patterns for different microbial groups. Cyanobacteria are the primary producers in this system providing energy, directly or indirectly, for the entire stromatolite microbial community. They also play key roles in the trapping and binding of sediments. Most of these species are highly motile and can adjust their position and orientation within the sediment matrix in order to optimize their access to irradiance and nutrients. As individual species have different physical and metabolic properties, this motility generally results in segregated distributions of species, which in turn contributes to the laminated textures observed in these actively forming stromatolites. Increasingly our studies suggest that the activities and locations of various cyanobacterial species also contribute greatly to the localization of new mineral precipitation through a variety of processes. We are investigating these contributions using an integrated approach combining detailed observations of field samples with manipulative experiments using both field samples and cultures of specific organisms isolated from these stromatolites. Experiments are conducted both in standard laboratory conditions and in outdoor running seawater flumes. A variety of standard techniques; SEM (scanning electron microscopy), petrographic analyses, TEM (transmission electron microscopy), are used to compare mineralization processes in field samples with those generated in laboratory-flume simulations. Using this approach we are able to more thoroughly investigate the effects of irradiance, CaCO3 saturation, and hydrodynamic regime on cyanobacterial distribution, trapping and binding and mineral precipitation. Simulation results will be presented and compared with community and mineralization distribution patterns seen in the field samples from which these communities were isolated.
The effects of cold plasma exposure on Deinococcus radiodurans, plasmid DNA and model proteins were assessed using microbiological, spectrometric and biochemical techniques. Exposure of D. radiodurans, an extremely radiation resistant microbe, to O2 plasma (less than or equal to 25 W, approx. 45 mTorr, 90 min) yielded a approx. 99.999 % sterilization and the sterilization rate was increased approx. 10-fold at 100 W and 500 mTorr. AFM images shows that the exposed cells are significantly deformed and possess 50-70 nm concavities. IR analysis indicates the chemical degradation of lipids, proteins and carotenoids of the cell wall and membrane. Intracellular damage was indicated by major absorbance loss at 1245, 1651 and 1538/cm corresponding to degradation of DNA and proteins, respectively. Biochemical experiments demonstrate that plasmas induce strand scissions and crosslinking of plasmid DNA, and reduction of enzyme activity; the degradation is power dependent with total sample loss occurring in 60 s at 200 W and 500 mTorr. Emission spectroscopy shows that D. radiodurans is volatilized into CO2, CO, N2 and H2O confirming the removal of biological matter from contaminated surfaces. The O2 plasma impacts several cellular components predominantly through chemical degradation by atomic oxygen. A CO2, plasma, however, was not effective at degrading D. radiodurans, revealing the importance of plasma composition, which has implications for planetary protection and the contamination of Mars.
Tholin is a word coined to describe the entire class of complex organic solids produced in laboratory experiments where pre-biotic gaseous chemicals are subject to bombardment by high energy. The atomic composition of Titan tholin produced from 10 percent CH4 and 90 percent N2 in a simulation of Titan atmosphere irradiated by charged particles trapped in the magnetosphere of Saturn gave 67 percent C and 33 percent N. Hydrolysis of Titan tholin with 6N HCl produced a racemic mixture of biological and non-biological amino acids that was confirmed by GC/MS. Other tholins, that revealed the presence of amino acids, were UV tholin produced under possible primitive Earth conditions by irradiation of a mixture of gases (CH4, C2H6, NH3, H2S and liquid H2O) with long-wavelength ultraviolet light, representing the most abundant useful energy source for prebiological organic synthesis; Spark tholin in a crude simulation of Jupiter atmosphere using electrical discharge through a mixture of CH4, NH3, and H2O vapor. Pyrolytic GC/MS of Titan tholin produced more than one hundred organic compounds including saturated and unsaturated aliphatic hydrocarbons, substituted polycyclic aromatics, nitriles, amines, pyrroles, pyrazines, pyridines, pyrimidines, and the purine, adenine. Similar rich pyrolytic products were obtained with UV as well as Spark tholins. A range of two to four ring PAHs (Polycyclic Aromatic Hydrocarbons) in Spark as well as Titan tholins, some with one to four alkylation sites, were identified by two-step laser desorption/multiphoton ionization mass spectrometry and also confirmed by the synchronous fluorescence technique. Previous studies have demonstrated the potential for use of tholins as a source of carbon and energy by microbes. This paper describes studies that evaluate the potential for using different types of tholins as (a) a substrate for growing plants and (b) a source of carbon and nitrogen for plants. The data are interpreted in terms of the potential for using such tholins to grow plants for food in extraterrestrial habitats and also to speculate on the possibilities of abiotic evolution of plants.
This lecture will introduce the concept of biogeochemical cycling. The roles of microbes in the cycling of nutrients, production and consumption of trace gases, and mineralization will be briefly introduced.