The Preservation and Spectral Detection of Historic Museum Specimen Microbial Mat Biosignatures Within Martian Dust: Lessons Learned for Mars Exploration and Sample Return.

The key building blocks for life on Mars could be preserved within potentially habitable paleo-depositional settings with their detection possible by utilizing mid-infrared spectroscopy; however, a definite identification and confirmation of organic or even biological origin will require the samples to be returned to Earth. In the present study, Fourier-transform infrared (FTIR) spectroscopic techniques were used to characterize both mineralogical and organic materials within Mars dust simulant JSC Mars-1 and ancient Antarctic cyanobacterial microbial mats from 1901 to 1904 Discovery Expedition. When FTIR spectroscopy is applied to cyanobacterial microbial mat communities, the resulting spectra will reflect the average biochemical composition of the mats rather than taxa-specific spectral patterns of the individual organisms and can thus be considered as a total chemical analysis of the mat colony. This study also highlights the potential difficulties in the detection of these communities on Mars and which spectral biosignatures will be most detectable within geological substrates. Through the creation and analysis of a suite of dried microbial mat material and Martian dust simulant mixtures, the spectral signatures and wavenumber positions of CHx aliphatic hydrocarbons and the C-O and O-H bands of polysaccharides remained detectable and may be detectable within sample mixtures obtained through Mars Sample Return activities.

Variable and Large Losses of Diagnostic Biomarkers After Simulated Cosmic Radiation Exposure in Clay- and Carbonate-Rich Mars Analog Samples.

Mars has been exposed to ionizing radiation for several billion years, and as part of the search for life on the Red Planet, it is crucial to understand the impact of radiation on biosignature preservation. Several NASA and ESA missions are looking for evidence of ancient life in samples collected at depths shallow enough that they have been impacted by galactic cosmic rays (GCRs). In this study, we exposed a diverse set of Mars analog samples to 0.9 Megagray (MGy) of gamma radiation to mimic 15 million years of exposure on the Martian surface. We measured no significant impact of GCRs on the total organic carbon (TOC) and bulk stable C isotopes in samples with initial TOC concentration > 0.1 wt. %; however, diagnostic molecular biosignatures presented a wide range of degradation that didn't correlate to factors like mineralogy, TOC, water content, and surface area. Exposure dating suggests that the surface of Gale crater has been irradiated at more than five times our dose, yet using this relatively low dose and "best-case scenario" geologically recalcitrant biomarkers, large and variable losses were nevertheless evident. Our results empasize the importance of selecting sampling sites at depth or recently exposed at the Martian surface.

Radiolytic Effects on Biological and Abiotic Amino Acids in Shallow Subsurface Ices on Europa and Enceladus.

Europa and Enceladus are key targets to search for evidence of life in our solar system. However, the surface and shallow subsurface of both airless icy moons are constantly bombarded by ionizing radiation that could degrade chemical biosignatures. Therefore, sampling of icy surfaces in future life detection missions to Europa and Enceladus requires a clear understanding of the necessary ice depth where unaltered organic biomolecules might be present. We conducted radiolysis experiments by exposing individual amino acids in ices and amino acids from dead microorganisms in ices to gamma radiation to simulate conditions on these icy worlds. In the pure amino acid samples, glycine did not show a detectable decrease in abundance, whereas the abundance of isovaline decreased by 40% after 4 MGy of exposure. Amino acids in dead Escherichia coli (E. coli) organic matter exhibited a gradual decline in abundances with the increase of exposure dosage, although at much slower rates than individual amino acids. The majority of amino acids in dead A. woodii samples demonstrated a step function decline as opposed to a gradual decline. After the initial drop in abundance with 1 MGy of exposure, those amino acids did not display further decreases in abundance after exposure up to 4 MGy. New radiolysis constants for isolated amino acids and amino acids in dead E. coli material for Europa/Enceladus-like conditions have been derived. Slow rates of amino acid destruction in biological samples under Europa and Enceladus-like surface conditions bolster the case for future life detection measurements by Europa and Enceladus lander missions. Based on our measurements, the "safe" sampling depth on Europa is ∼20 cm at high latitudes of the trailing hemisphere in the area of little impact gardening. Subsurface sampling is not required for the detection of amino acids on Enceladus-these molecules will survive radiolysis at any location on the Enceladus surface. If the stability of amino acids observed in A. woodii organic materials is confirmed in other microorganisms, then the survival of amino acids from a potential biosphere in Europa ice would be significantly increased.

