Immunoanalytical Detection of Conserved Peptides: Refining the Universe of Biomarker Targets in Planetary Exploration.

Ancient peptides are remnants of early biochemistry that continue to play pivotal roles in current proteins. They are simple molecules yet complex enough to exhibit independent functions, being products of an evolved biochemistry at the interface of life and nonlife. Their adsorption to minerals may contribute to their stabilization and preservation over time. To investigate the feasibility of conserved peptide sequences and structures as target biomarkers for the search for life on Mars or other planetary bodies, we conducted a bioinformatics selection of well-conserved ancient peptides and produced polyclonal antibodies for their detection using fluorescence microarray immunoassays. Additionally, we explored how adsorbing peptides to Mars-representative minerals to form organomineral complexes could affect their immunological detection. The results demonstrated that the selected peptides exhibited autonomous folding, with some of them regaining their structure, even after denaturation. Furthermore, their cognate antibodies detected their conformational features regardless of amino acid sequences, thereby broadening the spectrum of target peptide sequences. While certain antibodies displayed unspecific binding to bare minerals, we validated that peptide-mineral complexes can be detected using sandwich immunoassays, as confirmed through desorption and competitive assays. Consequently, we conclude that the diversity of peptide sequences and structures suitable for use as target biomarkers in astrobiology can be constrained to a few well conserved sets, and they can be detected even if they are adsorbed in organomineral complexes.

In Situ Real-Time Monitoring for Aseptic Drilling: Lessons Learned from the Atacama Rover Astrobiology Drilling Studies Contamination Control Strategy and Implementation and Application to the Icebreaker Mars Life Detection Mission.

In 2019, the Atacama Rover Astrobiology Drilling Studies (ARADS) project field-tested an autonomous rover-mounted robotic drill prototype for a 6-Sol life detection mission to Mars (Icebreaker). ARADS drilled Mars-like materials in the Atacama Desert (Chile), one of the most life-diminished regions on Earth, where mitigating contamination transfer into life-detection instruments becomes critical. Our Contamination Control Strategy and Implementation (CCSI) for the Sample Handling and Transfer System (SHTS) hardware (drill, scoop and funnels) included out-of-simulation protocol testing (out-of-sim) for hardware decontamination and verification during the 6-Sol simulation (in-sim). The most effective five-step decontamination combined safer-to-use sterilants (3%_hydrogen-peroxide-activated 5%_sodium-hypochlorite), and in situ real-time verification by adenosine triphosphate (ATP) and Signs of Life Detector (SOLID) Fluorescence Immunoassay for characterization hardware bioburden and airborne contaminants. The 20- to 40-min protocol enabled a 4-log bioburden reduction down to <0.1 fmoles ATP detection limit (funnels and drill) to 0.2-0.7 fmoles (scoop) of total ATP. The (post-cleaning) hardware background was 0.3 to 1-2 attomoles ATP/cm2 (cleanliness benchmark background values) equivalent to ca. 1-10 colony forming unit (CFU)/cm2. Further, 60-100% of the in-sim hardware background was ≤3-4 bacterial cells/cm2, the threshold limit for Class <7 aseptic operations. Across the six Sols, the flux of airborne contaminants to the drill sites was ∼5 and ∼22 amoles ATP/(cm2·day), accounting for an unexpectedly high Fluorescence Intensity (FI) signal (FI: ∼6000) against aquatic cyanobacteria, but negligible anthropogenic contribution. The SOLID immunoassay also detected microorganisms from multiple habitats across the Atacama Desert (anoxic, alkaline/acidic microenvironments in halite fields, playas, and alluvial fans) in both airborne and post-cleaning hardware background. Finally, the hardware ATP background was 40-250 times lower than the ATP in cores. Similarly, the FI peaks (FImax) against the microbial taxa and molecular biomarkers detected in the post-cleaned hardware (FI: ∼1500-1600) were 5-10 times lower than biomarkers in drilled sediments, excluding significant interference with putative biomarker found in cores. Similar protocols enable the acquisition of contamination-free materials for ultra-sensitive instruments analysis and the integrity of scientific results. Their application can augment our scientific knowledge of the distribution of cryptic life on Mars-like grounds and support life-detection robotic and human-operated missions to Mars.

