Experimental Identification of Potential Martian Biosignatures in Open and Closed Systems.

NASA's Perseverance and ESA's Rosalind Franklin rovers have the scientific goal of searching for evidence of ancient life on Mars. Geochemical biosignatures that form because of microbe-mineral interactions could play a key role in achieving this, as they can be preserved for millions of years on Earth, and the same could be true for Mars. Previous laboratory experiments have explored the formation of biosignatures under closed systems, but these do not represent the open systems that are found in natural martian environments, such as channels and lakes. In this study, we have conducted environmental simulation experiments using a global regolith simulant (OUCM-1), a thermochemically modelled groundwater, and an anaerobic microbial community to explore the formation of geochemical biosignatures within plausible open and closed systems on Mars. This initial investigation showed differences in the diversity of the microbial community developed after 28 days. In an open-system simulation (flow-through experiment), the acetogenic Acetobacterium (49% relative abundance) and the sulfate reducer Desulfosporomusa (43% relative abundance) were the dominant genera. Whereas in the batch experiment, the sulfate reducers Desulfovibrio, Desulfomicrobium, and Desulfuromonas (95% relative abundance in total) were dominant. We also found evidence of enhanced mineral dissolution within the flow-through experiment, but there was little evidence of secondary deposits in the presence of biota. In contrast, SiO2 and Fe deposits formed within the batch experiment with biota but not under abiotic conditions. The results from these initial experiments indicate that different geochemical biosignatures can be generated between open and closed systems, and therefore, biosignature formation in open systems warrants further investigation.

Habitability and Biosignature Formation in Simulated Martian Aqueous Environments.

Water present on early Mars is often assumed to have been habitable. In this study, experiments were performed to investigate the habitability of well-defined putative martian fluids and to identify the accompanying potential formation of biosignatures. Simulated martian environments were developed by combining martian fluid and regolith simulants based on the chemistry of the Rocknest sand shadow at Gale Crater. The simulated chemical environment was inoculated with terrestrial anoxic sediment from the Pyefleet mudflats (United Kingdom). These enrichments were cultured for 28 days and subsequently subcultured seven times to ensure that the microbial community was solely grown on the defined, simulated chemistry. The impact of the simulated chemistries on the microbial community was assessed by cell counts and sequencing of 16S rRNA gene profiles. Associated changes to the fluid and precipitate chemistries were established by using ICP-OES, IC, FTIR, and NIR. The fluids were confirmed as habitable, with the enriched microbial community showing a reduction in abundance and diversity over multiple subcultures relating to the selection of specific metabolic groups. The final community comprised sulfate-reducing, acetogenic, and other anaerobic and fermentative bacteria. Geochemical characterization and modeling of the simulant and fluid chemistries identified clear differences between the biotic and abiotic experiments. These differences included the elimination of sulfur owing to the presence of sulfate-reducing bacteria and more general changes in pH associated with actively respiring cells that impacted the mineral assemblages formed. This study confirmed that a system simulating the fluid chemistry of Gale Crater could support a microbial community and that variation in chemistries under biotic and abiotic conditions can be used to inform future life-detection missions.

Oligotrophic Growth of Nitrate-Dependent Fe2+-Oxidising Microorganisms Under Simulated Early Martian Conditions.

