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Project Reports

• Project 1A: Ground Control Experiments for the OREOcube Mission

  The OREOcube (ORganics Exposure in Orbit cube) experiment has been designed to study the effects of solar and cosmic radiation on astrobiology relevant organics when associated with mineral surfaces. The goal of this project is to investigate the (photo)chemical evolution and processing of organic matter in simulated proto-planetary and planetary microenvironments in the laboratory and in space. Testing the photostabitity and evolution of organics in actual space environments on the International Space Station (ISS) provides a powerful tool to study the combined impact of several relevant parameters (e.g., ultraviolet and comic radiation, microgravity) simultaneously. The OREOcube capability to measure in-situ changes of samples exposed to space radiation using an UV-visible-NIR spectrometer as a function of time in Low Earth Orbit will confer significant advantages relative to more basic ISS exposure facilities for which sample measurements are made on Earth prior to and at the end of a space mission. The innovative aspect of this OREOcube experiments is the capability of in-situ monitoring of flight samples, which has not yet been achieved on the ISS. A comparison of measurements performed with OREOcube with recent LEO data (EXPOSE-E, EXPOSE-R, O/OREOS Sat) and ground-based laboratory data will reveal how those combined data from different exposure facilities can be effectively used to investigate the evolution of organic matter in space.

  ROADMAP OBJECTIVES: 3.1

• Project 1B: Recovery of Mineral-Associated Amino Acids

  As the single most abundant molecules in a cell (apart from water), amino acids are among the most widespread biomolecules on Earth. Amino acids are the building blocks of proteins and are also used for countless other functions, such as signal transduction, in all domains of life. Of special interest are the α-amino acids that have both the amine and carboxylic acid group attached to the first carbon (α) atom and have an organic substituent as the functional side chain. These amino acids include the 23 proteomic amino acids which are linked together by protein translating machinery into peptides resulting in the formation of proteins. Many amino acids display isomerism resulting in the appearance of L- and D- amino acids. Amino acids have been detected in numerous extreme environments as well as in extraterrestrial meteorites. But they are fragile and can be rapidly degraded when exposed to UV radiation and other oxidizing conditions. The differences in side chains between the amino acids have an effect on their charge and polarity. The resulting differences between amino acids makes their interaction with minerals variable. This may result in different extraction rates when recovering them from soils. On the other hand, the structural molecular resemblance of amino acids makes them hard to separate and detect individually by techniques such as HPLC or LCMS. Derivatization methods have been produced to optimize separation and thus detection. Due to their high occurrence on Earth (and in specific space environments) and their role in biological processes, amino acids are good molecular biomarkers. The aim of this study was to determine the extraction efficiency of amino acids from a range of minerals that are found on Mars.

  ROADMAP OBJECTIVES: None Selected

• Project 1C: Analysis of Dissimilatory Iron-Reducing Microbial Communities in Chocolate Pots Hot Spring, Yellowstone National Park

  This study represents the first targeted exploration of the active microbial community at source vent of Chocolate Pots hot springs (CP), a warm, circumneutral pH hot spring in Yellowstone National Park. This work was motivated by previous in vitro dissimilatory iron reducing (DIR) incubations of the native microbial community present in the Fe(III) oxide deposits (hereafter referred to as “CP oxides”) near the vent. DIR has the potential to generate distinct signatures of microbial Fe redox metabolism, and identification of the microbial assemblages involved in this metabolism is important for making a concrete linkage between biological metabolism and the generation of geochemical and isotopic biosignatures in relation to redox gradients on Earth and other rocky planets. The central goal of this study was to obtain a phylogenetic and metagenomic characterization of the active acetate-oxidizing DIR community at CP using ¹³C stable isotope probing (SIP) techniques. CP oxide sediments and spring water were collected from the CP vent source and used to initiate in vitro SIP incubations using labeled (¹³C) and unlabeled acetate. Incubations targeted the active microbial community which is capable of coupling the oxidation of acetate to DIR. The SIP results allowed us to clearly separate the active acetate-metabolizing microbial community from the rest of the community and identify which organisms native to CP make up this population. The role of some members of this community can be inferred with reasonable confidence from the phylogeny of the OTUs from the amplicon libraries (e.g. Geobacter, Ignavibacteria), and the design of the incubations. The metabolic role of other dominant taxa is less well understood at this point and we are preparing to submit samples for shotgun metagenomic sequencing to address these questions.