Traces of Ancient Life in Oceanic Basalt Preserved as Iron-Mineralized Ultrastructures: Implications for Detecting Extraterrestrial Biosignatures.

Benefiting from their adaptability to extreme environments, subsurface microorganisms have been discovered in sedimentary and igneous rock environments on Earth and have been advocated as candidates in the search for extraterrestrial life. In this article, we study iron-mineralized microstructures in calcite-filled veins within basaltic pillows of the late Ladinian Fernazza group (Middle Triassic, 239 Ma) in Italy. These microstructures represent diverse morphologies, including filaments, globules, nodules, and micro-digitate stromatolites, which are similar to extant iron-oxidizing bacterial communities. In situ analyses including Raman spectroscopy have been used to investigate the morphological, elemental, mineralogical, and bond-vibrational modes of the microstructures. According to the Raman spectral parameters, iron minerals preserve heterogeneous ultrastructures and crystallinities, coinciding with the morphologies and precursor microbial activities. The degree of crystallinity usually represents a microscale gradient decreasing toward previously existing microbial cells, revealing a decline of mineralization due to microbial activities. This study provides an analog of possible rock-dwelling subsurface life on Mars or icy moons and advocates Raman spectroscopy as an efficient tool for in situ analyses. We put forward the concept that ultrastructural characteristics of minerals described by Raman spectral parameters corresponding to microscale morphologies could be employed as carbon-lean biosignatures in future space missions.

Impact of M Dwarfs Ultraviolet Radiation on Prebiotic Chemistry: The Case of Adenine.

To date, several exoplanets have been found to orbit within the habitable zone of main sequence M stars (M dwarfs). These stars exhibit different levels of chromospheric activity that produces ultraviolet (UV) radiation. UV may be harmful to life, but it can also trigger reactions of prebiotic importance on the surface of a potentially habitable planet (PHP). We created a code to obtain the adenine yield for a known adenine synthesis route from diaminomaleonitrile (DAMN). We used computational methods to calculate the reaction coefficient rates (photolysis rate J and rate constant K) for the intermediate molecules DAMN, diaminofumaronitrile (DAFN), and 4-aminoimidazole-5-carbonitrile (AICN) of the adenine synthesis route. We used stellar UV sources and a mercury lamp to compare the theoretical results with experiments performed with lamps. The surface UV flux of planets in the habitable zone of two active M dwarfs (Proxima Centauri and AD Leonis) and the prebiotic Earth was calculated using the photochemical model ATMOS, considering a CO2-N2-H2O atmosphere. We obtained UV absorption coefficients for DAMN and DAFN and thermodynamic parameters that are useful for prebiotic chemistry studies. According to our results, experiments using UV lamps may underestimate the photolysis production of molecules of prebiotic importance. Our results indicate that photolysis reactions are fast with a yield of 50% of AICN in 10 s for the young Sun and ∼1 h for Proxima Centauri b. Planets around active M dwarfs may provide the most favorable environment for UV-mediated production of compounds relevant to the origins of life. The kinetic reaction AICN + HCN  adenine is the bottleneck of the pathway with reaction rates <10-22 L/(mol·s).

The COSPAR Planetary Protection Policy for robotic missions to Mars: A review of current scientific knowledge and future perspectives.

Planetary protection guidance for martian exploration has become a notable point of discussion over the last decade. This is due to increased scientific interest in the habitability of the red planet with updated techniques, missions becoming more attainable by smaller space agencies, and both the private sector and governments engaging in activities to facilitate commercial opportunities and human-crewed missions. The international standards for planetary protection have been developed through consultation with the scientific community and the space agencies by the Committee on Space Research's (COSPAR) Panel on Planetary Protection, which provides guidance for compliance with the Outer Space Treaty of 1967. In 2021, the Panel evaluated recent scientific data and literature regarding the planetary protection requirements for Mars and the implications of this on the guidelines. In this paper, we discuss the COSPAR Planetary Protection Policy for Mars, review the new scientific findings and discuss the next steps required to enable the next generation of robotic missions to Mars.