Nitrate-dependent Fe2+ oxidation (NDFO) is a microbially mediated process observed in many anaerobic, low-nutrient (oligotrophic) neutral-alkaline environments on Earth, which describes oxidation of Fe2+ to Fe3+ in tandem with microbial nitrate reduction. Evidence suggests that similar environments existed on Mars during the Noachian epoch (4.1-3.7 Ga) and in periodic, localised environments more recently, indicating that NDFO metabolism could have played a role in a potential early martian biosphere. In this paper, three NDFO microorganisms, Acidovorax sp. strain BoFeN1, Pseudogulbenkiania sp. strain 2002 and Paracoccus sp. strain KS1, were assessed for their ability to grow oligotrophically in simulated martian brines and in a minimal medium with olivine as a solid Fe2+ source. These simulant-derived media were developed from modelled fluids based on the geochemistry of Mars sample locations at Rocknest (contemporary Mars soil), Paso Robles (sulphur-rich soil), Haematite Slope (haematite-rich soil) and a Shergottite meteorite (common basalt). The Shergottite medium was able to support growth of all three organisms, while the contemporary Mars medium supported growth of Acidovorax sp. strain BoFeN1 and Pseudogulbenkiania sp. strain 2002; however, growth was not accompanied by significant Fe2+ oxidation. Each of the strains was also able to grow in oligotrophic minimal media with olivine as the sole Fe2+ source. Biomineralised cells of Pseudogulbenkiania sp. strain 2002 were identified on the surface of the olivine, representing a potential biosignature for NDFO microorganisms in martian samples. The results suggest that NDFO microorganisms could have thrived in early martian groundwaters under oligotrophic conditions, depending on the local lithology. This can guide missions in identifying palaeoenvironments of interest for biosignature detection. Indeed, biomineralised cells identified on the olivine surface provide a previously unexplored mechanism for the preservation of morphological biosignatures in the martian geological record.

Synthesis of Organosilicon Ligands for Europium (III) and Gadolinium (III) as Potential Imaging Agents.

The relaxivity of MRI contrast agents can be increased by increasing the size of the contrast agent and by increasing concentration of the bound gadolinium. Large multi-site ligands able to coordinate several metal centres show increased relaxivity as a result. In this paper, an "aza-type Michael" reaction is used to prepare cyclen derivatives that can be attached to organosilicon frameworks via hydrosilylation reactions. A range of organosilicon frameworks were tested including silsesquioxane cages and dimethylsilylbenzene derivatives. Michael donors with strong electron withdrawing groups could be used to alkylate cyclen on three amine centres in a single step. Hydrosilylation successfully attached these to mono-, di-, and tri-dimethylsilyl-substituted benzene derivatives. The europium and gadolinium complexes were formed and studied using luminescence spectroscopy and relaxometry. This showed the complexes to contain two bound water moles per lanthanide centre and T1 relaxation time measurements demonstrated an increase in relaxivity had been achieved, in particular for the trisubstituted scaffold 1,3,5-tris((pentane-sDO3A)dimethylsilyl)benzene-Gd3. This showed a marked increase in the relaxivity (13.1 r1p/mM-1s-1).

The Microbial Community of a Terrestrial Anoxic Inter-Tidal Zone: A Model for Laboratory-Based Studies of Potentially Habitable Ancient Lacustrine Systems on Mars.

Evidence indicates that Gale crater on Mars harboured a fluvio-lacustrine environment that was subjected to physio-chemical variations such as changes in redox conditions and evaporation with salinity changes, over time. Microbial communities from terrestrial environmental analogues sites are important for studying such potential habitability environments on early Mars, especially in laboratory-based simulation experiments. Traditionally, such studies have predominantly focused on microorganisms from extreme terrestrial environments. These are applicable to a range of Martian environments; however, they lack relevance to the lacustrine systems. In this study, we characterise an anoxic inter-tidal zone as a terrestrial analogue for the Gale crater lake system according to its chemical and physical properties, and its microbiological community. The sub-surface inter-tidal environment of the River Dee estuary, United Kingdom (53°21'15.40″ N, 3°10'24.95″ W) was selected and compared with available data from Early Hesperian-time Gale crater, and temperature, redox, and pH were similar. Compared to subsurface 'groundwater'-type fluids invoked for the Gale subsurface, salinity was higher at the River Dee site, which are more comparable to increases in salinity that likely occurred as the Gale crater lake evolved. Similarities in clay abundance indicated similar access to, specifically, the bio-essential elements Mg, Fe and K. The River Dee microbial community consisted of taxa that were known to have members that could utilise chemolithoautotrophic and chemoorganoheterotrophic metabolism and such a mixed metabolic capability would potentially have been feasible on Mars. Microorganisms isolated from the site were able to grow under environment conditions that, based on mineralogical data, were similar to that of the Gale crater's aqueous environment at Yellowknife Bay. Thus, the results from this study suggest that the microbial community from an anoxic inter-tidal zone is a plausible terrestrial analogue for studying habitability of fluvio-lacustrine systems on early Mars, using laboratory-based simulation experiments.

