Xeropreservation of functionalized lipid biomarkers in hyperarid soils in the Atacama Desert.

Our understanding of long-term organic matter preservation comes mostly from studies in aquatic systems. In contrast, taphonomic processes in extremely dry environments are relatively understudied and are poorly understood. We investigated the accumulation and preservation of lipid biomarkers in hyperarid soils in the Yungay region of the Atacama Desert. Lipids from seven soil horizons in a 2.5 m vertical profile were extracted and analyzed using GC-MS and LC-MS. Diagnostic functionalized lipids and geolipids were detected and increased in abundance and diversity with depth. Deeper clay units contain fossil organic matter (radiocarbon dead) that has been protected from rainwater since the onset of hyperaridity. We show that these clay units contain lipids in an excellent state of structural preservation with functional groups and unsaturated bonds in carbon chains. This indicates that minimal degradation of lipids has occurred in these soils since the time of their deposition between >40,000 and 2 million years ago. The exceptional structural preservation of biomarkers is likely due to the long-term hyperaridity that has minimized microbial and enzymatic activity, a taphonomic process we term xeropreservation (i.e. preservation by drying). The degree of biomarker preservation allowed us to reconstruct major changes in ecology in the Yungay region that reflect a shift in hydrological regime from wet to dry since the early Quaternary. Our results suggest that hyperarid environments, which comprise 7.5% of the continental landmass, could represent a rich and relatively unexplored source of paleobiological information on Earth.

Active microbiota persist in dry permafrost and active layer from Elephant Head, Antarctica.

Dry permafrost is a challenging environment for microbial life due to cold, dry, and often oligotrophic conditions. In 2016, Elephant Head, Antarctica, was confirmed as the second site on Earth to contain dry permafrost. It is geographically distinct from the McMurdo Dry Valleys where dry permafrost has been studied previously. Here, we present the first study of the microbial activity, diversity, and functional potential of Elephant Head dry permafrost. Microbial activity was measured using radiorespiration assays with radiolabeled acetate as a carbon source at 5, 0, and -5°C. Low, but detectable, rates of microbial activity were measured in some samples at 0 and -5°C. This is distinct from previous studies of McMurdo Dry Valley dry permafrost which concluded that dry permafrost represents a cold-arid limit to life on the planet. The isolation of cold-adapted organisms from these soils, including one capable of subzero growth, further supports that the Elephant Head dry active layer and dry permafrost harbor viable microbial life, which may be active in situ. Metagenomic, 16S rRNA gene, and internal transcribed spacer and amplicon sequencing identified similar microbial communities to other Antarctic and cold environments. The Elephant Head microbial community appears to be adapted for survival in cold, dry, and oligotrophic conditions based on the presence of cold adaptation and stress response genes in the metagenomes. Together, our results show that dry permafrost environments do not exclude active microbial life at subzero temperatures, suggesting that the cold, dry soils of Mars may also not be as inhospitable as previously thought.

Quantifying Global Origin-Diagnostic Features and Patterns in Biotic and Abiotic Acyclic Lipids for Life Detection.

Lipids are a geologically robust class of organics ubiquitous to life as we know it. Lipid-like soluble organics are synthesized abiotically and have been identified in carbonaceous meteorites and on Mars. Ascertaining the origin of lipids on Mars would be a profound astrobiological achievement. We enumerate origin-diagnostic features and patterns in two acyclic lipid classes, fatty acids (i.e., carboxylic acids) and acyclic hydrocarbons, by collecting and analyzing molecular data reported in over 1500 samples from previously published studies of terrestrial and meteoritic organics. We identify 27 combined (15 for fatty acids, 12 for acyclic hydrocarbons) molecular patterns and structural features that can aid in distinguishing biotic from abiotic synthesis. Principal component analysis (PCA) demonstrates that multivariate analyses of molecular features (16 for fatty acids, 14 for acyclic hydrocarbons) can potentially indicate sample origin. Terrestrial lipids are dominated by longer straight-chain molecules (C4-C34 fatty acids, C14-C46 acyclic hydrocarbons), with predominance for specific branched and unsaturated isomers. Lipid-like meteoritic soluble organics are shorter, with random configurations. Organic solvent-extraction techniques are most commonly reported, motivating the design of our novel instrument, the Extractor for Chemical Analysis of Lipid Biomarkers in Regolith (ExCALiBR), which extracts lipids while preserving origin-diagnostic features that can indicate biogenicity.