Selection and Analytical Performances of the Dragonfly Mass Spectrometer Gas Chromatographic Columns to Support the Search for Organic Molecules of Astrobiological Interest on Titan.

Titan is a key planetary body for astrobiology, with the presence of a subsurface ocean and a dense atmosphere, in which complex chemistry is known to occur. Approximately 1-Titan-year after the Cassini-Huygens mission arrived in the saturnian system, Dragonfly rotorcraft will land on Titan's surface by 2034 for an exhaustive geophysical and chemical investigation of the Shangri-La organic sand sea region. Among the four instruments onboard Dragonfly, the Dragonfly Mass Spectrometer (DraMS) is dedicated to analyze the chemical composition of surface samples and noble gases in the atmosphere. One of the DraMS analysis modes, the Gas Chromatograph-Mass Spectrometer (GC-MS), is devoted to the detection and identification of organic molecules that could be involved in the development of a prebiotic chemistry or even representative of traces of past or present life. Therefore, DraMS-GC subsystem should be optimized to detect and identify relevant organic compounds to meet this objective. This work is focused on the experimental methods employed to select the chromatographic column to be integrated in DraMS-GC, to assess the analytical performances of the column selected, and also to assess the performances of the second DraMS-GC column, which is devoted to the separation of organic enantiomers. Four different stationary phases have been tested to select the most relevant one for the separation of the targeted chemical species. The results show that the stationary phase composed of polymethyl (95%) diphenyl (5%) siloxane is the best compromise in terms of efficiency, robustness, and retention times of the molecules. The combination of the general and the chiral columns in DraMS is perfectly suited to in situ chemical analysis on Titan and for the detection of expected diverse and complex organic compounds.

Laboratory experiment of ATP measurement using Mars soil simulant: as a method for extraterrestrial life detection.

We present a laboratory experiment of ATP measurement using Mars soil simulant and Escherichia coli (E. coli) with a point of view for future application to searching extraterrestrial life. We used Mars Global Simulant MGS-1 (Exolith Lab) as soil simulant, added E. coli suspension to it, then the soil simulant with E. coli was dried. Various configurations of samples with different E. coli density, 1.75 × 102, 1.75 × 103, 1.75 × 104, 1.75 × 105, and 1.75 × 106 cells (g soil)-1, were prepared together with controls. ATP extraction reagent and luminescence reagent were added to the sample, and bioluminescence measurement was performed. The result suggests significant detection of ATP for samples with E. coli density used in this work. Similar experiments but without the soil simulant were carried out, and results with and without the soil simulant are compared. Based on the ATP measurement studied in this work, we discussed extraterrestrial life search, planetary protection, relation with the panspermia hypothesis, and also other applications.

An easy-to-use function to assess deep space radiation in human brains.

Health risks from radiation exposure in space are an important factor for astronauts' safety as they venture on long-duration missions to the Moon or Mars. It is important to assess the radiation level inside the human brain to evaluate the possible hazardous effects on the central nervous system especially during solar energetic particle (SEP) events. We use a realistic model of the head/brain structure and calculate the radiation deposit therein by realistic SEP events, also under various shielding scenarios. We then determine the relation between the radiation dose deposited in different parts of the brain and the properties of the SEP events and obtain some simple and ready-to-use functions which can be used to quickly and reliably forecast the event dose in the brain. Such a novel tool can be used from fast nowcasting of the consequences of SEP events to optimization of shielding systems and other mitigation strategies of astronauts in space.

The Radiation Stability of Thymine in Solid H2O.

