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The waters that were present on early Mars may have been habitable. Characterising environments analogous to these waters and investigating the viability of their microbes under simulated martian chemical conditions is key to developing hypotheses on this habitability and potential biosignature formation. In this study, we examined the viability of microbes from the Anderton Brine Springs (United Kingdom) under simulated martian chemistries designed to simulate the chemical conditions of water that may have existed during the Hesperian. Associated changes in the fluid chemistries were also tested using inductively coupled plasma-optical emission spectroscopy (ICP-OES). The tested Hesperian fluid chemistries were shown to be habitable, supporting the growth of all of the Anderton Brine Spring isolates. However, inter and intra-generic variation was observed both in the ability of the isolates to tolerate more concentrated fluids and in their impact on the fluid chemistry. Therefore, whilst this study shows microbes from fluctuating brines can survive and grow in simulated martian water chemistry, further investigations are required to further define the potential habitability under past martian conditions.

The identification of geochemical bio-signatures is important for assessing whether life existed on early Mars. In this paper, experimental microbiology and thermochemical modelling were combined to identify potential inorganic bio-signatures for life detection on early Mars. An analogue mixed microbial community from an analogue terrestrial fluvio-lacustrine environment similar to an ancient lacustrine system at Gale Crater was used to study microbial dissolution of a basalt regolith simulant and the formation of bio-signatures over a short time frame (1°month) at 14°C, 2 bar. Microbial growth influenced element dissolution (Mg, Fe, Mn, Ca and K) and the formation of morphologies and Fe-Si amorphous layers on mineral surfaces. Thermochemical models were performed at 14°C, 2 bar; the results were compared with experimental data to predict bio-signatures that would occur over geological timescales. The pH was varied to simulate abiotic and biotic experimental conditions. Model results suggest that, at water to rock ratios of 100 to 38, a less complex secondary mineral assemblage forms during biotic dissolution compared to abiotic weathering. Carbonates, quartz, pyrite and hydroxyapatite form under biotic conditions, whereas in the abiotic system magnetite and phyllosilicates would also precipitate. These results could be used to distinguish between abiotic and biotic basalt weathering processes, aiding the interpretation of data from Mars exploration missions.

Water present on the surface of early Mars (>3.0 Ga) may have been habitable. Characterising analogue environments and investigating the aspects of their microbiome best suited for growth under simulated martian chemical conditions is key to understanding potential habitability. Experiments were conducted to investigate the viability of microbes from a Mars analogue environment, Colour Peak Springs (Axel Heiberg Island, Canadian High Arctic), under simulated martian chemistries. The fluid was designed to emulate waters thought to be typical of the late Noachian, in combination with regolith simulant material based on two distinct martian geologies. These experiments were performed with a microbial community from Colour Peak Springs sediment. The impact on the microbes was assessed by cell counting and 16S rRNA gene amplicon sequencing. Changes in fluid chemistries were tested using ICP-OES. Both chemistries were shown to be habitable, with growth in both chemistries. Microbial communities exhibited distinct growth dynamics and taxonomic composition, comprised of sulfur-cycling bacteria, represented by either sulfate-reducing or sulfur-oxidising bacteria, and additional heterotrophic halophiles. Our data support the identification of Colour Peak Springs as an analogue for former martian environments, with a specific subsection of the biota able to survive under more accurate proxies for martian chemistries.

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.

The Perseverance rover is meant to collect samples of the martian surface for eventual return to Earth. The headspace gas present over the solid samples within the sample tubes will be of significant scientific interest for what it reveals about the interactions of the solid samples with the trapped atmosphere and for what it will reveal about the martian atmosphere itself. However, establishing the composition of the martian atmosphere will require other dedicated samples. The headspace gas as the sole atmospheric sample is problematic for many reasons. The quantity of gas present within the sample tube volume is insufficient for many investigations, and there will be exchange between solid samples, headspace gas, and tube walls. Importantly, the sample tube materials and preparation were not designed for optimal Mars atmospheric gas collection and storage as they were not sent to Mars in a degassed evacuated state and have been exposed to both Earth's and Mars' atmospheres. Additionally, there is a risk of unconstrained seal leakage in transit back to Earth, which would allow fractionation of the sample (leak-out) and contamination (leak-in). The science return can be improved significantly (and, in some cases, dramatically) by adding one or more of several strategies listed here in increasing order of effectiveness and difficulty of implementation: (1) Having Perseverance collect a gas sample in an empty sample tube, (2) Collecting gas in a newly-designed, valved, sample-tube-sized vessel that is flown on either the Sample Fetch Rover (SFR) or the Sample Retrieval Lander (SRL), (3) Adding a larger (50-100 cc) dedicated gas sampling volume to the Orbiting Sample container (OS), (4) Adding a larger (50-100 cc) dedicated gas sampling volume to the OS that can be filled with compressed martian atmosphere., Astrobiology, 22 (S1), ISSN:1531-1074, ISSN:1557-8070