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.

Exploring the evidence of Middle Amazonian aquifer sedimentary outburst residues in a Martian chaotic terrain.

The quest for past Martian life hinges on locating surface formations linked to ancient habitability. While Mars' surface is considered to have become cryogenic ~3.7 Ga, stable subsurface aquifers persisted long after this transition. Their extensive collapse triggered megafloods ~3.4 Ga, and the resulting outflow channel excavation generated voluminous sediment eroded from the highlands. These materials are considered to have extensively covered the northern lowlands. Here, we show evidence that a lacustrine sedimentary residue within Hydraotes Chaos formed due to regional aquifer upwelling and ponding into an interior basin. Unlike the northern lowland counterparts, its sedimentary makeup likely consists of aquifer-expelled materials, offering a potential window into the nature of Mars' subsurface habitability. Furthermore, the lake's residue's estimated age is ~1.1 Ga (~3.2 Ga post-peak aquifer drainage during the Late Hesperian), enhancing the prospects for organic matter preservation. This deposit's inferred fine-grained composition, coupled with the presence of coexisting mud volcanoes and diapirs, suggest that its source aquifer existed within abundant subsurface mudstones, water ice, and evaporites, forming part of the region's extremely ancient (~ 4 Ga) highland stratigraphy. Our numerical models suggest that magmatically induced phase segregation within these materials generated enormous water-filled chambers. The meltwater, originating from varying thermally affected mudstone depths, could have potentially harbored diverse biosignatures, which could have become concentrated within the lake's sedimentary residue. Thus, we propose that Hydraotes Chaos merits priority consideration in future missions aiming to detect Martian biosignatures.

0.25 Ga Salt Deposits Preserve Signatures of Habitable Conditions and Ancient Lipids.

Polygonal features in a ∼250 million-year-old Permian evaporitic deposit were investigated for their geological and organic content to test the hypothesis that they could preserve the signature of ancient habitable conditions and biological activity. Investigations on evaporitic rock were carried out as part of the MIne Analog Research (MINAR) project at Boulby Mine, the United Kingdom. The edges of the polygons have a higher clay content and contain higher abundances of minerals such as quartz and microcline, and clays such as illite and chlorite, compared with the interior of polygons, suggesting that the edges were preferred locations for the accumulation of weathering products during their formation. The mineral content and its strontium isotope ratio suggest that the material is from continental weathering at the borders of the Permian Zechstein Sea. The edges of the polygons contain material with mean δ13C and δ15N values of -20.8 and 5.3, respectively. Lipids, including alkanes and hopanes, were extracted from the interior and edges of the polygons, which are inferred to represent organic material entrained in the evaporites when they were formed. The presence of long-chain alkanes (C20-C35) that lack a carbon preference, low abundances of C23-C29 hopanes, and lack of marine, evaporitic, or thermal maturity indicators show that lipid biomarkers were, at least in part, potentially derived from a continental source and have not undergone significant thermal maturation since deposition. Lipid extractions using weak acids revealed significantly more lipids than those without acid, potentially indicating that encapsulation was not the only type of preservation mechanism occurring in Boulby salts. These data demonstrate the potential for ancient evaporites and their polygons to preserve information on local geological conditions, ancient habitability, and evidence of life. The data show that analogous martian evaporitic deposits are good targets for future life detection missions and the investigation of ancient martian habitability.

The Oldest Highlands of Mars May Be Massive Dust Fallout Deposits.

The oldest terrains of Mars are cratered landscapes, in which extensive valleys and basins are covered by ubiquitous fluvial plains. One current paradigm maintains that an impact-generated megaregolith underlies these sediments. This megaregolith was likely largely generated during the Early Noachian (~4.1 to ~3.94 Ga) when most Martian impact basins formed. We examined the geologic records of NW Hellas and NW Isidis, which include this epoch's most extensive circum-basin outcrops. Here, we show that these regions include widespread, wind-eroded landscapes, crater rims eroded down by several hundred meters, pitted plains, and inverted fluvial and crater landforms. These surfaces exhibit few fresh craters, indicating geologically recent wind erosion. The deep erosion, topographic inversions, and an absence of dunes on or near talus across these regions suggest that sediments finer than sand compose most of these highland materials. We propose that basin-impact-generated hurricane-force winds created sediment-laden atmospheric conditions, and that muddy rains rapidly settled suspended sediments to construct extensive Early Noachian highlands. The implied high abundance of fine-grained sediments before these impacts suggests large-scale glacial silt production and supports the previously proposed Noachian "icy highlands" hypothesis. We suggest that subglacial meltwater interactions with the sedimentary highlands could have promoted habitability, particularly in clay strata.