Nucleobases are of significant importance to all known organisms, may be an important building block of life, and could be important biosignatures of current or past life. Given their potential significance to the field of astrobiology, it is important to understand the survival of these molecules when subjected to ionizing radiation as is present in a range of extraterrestrial environments. In this work, we present data on the kinetics of the radiolytic destruction of pure thymine and water + thymine ice mixtures at temperatures from 13 to 150 K. Rate constants were measured using in situ infrared spectroscopy, and radiolytic half-lives for thymine were computed for different planetary and interstellar environments. Our results demonstrate that the survival of thymine decreases as the dilution of thymine in water increases. Additionally, we find that thymine survival increases with ice temperature and that this decrease may be related to structure of the ice matrix.

Simulating Mars Drilling Mission for Searching for Life: Ground-Truthing Lipids and Other Complex Microbial Biomarkers in the Iron-Sulfur Rich Río Tinto Analog.

Sulfate and iron oxide deposits in Río Tinto (Southwestern Spain) are a terrestrial analog of early martian hematite-rich regions. Understanding the distribution and drivers of microbial life in iron-rich environments can give critical clues on how to search for biosignatures on Mars. We simulated a robotic drilling mission searching for signs of life in the martian subsurface, by using a 1m-class planetary prototype drill mounted on a full-scale mockup of NASA's Phoenix and InSight lander platforms. We demonstrated fully automated and aseptic drilling on iron and sulfur rich sediments at the Río Tinto riverbanks, and sample transfer and delivery to sterile containers and analytical instruments. As a ground-truth study, samples were analyzed in the field with the life detector chip immunoassay for searching microbial markers, and then in the laboratory with X-ray diffraction to determine mineralogy, gas chromatography/mass spectrometry for lipid composition, isotope-ratio mass spectrometry for isotopic ratios, and 16S/18S rRNA genes sequencing for biodiversity. A ubiquitous presence of microbial biomarkers distributed along the 1m-depth subsurface was influenced by the local mineralogy and geochemistry. The spatial heterogeneity of abiotic variables at local scale highlights the importance of considering drill replicates in future martian drilling missions. The multi-analytical approach provided proof of concept that molecular biomarkers varying in compositional nature, preservation potential, and taxonomic specificity can be recovered from shallow drilling on iron-rich Mars analogues by using an automated life-detection lander prototype, such as the one proposed for NASA's IceBreaker mission proposal.

Testing Flight-like Pyrolysis Gas Chromatography-Mass Spectrometry as Performed by the Mars Organic Molecule Analyzer Onboard the ExoMars 2020 Rover on Oxia Planum Analog Samples.

The Mars Organic Molecule Analyzer (MOMA) onboard the ExoMars 2020 rover (to be landed in March 2021) utilizes pyrolysis gas chromatography-mass spectrometry (GC-MS) with the aim to detect organic molecules in martian (sub-) surface materials. Pyrolysis, however, may thermally destroy and transform organic matter depending on the temperature and nature of the molecules, thus altering the original molecular signatures. In this study, we tested MOMA flight-like pyrolysis GC-MS without the addition of perchlorates on well-characterized natural mineralogical analog samples for Oxia Planum, the designated ExoMars 2020 landing site. Experiments were performed on an iron-rich shale (that is rich in Fe-Mg-smectites) and an opaline chert, with known organic matter compositions, to test pyrolytic effects related to heating in the MOMA oven. Two hydrocarbon standards (n-octadecane and phytane) were also analyzed. The experiments show that during stepwise pyrolysis (300°C, 500°C, and 700°C), (1) low-molecular-weight hydrocarbon biomarkers (such as acyclic isoprenoids and aryl isoprenoids) can be analyzed intact, (2) discrimination between free and complex molecules (macromolecules) is principally possible, (3) secondary pyrolysis products and carryover may affect the 500°C and 700°C runs, and (4) the type of the organic matter (functionalized vs. defunctionalized) governs the pyrolysis outcome rather than the difference in mineralogy. Although pyrosynthesis reactions and carryover clearly have to be considered in data interpretation, our results demonstrate that pyrolysis GC-MS onboard MOMA operated under favorable conditions (e.g., no perchlorates) will be capable of providing important structural information on organic matter found on Mars, particularly when used in conjunction with other techniques on MOMA, including derivatization and thermochemolysis GC-MS and laser desorption/ionization-MS.