The surface of Mars cannot sustain liquid water today, but there is evidence water was present during the Noachian era. The transition of the martian climate from the wet Noachian to the dry Hesperian would have resulted in saline and sulfur rich surface waters . Terrestrial analogue environments that possess a chemistry like these proposed waters can be used to develop an understanding of organisms that could have persisted under such conditions. Here we present the chemistry and microbiome of the analogue environment Colour Peak, a sulfidic and saline spring system located within the Canadian High Arctic.\ud \ud In this study, molecular and geochemical techniques were used to investigate the sediment of the Colour Peak springs. Nucleic acids were extracted from the microbes in the sediments and the microbiome was characterised by the amplification and sequencing of 16S rRNA gene amplicons. The elemental composition of the fluids and sediment was determined by ICP-OES and compared with brines determined from the chemistry of the “Rocknest” sand sample at Yellowknife Bay, Gale Crater (Mars) by thermochemical modelling. Gibbs energy values were calculated from this fluid chemistry to identify potentially viable metabolisms. \ud \ud Analysis of the chemistries of the Colour Peak fluids confirmed a chemical composition like the thermochemically modelled fluid, with this justifying the classification of Colour Peak as an appropriate analogue environment to investigate the habitability of former martian aqueous environments. 16S rRNA gene profiling of the Colour Peak microbial community revealed it was dominated by bacteria associated with oxidation of reduced sulfur species and carbon dioxide fixation. Gibbs energy values calculated using the chemistry of the modelled martian fluid demonstrated that the oxidation of reduced sulfur species was also viable in this chemical environment under aerobic and anaerobic conditions. These results demonstrate that microbial sulfide oxidation is thermodynamically viable using both modelled and environmental proxies for former martian aqueous environments. \ud \ud This study highlights that metabolisms utilising the oxidation of reduced sulfur species could have been thermodynamically viable in ancient martian aqueous environments. Further work is needed to assess this proposed viability and the potential for unambiguous biosignature formation.

The martian surface cannot sustain liquid water today, but there is evidence water was present during the Noachian era. The transition of the martian climate into the Hesperian would have resulted in saline and sulfuric waters. Terrestrial analogue environments that possess a chemistry like these proposed waters can be used to develop an understanding of organisms that could have persisted. Here we present the chemistry and microbiome of Colour Peak, a sulfidic and saline spring system located within the Canadian High Arctic. \ud Nucleic acids were extracted from the microbes in the sediments and the microbiome was characterised by the amplification and sequencing of 16S rRNA gene amplicons. The elemental composition of the fluids and sediment was determined by ICP-OES and compared with brines determined from the chemistry of the “Rocknest” sample at Yellowknife Bay, Gale Crater (Mars) by thermochemical modelling. Gibbs energy values were calculated from this fluid chemistry to identify potentially viable metabolisms. \ud Analysis of the chemistries of the Colour Peak fluids confirmed a composition like the thermochemically modelled fluid, providing justification for the classification of Colour Peak as an appropriate analogue environment to investigate the habitability of former martian waters. Profiling of the Colour Peak microbial community revealed domination by bacteria associated with oxidation of reduced sulfur species and carbon dioxide fixation. Gibbs energy values calculated using the modelled martian fluid chemistry demonstrated that oxidation of reduced sulfur species was also viable in this chemical environment under aerobic and anaerobic conditions. These results demonstrate microbial sulfide oxidation is thermodynamically viable using both modelled and environmental proxies for former martian aqueous environments. \ud This study highlights that metabolisms utilising the oxidation of reduced sulfur species could have been thermodynamically viable in ancient martian aqueous environments. Further work is needed to test this viability and the subsequent potential for biosignature formation.

Habitability of martian waters would have been partially determined by the chemistry arising from interactions with martian lithologies. In this study, the habitability of groundwater chemistries (based on basaltic, iron- and sulfur-enriched lithologies) and the resulting variation in biosignatures was investigated, with microbes from anaerobic estuarine sediment used as an inoculum. The microbial community was monitored by cell counts and 16S rRNA gene profiling. Changes in fluid and precipitate chemistries were measured using ICP-OES and IC, with changes over geological timescales modelled using CHIM-XPT. The fluid chemistries were shown to be habitable, with distinct patterns in cell abundance and growth phases between the chemistries. However, the same genera dominated (Acetobacterium, Desulfovibrio and Desulfosporomusa) regardless of the initial fluid chemistry. In the biotic test group, changes in fluid chemistry were the same in the three chemistries, with an enhanced concentration of aluminium and iron and the removal of sulfate. However, geochemical modelling of the fluids under abiotic conditions over geological timescales revealed similar changes to those in the biotic test groups. Therefore, these samples require further analysis to assess whether we can identify any potentially unambiguous biosignatures that could develop between geologically distinct sites.

Microbes have multiple ways of producing energy. Which of these methods are possible depends on the chemistry of the environment the microbes are in (e.g. not enough of a metal or too much salt), with only specific methods working in certain environments. The same would be true of any waters that might continue to exist on Mars. To narrow down which methods of producing energy would be theoretically possible we simulated martian waters using a collection of minerals that are chemically similar to the chemistry measured by the Mars rover Curiosity in a crater on Mars. We added mud from an estuary to the simulated martian water and identified which microbes were able to grow. We then repeatedly transferred the growing microbes to fresh “martian” water to dilute out the nutrients from the mud. Over time we observed that most of the microbes from the mud have been lost but a few specific microbes were growing well. From this we hope to investigate changes in the chemistry of the water that happen because of these microbes, to try and identify specific chemistries that can be looked for by the future rover missions on Mars seeking evidence of life.