Siderite and vivianite as energy sources for the extreme acidophilic bacterium Acidithiobacillus ferrooxidans in the context of mars habitability.

Past and present habitability of Mars have been intensely studied in the context of the search for signals of life. Despite the harsh conditions observed today on the planet, some ancient Mars environments could have harbored specific characteristics able to mitigate several challenges for the development of microbial life. In such environments, Fe2+ minerals like siderite (already identified on Mars), and vivianite (proposed, but not confirmed) could sustain a chemolithoautotrophic community. In this study, we investigate the ability of the acidophilic iron-oxidizing chemolithoautotrophic bacterium Acidithiobacillus ferrooxidans to use these minerals as its sole energy source. A. ferrooxidans was grown in media containing siderite or vivianite under different conditions and compared to abiotic controls. Our experiments demonstrated that this microorganism was able to grow, obtaining its energy from the oxidation of Fe2+ that came from the solubilization of these minerals under low pH. Additionally, in sealed flasks without CO2, A. ferrooxidans was able to fix carbon directly from the carbonate ion released from siderite for biomass production, indicating that it could be able to colonize subsurface environments with little or no contact with an atmosphere. These previously unexplored abilities broaden our knowledge on the variety of minerals able to sustain life. In the context of astrobiology, this expands the list of geomicrobiological processes that should be taken into account when considering the habitability of environments beyond Earth, and opens for investigation the possible biological traces left on these substrates as biosignatures.

Acclimation mechanism of microalgal photosynthetic apparatus under low atmospheric pressures - new astrobiological perspectives in a Mars-like atmosphere.

This study reveals a new acclimation mechanism of the eukaryotic unicellular green alga Chlorella vulgaris in terms of the effect of varying atmospheric pressures on the structure and function of its photosynthetic apparatus using fluorescence induction measurements (JIP-test). The results indicate that low (400mbar) and extreme low (2 atmosphere (simulating the Mars atmosphere), reveals that the impact of extremely low atmospheric pressure on PQ mobility within the photosynthetic membrane, coupled with the low density of an almost 100% CO2 Mars-like atmosphere, results to a similar photosynthetic efficiency to that on Earth. These findings pave the way for the identification of novel functional acclimation mechanisms of microalgae to extreme environments that are vastly distinct from those found on Earth.

Rebuilding the Habitable Zone from the Bottom up with Computational Zones.

Computation, if treated as a set of physical processes that act on information represented by states of matter, encompasses biological systems, digital systems, and other constructs and may be a fundamental measure of living systems. The opportunity for biological computation, represented in the propagation and selection-driven evolution of information-carrying organic molecular structures, has been partially characterized in terms of planetary habitable zones (HZs) based on primary conditions such as temperature and the presence of liquid water. A generalization of this concept to computational zones (CZs) is proposed, with constraints set by three principal characteristics: capacity (including computation rates), energy, and instantiation (or substrate, including spatial extent). CZs naturally combine traditional habitability factors, including those associated with biological function that incorporate the chemical milieu, constraints on nutrients and free energy, as well as element availability. Two example applications are presented by examining the fundamental thermodynamic work efficiency and Landauer limit of photon-driven biological computation on planetary surfaces and of generalized computation in stellar energy capture structures (a.k.a. Dyson structures). It is suggested that CZs that involve nested structures or substellar objects could manifest unique observational signatures as cool far-infrared emitters. While these latter scenarios are entirely hypothetical, they offer a useful, complementary introduction to the potential universality of CZs.