Analog Experiments for the Identification of Trace Biosignatures in Ice Grains from Extraterrestrial Ocean Worlds.

Reliable identification of biosignatures, such as amino acids, fatty acids, and peptides, on extraterrestrial ocean worlds is a key prerequisite for space missions that search for life or its emergence on these worlds. One promising approach is the use of high-performance in situ impact ionization mass spectrometers to sample water ice grains emerging from ocean-bearing moons such as Europa or Enceladus. A predecessor of such detectors, the Cosmic Dust Analyzer on board the Cassini spacecraft, has proven to be very successful in analyzing inorganic and organic ocean constituents and with that characterizing the habitability of Enceladus ocean. However, biosignatures have not been definitively identified in extraterrestrial ocean environments so far. Here, we investigate with an analog experiment the spectral appearance of amino acids, fatty acids, and peptides in water ice grains, together with their detection limits, as applicable to spaceborne mass spectrometers. We employ a laboratory-based laser induced liquid beam ion desorption technique, proven to simulate accurately the impact ionization mass spectra of water ice grains over a wide range of impact speeds. The investigated organics produce characteristic mass spectra, with molecular peaks as well as clearly identifiable, distinctive fragments. We find the detection limits of these key biosignatures to be at the µM or nM level, depending on the molecular species and instrument polarity, and infer that impact ionization mass spectrometers are most sensitive to the molecular peaks of these biosignatures at encounter velocities of 4-6 km/s.

Phosphine as a Biosignature Gas in Exoplanet Atmospheres.

A long-term goal of exoplanet studies is the identification and detection of biosignature gases. Beyond the most discussed biosignature gas O2, only a handful of gases have been considered in detail. In this study, we evaluate phosphine (PH3). On Earth, PH3 is associated with anaerobic ecosystems, and as such, it is a potential biosignature gas in anoxic exoplanets. We simulate the atmospheres of habitable terrestrial planets with CO2- and H2-dominated atmospheres and find that PH3 can accumulate to detectable concentrations on planets with surface production fluxes of 1010 to 1014 cm-2 s-1 (corresponding to surface concentrations of 10s of ppb to 100s of ppm), depending on atmospheric composition and ultraviolet (UV) irradiation. While high, the surface flux values are comparable to the global terrestrial production rate of methane or CH4 (1011 cm-2 s-1) and below the maximum local terrestrial PH3 production rate (1014 cm-2 s-1). As with other gases, PH3 can more readily accumulate on low-UV planets, for example, planets orbiting quiet M dwarfs or with a photochemically generated UV shield. PH3 has three strong spectral features such that in any atmosphere scenario one of the three will be unique compared with other dominant spectroscopic molecules. Phosphine's weakness as a biosignature gas is its high reactivity, requiring high outgassing rates for detectability. We calculate that tens of hours of JWST (James Webb Space Telescope) time are required for a potential detection of PH3. Yet, because PH3 is spectrally active in the same wavelength regions as other atmospherically important molecules (such as H2O and CH4), searches for PH3 can be carried out at no additional observational cost to searches for other molecular species relevant to characterizing exoplanet habitability. Phosphine is a promising biosignature gas, as it has no known abiotic false positives on terrestrial planets from any source that could generate the high fluxes required for detection.

Investigating the Effect of Perchlorate on Flight-like Gas Chromatography-Mass Spectrometry as Performed by MOMA on board the ExoMars 2020 Rover.