Nitrate-Dependent Iron Oxidation: A Potential Mars Metabolism.

This work considers the hypothetical viability of microbial nitrate-dependent Fe2+ oxidation (NDFO) for supporting simple life in the context of the early Mars environment. This draws on knowledge built up over several decades of remote and in situ observation, as well as recent discoveries that have shaped current understanding of early Mars. Our current understanding is that certain early martian environments fulfill several of the key requirements for microbes with NDFO metabolism. First, abundant Fe2+ has been identified on Mars and provides evidence of an accessible electron donor; evidence of anoxia suggests that abiotic Fe2+ oxidation by molecular oxygen would not have interfered and competed with microbial iron metabolism in these environments. Second, nitrate, which can be used by some iron oxidizing microorganisms as an electron acceptor, has also been confirmed in modern aeolian and ancient sediment deposits on Mars. In addition to redox substrates, reservoirs of both organic and inorganic carbon are available for biosynthesis, and geochemical evidence suggests that lacustrine systems during the hydrologically active Noachian period (4.1-3.7 Ga) match the circumneutral pH requirements of nitrate-dependent iron-oxidizing microorganisms. As well as potentially acting as a primary producer in early martian lakes and fluvial systems, the light-independent nature of NDFO suggests that such microbes could have persisted in sub-surface aquifers long after the desiccation of the surface, provided that adequate carbon and nitrates sources were prevalent. Traces of NDFO microorganisms may be preserved in the rock record by biomineralization and cellular encrustation in zones of high Fe2+ concentrations. These processes could produce morphological biosignatures, preserve distinctive Fe-isotope variation patterns, and enhance preservation of biological organic compounds. Such biosignatures could be detectable by future missions to Mars with appropriate instrumentation.

Determination of Geochemical Bio-Signatures in Mars-Like Basaltic Environments.

Bio-signatures play a central role in determining whether life existed on early Mars. Using a terrestrial basalt as a compositional analog for the martian surface, we applied a combination of experimental microbiology and thermochemical modeling techniques to identify potential geochemical bio-signatures for life on early Mars. Laboratory experiments were used to determine the short-term effects of biota on the dissolution of terrestrial basalt, and the formation of secondary alteration minerals. The chemoorganoheterotrophic bacterium, Burkholderia sp. strain B_33, was grown in a minimal growth medium with and without terrestrial basalt as the sole nutrient source. No growth was detected in the absence of the basalt. In the presence of basalt, during exponential growth, the pH decreased rapidly from pH 7.0 to 3.6 and then gradually increased to a steady-state of equilibrium of between 6.8 and 7.1. Microbial growth coincided with an increase in key elements in the growth medium (Si, K, Ca, Mg, and Fe). Experimental results were compared with theoretical thermochemical modeling to predict growth of secondary alteration minerals, which can be used as bio-signatures, over a geological timescale. We thermochemically modeled the dissolution of the basalt (in the absence of biota) in very dilute brine at 25°C, 1 bar; the pH was buffered by the mineral dissolution and precipitation reactions. Preliminary results suggested that at the water to rock ratio of 1 × 107, zeolite, hematite, chlorite, kaolinite, and apatite formed abiotically. The biotic weathering processes were modeled by varying the pH conditions within the model to adjust for biologic influence. The results suggested that, for a basaltic system, the microbially-mediated dissolution of basalt would result in "simpler" secondary alteration, consisting of Fe-hydroxide and kaolinite, under conditions where the abiotic system would also form chlorite. The results from this study demonstrate that, by using laboratory-based experiments and thermochemical modeling, it is possible to identify secondary alteration minerals that could potentially be used to distinguish between abiotic and biotic weathering processes on early Mars. This work will contribute to the interpretation of data from past, present, and future life detection missions to Mars.