Draft Genome Sequence from a Putative New Genus and Species in the Family Methanoregulaceae Isolated from the Anoxic Basin of Lake Untersee in East Antarctica.

Here, we report the draft genome sequence for a new putative genus and species in the Methanoregulaceae family, whose members are generally slow-growing rod-shaped or coccoid methanogenic archaea. The information on this sediment-dwelling organism sheds light on the prokaryotes inhabiting isolated, deep, and extremely cold methane-rich environments.

Recovery of Fatty Acids from Mineralogic Mars Analogs by TMAH Thermochemolysis for the Sample Analysis at Mars Wet Chemistry Experiment on the Curiosity Rover.

The Mars Curiosity rover carries a diverse instrument payload to characterize habitable environments in the sedimentary layers of Aeolis Mons. One of these instruments is Sample Analysis at Mars (SAM), which contains a mass spectrometer that is capable of detecting organic compounds via pyrolysis gas chromatography mass spectrometry (py-GC-MS). To identify polar organic molecules, the SAM instrument carries the thermochemolysis reagent tetramethylammonium hydroxide (TMAH) in methanol (hereafter referred to as TMAH). TMAH can liberate fatty acids bound in macromolecules or chemically bound monomers associated with mineral phases and make these organics detectable via gas chromatography mass spectrometry (GC-MS) by methylation. Fatty acids, a type of carboxylic acid that contains a carboxyl functional group, are of particular interest given their presence in both biotic and abiotic materials. This work represents the first analyses of a suite of Mars-analog samples using the TMAH experiment under select SAM-like conditions. Samples analyzed include iron oxyhydroxides and iron oxyhydroxysulfates, a mixture of iron oxides/oxyhydroxides and clays, iron sulfide, siliceous sinter, carbonates, and shale. The TMAH experiments produced detectable signals under SAM-like pyrolysis conditions when organics were present either at high concentrations or in geologically modern systems. Although only a few analog samples exhibited a high abundance and variety of fatty acid methyl esters (FAMEs), FAMEs were detected in the majority of analog samples tested. When utilized, the TMAH thermochemolysis experiment on SAM could be an opportunity to detect organic molecules bound in macromolecules on Mars. The detection of a FAME profile is of great astrobiological interest, as it could provide information regarding the source of martian organic material detected by SAM.

Constraints on the Metabolic Activity of Microorganisms in Atacama Surface Soils Inferred from Refractory Biomarkers: Implications for Martian Habitability and Biomarker Detection.

Dryness is one of the main environmental challenges to microbial survival. Understanding the threshold of microbial tolerance to extreme dryness is relevant to better constrain the environmental limits of life on Earth and critically evaluate long-term habitability models of Mars. Biomolecular proxies for microbial adaptation and growth were measured in Mars-like hyperarid surface soils in the Atacama Desert that experience only a few millimeters of precipitation per decade, and in biologically active soils a few hundred kilometers away that experience two- to fivefold more precipitation. Diversity and abundance of lipids and other biomolecules decreased with increasing dryness. Cyclopropane fatty acids (CFAs), which are indicative of adaptive response to environmental stress and growth in bacteria, were only detected in the wetter surface soils. The ratio of trans to cis isomers of an unsaturated fatty acid, another bacterial stress indicator, decreased with increasingly dry conditions. Aspartic acid racemization ratios increased from 0.01 in the wetter soils to 0.1 in the driest soils, which is indicative of racemization rates comparable to de novo biosynthesis over long timescales (∼10,000 years). The content and integrity of stress proteins profiled by immunoassays were additional indicators that biomass in the driest soils is not recycled at significant levels. Together, our results point to minimal or no in situ microbial growth in the driest surface soils of the Atacama, and any metabolic activity is likely to be basal for cellular repair and maintenance only. Our data add to a growing body of evidence that the driest Atacama surface soils represent a threshold for long-term habitability (i.e., growth and reproduction). These results place constraints on the potential for extant life on the surface of Mars, which is 100-1000 times drier than the driest regions in the Atacama. Key Words: Atacama Desert-Dryness-Growth-Habitability-Biomarker-Mars. Astrobiology 18, 955-966.