The Mars Organic Molecule Analyzer (MOMA) instrument on board ESA's ExoMars 2020 rover will be essential in the search for organic matter. MOMA applies gas chromatography-mass spectrometry (GC-MS) techniques that rely on thermal volatilization. Problematically, perchlorates and chlorates in martian soils and rocks become highly reactive during heating (>200°C) and can lead to oxidation and chlorination of organic compounds, potentially rendering them unidentifiable. Here, we analyzed a synthetic sample (alkanols and alkanoic acids on silica gel) and a Silurian chert with and without Mg-perchlorate to evaluate the applicability of MOMA-like GC-MS techniques to different sample types and assess the impact of perchlorate. We used a MOMA flight analog system coupled to a commercial GC-MS to perform MOMA-like pyrolysis, in situ derivatization, and in situ thermochemolysis. We show that pyrolysis can provide a sufficient overview of the organic inventory but is strongly affected by the presence of perchlorates. In situ derivatization facilitates the identification of functionalized organics but showed low efficiency for n-alkanoic acids. Thermochemolysis is shown to be an effective technique for the identification of both refractory and functional compounds. Most importantly, this technique was barely affected by perchlorates. Therefore, MOMA GC-MS analyses of martian surface/subsurface material may be less affected by perchlorates than commonly thought, in particular when applying the full range of available MOMA GC-MS techniques.

Metabolic Processes Preserved as Biosignatures in Iron-Oxidizing Microorganisms: Implications for Biosignature Detection on Mars.

Iron-oxidizing bacteria occupy a distinct environmental niche. These chemolithoautotrophic organisms require very little oxygen (when neutrophilic) or outcompete oxygen for access to Fe(II) (when acidophilic). The utilization of Fe(II) as an electron donor makes them strong analog organisms for any potential life that could be found on Mars. Despite their importance to the elucidation of early life on, and potentially beyond, Earth, many details of their metabolism remain unknown. By using on-line thermochemolysis and gas chromatography-mass spectrometry (GC-MS), a distinct signal for a low-molecular-weight molecule was discovered in multiple iron-oxidizing isolates as well as several iron-dominated environmental samples, from freshwater and marine environments and in both modern and older iron rock samples. This GC-MS signal was neither detected in organisms that did not use Fe(II) as an electron donor nor present in iron mats in which organic carbon was destroyed by heating. Mass spectral analysis indicates that the molecule bears the hallmarks of a pterin-bearing molecule. Genomic analysis has previously identified a molybdopterin that could be part of the electron transport chain in a number of lithotrophic iron-oxidizing bacteria, suggesting one possible source for this signal is the pterin component of this protein. The rock samples indicate the possibility that the molecule can be preserved within lithified sedimentary rocks. The specificity of the signal to organisms requiring iron in their metabolism makes this a novel biosignature with which to investigate both the evolution of life on ancient Earth and potential life on Mars.

Effects of Gamma and Electron Radiation on the Structural Integrity of Organic Molecules and Macromolecular Biomarkers Measured by Microarray Immunoassays and Their Astrobiological Implications.

High-energy ionizing radiation in the form of solar energetic particles and galactic cosmic rays is pervasive on the surface of planetary bodies with thin atmospheres or in space facilities for humans, and it may seriously affect the chemistry and the structure of organic and biological material. We used fluorescent microarray immunoassays to assess how different doses of electron and gamma radiations affect the stability of target compounds such as biological polymers and small molecules (haptens) conjugated to large proteins. The radiation effect was monitored by measuring the loss in the immunoidentification of the target due to an impaired ability of the antibodies for binding their corresponding irradiated and damaged epitopes (the part of the target molecule to which antibodies bind). Exposure to electron radiation alone was more damaging at low doses (1 kGy) than exposure to gamma radiation alone, but this effect was reversed at the highest radiation dose (500 kGy). Differences in the dose-effect immunoidentification patterns suggested that the amount (dose) and not the type of radiation was the main factor for the cumulative damage on the majority of the assayed molecules. Molecules irradiated with both types of radiation showed a response similar to that of the individual treatments at increasing radiation doses, although the pattern obtained with electrons only was the most similar. The calculated radiolysis constant did not show a unique pattern; it rather suggested a different behavior perhaps associated with the unique structure of each molecule. Although not strictly comparable with extraterrestrial conditions because the irradiations were performed under air and at room temperature, our results may contribute to understanding the effects of ionizing radiation on complex molecules and the search for biomarkers through bioaffinity-based systems in planetary exploration.

Instrumentation for Testing Whether the Icy Moons of the Gas and Ice Giants Are Inhabited.