  ROADMAP OBJECTIVES: 2.1 4.1 5.1 5.3

• Project 1D: Comparative Genomic Analysis of Chemolithotrophic Fe(II)-Oxidizing Bacteria

  A comparative genomic analysis was performed to identify candidate genes involved in extracellular electron transfer (EET) by Fe(II)-oxidizing bacteria (FeOB). The analysis included a variety of publically-available FeOB genomes, together with genomes from FeOB isolated from subsurface sediments, previously-isolated marine basalt-associated FeOB, and metagenomes from chemolithoautotrophic aerobic pyrite-oxidizing and nitrate-reducing Fe(II)-oxidizing enrichment cultures. We identified outer membrane multi-copper oxidase (MCO) genes homologous to proteins known to be involved in EET in several of the FeOB genomes, as well as homologs to the outer membrane c-type cytochrome (ctyc) Cyc2 known to be involved in bacterial Fe(II) oxidation by Acidithiobacillus ferrooxidans under acidic conditions. Further, we found gene clusters that may potentially encode novel “porin-cytochrome-c protein complex” (PCC) in the well-known neutral-pH FeOB S. lithotrophicus ES-1, and homologous operons were found in other recognized FeOB (Leptothrix cholodnii SP-6 and Leptothrix ochracea L12. Another gene cluster consisting of a porin and three periplasmic multiheme cytc was identified in Hyphomicrobium sp. genome retrieved from a pyrite-oxidizing enrichment culture, and its homologous gene clusters are also present in five marine Zetaproteobacterial FeOB genomes. Overall, this analysis, which is based on our current understanding of bacterial EET in Fe redox reactions, provides a list of candidate genes for further experimental and genomic studies.

  ROADMAP OBJECTIVES: 2.1 3.2 4.1 5.1 5.3

• Project 1E: Studies of Early-Evolved Enzymes in Modern Organisms May Reveal the History of Earth’s Ambient Temperature Over Geological Time

  By addressing a focused question — “Does the thermal stability of the reconstructed ancient enzymes of modern organisms provide evidence of the temperature of the environment in which the enzymes originated?” — this study asks a much broader question, namely, “can the biochemistry of extant life provide evidence of ancient environments?” In the geological record, there is virtually no mineralogical evidence to determine ambient surface temperature and data from other sources are ambiguous, contradictory and contentious. By analyzing the thermal stability of ancient reconstructed ancient enzymes, this work may pave the way to solve this fundamental problem and, by doing so, demonstrate a new way to understand the co-evolution of life and its planetary environment.

  ROADMAP OBJECTIVES: 4.1 6.1

• Project 1F: Study of Modern Fe-Mn Nodules in Green Bay Sediments as Analog to the Fe-Nodules and Fe-Oxyhydroxide Minerals on Mars

  Fe-oxide nodules and concretions are common in terrestrial sedimentary rocks and also occur in Martial sediments. The precursors of the hematite are nano-phases of Fe-oxyhydroxides. Modern Fe-Mn nodules from Green Bay sediments were investigated by in-situ XRD, scanning electron microscopy (SEM), high-resolution transmission electron microscopy (HRTEM), Z-contrast imaging, and ab-initio calculations using the density functional theory (DFT) method. Nano-phase minerals for hosting trace elements of As, P, Ba, Co, Ni, and Zn have been identified. Structural sites of the trace elements and their incorporation mechanisms are also proposed. The Fe-Mn nodules can be used as analog for understanding the Fe-nodules and Fe-oxyhydroxide minerals on Mars.