Investigating Microbial Biosignatures in Aeolian Environments Using Micro-X-Ray: Simulation of PIXL Instrument Analyses at Jezero Crater Onboard the Perseverance Mars 2020 Rover.

Assessing the past habitability of Mars and searching for evidence of ancient life at Jezero crater via the Perseverance rover are the key objectives of NASA's Mars 2020 mission. Onboard the rover, PIXL (Planetary Instrument for X-ray Lithochemistry) is one of the best suited instruments to search for microbial biosignatures due to its ability to characterize chemical composition of fine scale textures in geological targets using a nondestructive technique. PIXL is also the first micro-X-ray fluorescence (XRF) spectrometer onboard a Mars rover. Here, we present guidelines for identifying and investigating a microbial biosignature in an aeolian environment using PIXL-analogous micro-XRF (µXRF) analyses. We collected samples from a modern wet aeolian environment at Padre Island, Texas, that contain buried microbial mats, and we analyzed them using µXRF techniques analogous to how PIXL is being operated on Mars. We show via µXRF technique and microscope images the geochemical and textural variations from the surface to ∼40 cm depth. Microbial mats are associated with heavy-mineral lags and show specific textural and geochemical characteristics that make them a distinct biosignature for this environment. Upon burial, they acquire a diffuse texture due to the expansion and contraction of gas-filled voids, and they present a geochemical signature rich in iron and titanium, which is due to the trapping of heavy minerals. We show that these intrinsic characteristics can be detected via µXRF analyses, and that they are distinct from buried abiotic facies such as cross-stratification and adhesion ripple laminations. We also designed and conducted an interactive survey using the Padre Island µXRF data to explore how different users chose to investigate a biosignature-bearing dataset via PIXL-like sampling strategies. We show that investigating biosignatures via PIXL-like analyses is heavily influenced by technical constraints (e.g., the XRF measurement characteristics) and by the variety of approaches chosen by different scientists. Lessons learned for accurately identifying and characterizing this biosignature in the context of rover-mission constraints include defining relative priorities among measurements, favoring a multidisciplinary approach to the decision-making process of XRF measurements selection, and considering abiotic results to support or discard a biosignature interpretation. Our results provide guidelines for PIXL analyses of potential biosignature on Mars.

Martian microbes research and lessons learnt for forensic science.

A new method for looking for life outside the Earth is used as an example to demonstrate how ways of presenting complex scientific concepts to the general public, used in planetary science, could be used in forensic science. The work led to a pared down, practical definition of detectable Life for planetary exploration, An organised system capable of processing energy sources to its advantage. For nearly three quarters of Earth's history the only lifeforms were microbes, which are the target for looking for extraterrestrial life. Microbes are microscopic and may be sparsely distributed, but their metabolic products can form large, durable rocks, much easier to find and which may contain the organisms or their remains. There are similar challenges in presenting astrobiological and forensic science. Both may have to deal with very large or very small numbers which are not immediately comprehensible but can be understood by analogy. To increase the impact on the listener or reader, dramatic analogues are valuable, for example, referring to the mineralised microbial metabolic products as, "fossilised breath of bacteria" demands the audience's attention and engages them before more detailed explanations are given. The power of practical experiments or demonstrations is most important to reinforce what might otherwise be a fairly abstract concept. Surprisingly, most of these approaches can be made to work equally well in both spoken and written forms as well as in both sciences.

Solvent constraints for biopolymer folding and evolution in extraterrestrial environments.

We propose that spontaneous folding and molecular evolution of biopolymers are two universal aspects that must concur for life to happen. These aspects are fundamentally related to the chemical composition of biopolymers and crucially depend on the solvent in which they are embedded. We show that molecular information theory and energy landscape theory allow us to explore the limits that solvents impose on biopolymer existence. We consider 54 solvents, including water, alcohols, hydrocarbons, halogenated solvents, aromatic solvents, and low molecular weight substances made up of elements abundant in the universe, which may potentially take part in alternative biochemistries. We find that along with water, there are many solvents for which the liquid regime is compatible with biopolymer folding and evolution. We present a ranking of the solvents in terms of biopolymer compatibility. Many of these solvents have been found in molecular clouds or may be expected to occur in extrasolar planets.