Evidence of life beyond Earth may be closer than we think, given that the forthcoming missions to the jovian system will be equipped with instruments capable of probing Europa's icy surface for possible biosignatures, including chemical biomarkers, despite the strong radiation environment. Geochemical biomarkers may also exist beyond Europa on icy moons of the gas giants. Sulfur is proposed as a reliable geochemical biomarker for approved and forthcoming missions to the outer solar system. Key Words: JUICE mission-Clipper mission-Geochemical biomarkers-Europa-Moons of the ice giants-Geochemistry-Mass spectrometry. Astrobiology 17, 958-961.

Correlations Between Life-Detection Techniques and Implications for Sampling Site Selection in Planetary Analog Missions.

We conducted an analog sampling expedition under simulated mission constraints to areas dominated by basaltic tephra of the Eldfell and Fimmvörðuháls lava fields (Iceland). Sites were selected to be "homogeneous" at a coarse remote sensing resolution (10-100 m) in apparent color, morphology, moisture, and grain size, with best-effort realism in numbers of locations and replicates. Three different biomarker assays (counting of nucleic-acid-stained cells via fluorescent microscopy, a luciferin/luciferase assay for adenosine triphosphate, and quantitative polymerase chain reaction (qPCR) to detect DNA associated with bacteria, archaea, and fungi) were characterized at four nested spatial scales (1 m, 10 m, 100 m, and >1 km) by using five common metrics for sample site representativeness (sample mean variance, group F tests, pairwise t tests, and the distribution-free rank sum H and u tests). Correlations between all assays were characterized with Spearman's rank test. The bioluminescence assay showed the most variance across the sites, followed by qPCR for bacterial and archaeal DNA; these results could not be considered representative at the finest resolution tested (1 m). Cell concentration and fungal DNA also had significant local variation, but they were homogeneous over scales of >1 km. These results show that the selection of life detection assays and the number, distribution, and location of sampling sites in a low biomass environment with limited a priori characterization can yield both contrasting and complementary results, and that their interdependence must be given due consideration to maximize science return in future biomarker sampling expeditions. Key Words: Astrobiology-Biodiversity-Microbiology-Iceland-Planetary exploration-Mars mission simulation-Biomarker. Astrobiology 17, 1009-1021.

On the Habitability of Desert Varnish: A Combined Study by Micro-Raman Spectroscopy, X-ray Diffraction, and Methylated Pyrolysis-Gas Chromatography-Mass Spectrometry.

In 2020, the ESA ExoMars and NASA Mars 2020 missions will be launched to Mars to search for evidence of past and present life. In preparation for these missions, terrestrial analog samples of rock formations on Mars are studied in detail in order to optimize the scientific information that the analytical instrumentation will return. Desert varnishes are thin mineral coatings found on rocks in arid and semi-arid environments on Earth that are recognized as analog samples. During the formation of desert varnishes (which takes many hundreds of years), organic matter is incorporated, and microorganisms may also play an active role in the formation process. During this study, four complementary analytical techniques proposed for Mars missions (X-ray diffraction [XRD], Raman spectroscopy, elemental analysis, and pyrolysis-gas chromatography-mass spectrometry [Py-GC-MS]) were used to interrogate samples of desert varnish and describe their capacity to sustain life under extreme scenarios. For the first time, both the geochemistry and the organic compounds associated with desert varnish are described with the use of identical sets of samples. XRD and Raman spectroscopy measurements were used to nondestructively interrogate the mineralogy of the samples. In addition, the use of Raman spectroscopy instruments enabled the detection of ß-carotene, a highly Raman-active biomarker. The content and the nature of the organic material in the samples were further investigated with elemental analysis and methylated Py-GC-MS, and a bacterial origin was determined to be likely. In the context of planetary exploration, we describe the habitable nature of desert varnish based on the biogeochemical composition of the samples. Possible interference of the geological substrate on the detectability of pyrolysis products is also suggested. Key Words: Desert varnish-Habitability-Raman spectroscopy-Py-GC-MS-XRD-ExoMars-Planetary science. Astrobiology 17, 1123-1137.