  ROADMAP OBJECTIVES: 2.1 7.1

• Project 2A: Application of the 13C-18O Clumped Isotope Thermometer to the Ancient Earth

  Application of the clumped isotope thermometer in carbonates, which is based on preferential bonding between ¹³C and ¹⁸O, offers the possibility of testing controversial proposals based on conventional stable isotope thermometry that the early Earth’s oceans were hot. In this project, we report on the results of a study of clumped isotope thermometry of the Neoarchean Campbellrand carbonate platform of South Africa. This platform is the best preserved Archean carbonate sequence known and was subjected to only very low grades of metamorphism, and hence offers the best opportunity to determine seawater temperature for the late Archean oceans. Comparison of clumped isotope temperatures retrieved from co-existing calcite and dolomite confirm that resetting of clumped isotope temperatures occurs at different rates for these minerals, where dolomite is closest to preserving primary temperatures. Despite slower re-equilibration rates for dolomite, however, the fact that the Campbellrand platform was buried at temperatures up to 170 oC for ~2 b.y., prevents use of clumped isotope thermometry from preserving Archean seawater temperatures. This would likely not be the case for carbonates on Mars, where early Mars carbonates would have had a relatively low temperature thermal history, suggesting that clumped isotope thermometry for Mars carbonates is a promising approach for determining ancient surface temperatures on Mars.

  ROADMAP OBJECTIVES: 4.1 5.2 7.1

• Project 2B: Cation Content of Carbonate Minerals as a pCO2 Proxy: Inorganic Synthesis Experiments

  Atmospheric CO₂ content and temperature control reaction mechanisms and precipitation kinetics for carbonate minerals. As such, these parameters may indirectly influence the geochemistry and isotope composition of carbonate minerals. To better understand the significance of these geochemical fingerprints, a suite of laboratory experiments is being conducted to characterize the effect of PCO₂ (<1 to >90% CO₂) and aqueous ratios and low mol% MgCO₃ (<3). With the completion of upcoming Mg-isotope analyses and the establishment of a unified consensus on Mg isotope fractionation relations in calcite, Mg isotope systematics can be used to better understand the geochemical and environmental information archived in the rock record.

  ROADMAP OBJECTIVES: 7.1

• Project 2C: Investigation of the Role of Polysaccharide in the Dolomite Growth at Low Temperature by Using Atomistic Simulations

  Polysaccharides in microbial EPS can promote dolomite growth at room temperature. Our molecular dynamics modeling results show that adsorbed polysaccharides can lower activation energy for removing surface water molecules next to the polysaccharides and catalyze dolomite crystallization at low temperature. The process can lower the energy barrier by ~ 1 kcal / mole. Low temperature dolomite / sedimentary dolomite is a potential biosignature. The new finding also provides key to solving the “Dolomite Problem” that has puzzled geologists for decades.

  ROADMAP OBJECTIVES: 7.1 7.2

• Project 2D: Carbonate-Associated Sulfate (CAS) as a Tracer of Ancient Microbial Ecosystems

  Our aim is to investigate and understand microbial communities that flourished much earlier in the Earth’s history. We have adapted a method used to investigate the isotopic compositions of ancient oceans, by analyzing rocks formed at those times, but applied it to the pore-waters present in ancient sediments inhabited by in the contemporaneous microbial communities. The isotopic compositions we measure tell us about the extent and progress of microbial metabolic processes. We have applied this method very successfully to 12 million year old sediments. Most recently in order to test and calibrate the approach most fully, we have been examining recent deposits in which we can analyze microbiological communities, the pore-waters in which they live and the rocks forming there.

  ROADMAP OBJECTIVES: 5.2 6.1 7.1

• Project 3A: Apatitic Latest Precambrian and Early Cambrian Fossils Provide Direct Evidence of Concentrations of Environmental Oxygen

  Means are not currently available to asses either quantitatively or semi-quantitatively the concentration of oxygen in Earth’s atmosphere over geological time. Despite this, the environmental availability of O₂ has been repeatedly postulated to be a cause of major changes in Earth’s biota, most particularly at the Precambrian-Cambrian boundary-defining “Cambrian Explosion of Life,” a time in Earth history when large deposits of phosphate-rich apatite were deposited in shallow basins worldwide. This study shows that substitution of Sm+3 in the Ca I and Ca II sites of fossil-permineralizing, -infilling, and -encrusting apatite can differentiate between oxic, dysoxic, an anoxic settings of apatite formation. Further studies are underway to date such apatite and establish its REE-substitution as a quantitative O₂ paleobarometer.