Microbial preference for chlorate over perchlorate under simulated shallow subsurface Mars-like conditions.

The Martian surface and shallow subsurface lacks stable liquid water, yet hygroscopic salts in the regolith may enable the transient formation of liquid brines. This study investigated the combined impact of water scarcity, UV exposure, and regolith depth on microbial survival under Mars-like environmental conditions. Both vegetative cells of Debaryomyces hansenii and Planococcus halocryophilus, alongside with spores of Aspergillus niger, were exposed to an experimental chamber simulating Martian environmental conditions (constant temperatures of about - 11 °C, low pressure of approximately 6 mbar, a CO2 atmosphere, and 2 h of daily UV irradiation). We evaluated colony-forming units (CFU) and water content at three different regolith depths before and after exposure periods of 3 and 7 days, respectively. Each organism was tested under three conditions: one without the addition of salts to the regolith, one containing sodium chlorate, and one with sodium perchlorate. Our results reveal that the residual water content after the exposure experiments increased with regolith depth, along with the organism survival rates in chlorate-containing and salt-free samples. The survival rates of the three organisms in perchlorate-containing regolith were consistently lower for all organisms and depths compared to chlorate, with the most significant difference being observed at a depth of 10-12 cm, which corresponds to the depth with the highest residual water content. The postulated reason for this is an increase in the salt concentration at this depth due to the freezing of water, showing that for these organisms, perchlorate brines are more toxic than chlorate brines under the experimental conditions. This underscores the significance of chlorate salts when considering the habitability of Martian environments.

Emergent ribozyme behaviors in oxychlorine brines indicate a unique niche for molecular evolution on Mars.

Mars is a particularly attractive candidate among known astronomical objects to potentially host life. Results from space exploration missions have provided insights into Martian geochemistry that indicate oxychlorine species, particularly perchlorate, are ubiquitous features of the Martian geochemical landscape. Perchlorate presents potential obstacles for known forms of life due to its toxicity. However, it can also provide potential benefits, such as producing brines by deliquescence, like those thought to exist on present-day Mars. Here we show perchlorate brines support folding and catalysis of functional RNAs, while inactivating representative protein enzymes. Additionally, we show perchlorate and other oxychlorine species enable ribozyme functions, including homeostasis-like regulatory behavior and ribozyme-catalyzed chlorination of organic molecules. We suggest nucleic acids are uniquely well-suited to hypersaline Martian environments. Furthermore, Martian near- or subsurface oxychlorine brines, and brines found in potential lifeforms, could provide a unique niche for biomolecular evolution.

Hydrochemical and isotopic characteristics of water sources for biological activity across a massive evaporite basin on the Tibetan Plateau: Implications for aquatic environments on early Mars.

Covered by vast eolian landforms, gravel deposits, and playas, the worldwide typical evaporite deposit land, Qaidam Basin, in northwestern China is analogous to early Mars when the aridification process had lasted for millions of years since the end of a wetter climate. This study aims to investigate the chemical and isotopic characteristics of waters in an evaporite-rich environment, as well as the habitable conditions therein, that have undergone a transformation similar to early Mars. In May 2023, a total of 26 water samples were collected across the representative central axis of a longitudinal aridity gradient in the Qaidam Basin, including categories of meteoric water, freshwater, standing water accumulated after precipitation, salty lacustrine water, and hypersaline brines to inspect compounds made up of carbon, nitrogen, phosphorus, sulfur, halogen, and metallic elements. As evaporation intensified, the salt types transformed from HCO3-Ca·Na to Cl·SO4-Na or ClMg. The dominance of carbonate will gradually be replaced by sulfate and chloride, leaving much more dilute and less detectable contents. The presence of trace ClO4-, ClO3-, ClO2-, and BrO3- was confirmed in a few of the sampled Qaidam waters, indicating the preservation of oxyhalides in waters within an arid region and possibly the presence of relevant microbial enzymes. The isotopes of water, carbonaceous, and nitrogenous compounds provide valuable references for either abiogenic or biogenic signatures. With undetectable amount, phosphorus was found to be the limiting nutrient in evaporative aquatic environments but not necessarily antibiosignatures. Overall, these results suggest that the paleo-lacustrine environments on Mars are more likely to preserve biosignatures if they feature the dominance of carbonate minerals, bioavailable nitrate, phosphorus, and organic carbon, the presence of thermodynamically unstable oxyhalides, and isotope ratios that point to the involvement of biological activity.