  ROADMAP OBJECTIVES: 4.1 4.2

• Project 3B: Biologically Recycled Continental Iron Is a Major Component in Banded Iron Formations

  Combined Fe- and Nd-isotope signatures suggest that banded iron formations (BIFs) contain a major component of continentally derived iron that was mobilized by microbial iron reduction followed by transport through an iron shuttle to the site of BIF formation in deep basin environments. This Fe source is in addition to the widely accepted submarine hydrothermal source of Fe in BIFs, and the two sources of Fe may be comparable in importance, although their proportions change over time dependent on basin-scale circulation. These results document a vigorous, basin-scale biological cycle for Fe at least 2.5 b.y. ago.

  ROADMAP OBJECTIVES: 4.1 5.2 7.1

• Project 3C: A Redox-Stratified Ocean 3.2 Billion Years Ago

  A novel combination of stable Fe and radiogenic U–Th–Pb isotope data that demonstrate that significant oxygen contents existed in the shallow oceans at 3.2 Ga, based on analysis of the Manzimnyama Banded Iron Formation (BIF), Fig Tree Group, South Africa. This unit is exceptional in that proximal, shallow-water and distal, deep-water facies are preserved. When compared to the distal, deep-water facies, the proximal samples show elevated U concentrations and moderately positive 56Fe values, indicating vertical stratification in dissolved oxygen contents. Confirmation of oxidizing conditions using U abundances is robustly constrained using samples that have been closed to U and Pb mobility using U–Th–Pb geochronology. This documents the oldest known preserved marine redox gradient in the rock record. The relative enrichment of O₂ in the upper water column is likely due to the existence of oxygen-producing microorganisms such as cyanobacteria. These results provide a new approach for identifying free oxygen in Earth’s ancient oceans, including confirming the age of redox proxies, and indicate that cyanobacteria evolved prior to 3.2 Ga.

  ROADMAP OBJECTIVES: 4.1 5.2 7.1

• Project 3D: SIMS Analyses of Filamentous Fossil Microbes From the ~3,465 Ma-Old Apex Chert May Reveal Their Physiology

  Permineralized carbonaceous filamentous fossils of the ~3465 Ma Apex chert of Western Australia are among the oldest known in the geological record. Eleven taxa of Bacteria Incertae Sedis (microbes of uncertain systematic affinities) have been described from this assemblage, defined on the basis of the medial and terminal cell size and shape of 200 specimens. To assess their physiology and, thus, their biological affinities, 20 additional specimens, referable to six of the taxa, have been prepared in thin section. Samples will be cut out and repolished to expose each microfossil individually for δ¹³C analyses by use of secondary ion mass spectroscopy (SIMS). Their carbonaceous composition has been mapped by Raman spectroscopy and, as applicable, by confocal laser scanning microscopy, CLSM. This is the first study to use optical microscopy, Raman spectroscopy, SIMS and CLSM, to analyze multiple individual microscopic fossils from a single deposit — applied here to among the oldest fossils now known in the geological record.

  ROADMAP OBJECTIVES: 4.1

• Project 3E: Genesis of High-δ18O Archean Chert, Pilbara Craton, Australia

  The cherts of the Strelley Pool formation host the oldest generally accepted evidence of life, stromatolites, organic matter and microfossils. Oxygen isotope ratios provide independent evidence to evaluate conditions when quartz formed. These results support habitable conditions during formation of early quartz and late alteration for genesis of late high δ¹⁸O-quartz.

  ROADMAP OBJECTIVES: 1.1 4.1 7.1

• Project 3F: Searching for Ancient Impact Events Through Detrital Shocked Zircons

  Understanding how quickly planetary surface environments evolve on newly accreted worlds is critical for predicting when habitable conditions are established. The meteorite impact history of the inner solar system strongly indicates that the Earth was subject to a global impact bombardment during the first few hundred million years after accretion. The scope, timing, and consequences of this profound process are hotly debated. This project investigated populations of detrital zircons in Archean sedimentary rocks to search for tell-tale signs of impact processes in the form of shock-induced microstructures that are diagnostic of impact. Such features have been shown to survive in detrital shocked zircons eroded from known impact structures on Earth, including the Vredefort, Sudbury, and Santa Fe craters. We have investigated populations of 1,000 zircons per sample using backscattered electron imaging of grain exteriors with a scanning electron microscope. Thus far we have surveyed zircons separated from rocks collected from the Yilgarn craton (Australia), North China craton (China), Wyoming craton (USA), the Superior craton (Canada). While intriguing microstructures have been observed, thus far no confirmed shock microstructures have been encountered in our Archean sample suites. Our inability the identify shocked grains in populations of 1,000 zircons (per sample) does not necessarily mean shocked grains are absent; our results provide constraints that if they are present, they are in abundances of <0.1% in the detrital population of the rocks investigated. We are currently in the process of more in-depth surveying (e.g., >1000 grains/sample) to test for very low frequency occurrency events. Our detailed search continues…

  ROADMAP OBJECTIVES: 1.1 4.1 4.3

• Project 3G: A 3,400 Ma-Old Shallow Water Anaerobic Sulfuretum Evidences the Anoxic Archean Atmosphere

  Carbonaceous cherts of the ~3430 Ma Strelley Pool Formation contain innumerable “swirls” of fossilized sulfuretum bacteria encompassing quartz-replaced anhydrite nodules intermixed with layered assemblages of phototrophic filamentous fossil microbes. The geologic setting of the fossil-hosting unit, the preservation of the sulfuretum swirls adpressed to quartz pseudomorphs of precipitated anhydrite or gypsum, and the lack of physical disruption of the assemblage document its near-surface quiescent marine environment. The anaerobic physiology of the sulfuretum microbes indicates that Earth’s surface was anoxic. This exceedingly ancient biota is therefore interpreted to be composed of anaerobic H2S-producing sulfuretum microbes and H₂S-using anoxygenic phototrophic bacteria. As such, this first-identified fossil microbial consortium provides firm evidence of the anoxia of Earth’s early environment.

  ROADMAP OBJECTIVES: 4.1 5.2 6.2 7.2

• Project 4A: New in Situ Techniques (CLSM and Raman) Solve the Problem Presented by the Disaggregation of Acid-Macerated Organic-Walled Microfossils

  Because of the enormous costs involved in Mars Surface Sample Return Missions, the search for evidence of past life in rocks from Mars is likely to hinge on the use of “safe,” non-intrusive, non-destructive techniques to establish the biogenicity of any detected fossil-like objects by analyses of their cellular morphology and molecular composition. The most promising rock types to preserve such evidence are chemically precipitated sediments such as cherts, gypsums, carbonates and phosphates — all of which on Earth have been shown to be richly fossiliferous. The organic-walled microbes in such rocks are typically not amenable to investigation by the commonly used but rock-destroying technique of acid maceration. This study shows that the combined use of optical microscopy, confocal laser scanning microscopy, and Raman spectroscopy solves this problem, documenting effective means for the investigation of Mars rocks.

  ROADMAP OBJECTIVES: 7.2

• Project 4B: New Standards for Analysis of O and C Isotope Ratios in Ca-Mg-Fe Carbonates

  Stable isotope ratios are a powerful tool for determining the temperature and fluid conditions during formation of carbonates that host evidence for early life on Earth. However, sedimentary carbonates are often zoned at μm-scale and conventional analysis yields average values. We developed a suite of standards for the dolomite-ankerite series that allow us to make the first accurate SIMS (Secondary ion mass spectrometry) analyses oxygen and carbon isotope ratios at 1-10 μm-scale for these important carbonate minerals.

  ROADMAP OBJECTIVES: 1.1 4.1 4.3 7.1