The Concept of Life on Venus Informs the Concept of Habitability.

An enduring question in astrobiology is how we assess extraterrestrial environments as being suitable for life. We suggest that the most reliable assessments of the habitability of extraterrestrial environments are made with respect to the empirically determined limits to known life. We discuss qualitatively distinct categories of habitability: empirical habitability that is constrained by the observed limits to biological activity; habitability sensu stricto, which is defined with reference to the known or unknown limits to the activity of all known organisms; and habitability sensu lato (habitability in the broadest sense), which is circumscribed by the limit of all possible life in the universe, which is the most difficult (and perhaps impossible) to determine. We use the cloud deck of Venus, which is temperate but incompatible with known life, as an example to elaborate and hypothesize on these limits.

Quantifying the Potential for Nitrate-Dependent Iron Oxidation on Early Mars: Implications for the Interpretation of Gale Crater Organics.

Geological evidence and atmospheric and climate models suggest habitable conditions occurred on early Mars, including in a lake in Gale crater. Instruments aboard the Curiosity rover measured organic compounds of unknown provenance in sedimentary mudstones at Gale crater. Additionally, Curiosity measured nitrates in Gale crater sediments, which suggests that nitrate-dependent Fe2+ oxidation (NDFO) may have been a viable metabolism for putative martian life. Here, we perform the first quantitative assessment of an NDFO community that could have existed in an ancient Gale crater lake and quantify the long-term preservation of biological necromass in lakebed mudstones. We find that an NDFO community would have the capacity to produce cell concentrations of up to 106 cells mL-1, which is comparable to microbes in Earth's oceans. However, only a concentration of <104 cells mL-1, due to organisms that inefficiently consume less than 10% of precipitating nitrate, would be consistent with the abundance of organics found at Gale. We also find that meteoritic sources of organics would likely be insufficient as a sole source for the Gale crater organics, which would require a separate source, such as abiotic hydrothermal or atmospheric production or possibly biological production from a slowly turning over chemotrophic community.

A Novel Abiotic Pathway for Phosphine Synthesis over Acidic Dust in Venus' Atmosphere.

Recent ground-based observations of Venus have detected a single spectral feature consistent with phosphine (PH3) in the middle atmosphere, a gas which has been suggested as a biosignature on rocky planets. The presence of PH3 in the oxidized atmosphere of Venus has not yet been explained by any abiotic process. However, state-of-the-art experimental and theoretical research published in previous works demonstrated a photochemical origin of another potential biosignature-the hydride methane-from carbon dioxide over acidic mineral surfaces on Mars. The production of methane includes formation of the HC · O radical. Our density functional theory (DFT) calculations predict an energetically plausible reaction network leading to PH3, involving either HC · O or H· radicals. We suggest that, similarly to the photochemical formation of methane over acidic minerals already discussed for Mars, the origin of PH3 in Venus' atmosphere could be explained by radical chemistry starting with the reaction of ·PO with HC·O, the latter being produced by reduction of CO2 over acidic dust in upper atmospheric layers of Venus by ultraviolet radiation. HPO, H2P·O, and H3P·OH have been identified as key intermediate species in our model pathway for phosphine synthesis.