Perchlorate-specific proteomic stress responses of Debaryomyces hansenii could enable microbial survival in Martian brines.

If life exists on Mars, it would face several challenges including the presence of perchlorates, which destabilize biomacromolecules by inducing chaotropic stress. However, little is known about perchlorate toxicity for microorganisms on the cellular level. Here, we present the first proteomic investigation on the perchlorate-specific stress responses of the halotolerant yeast Debaryomyces hansenii and compare these to generally known salt stress adaptations. We found that the responses to NaCl and NaClO4 -induced stresses share many common metabolic features, for example, signalling pathways, elevated energy metabolism, or osmolyte biosynthesis. Nevertheless, several new perchlorate-specific stress responses could be identified, such as protein glycosylation and cell wall remodulations, presumably in order to stabilize protein structures and the cell envelope. These stress responses would also be relevant for putative life on Mars, which-given the environmental conditions-likely developed chaotropic defence strategies such as stabilized confirmations of biomacromolecules or the formation of cell clusters.

Transitory microbial habitat in the hyperarid Atacama Desert.

Traces of life are nearly ubiquitous on Earth. However, a central unresolved question is whether these traces always indicate an active microbial community or whether, in extreme environments, such as hyperarid deserts, they instead reflect just dormant or dead cells. Although microbial biomass and diversity decrease with increasing aridity in the Atacama Desert, we provide multiple lines of evidence for the presence of an at times metabolically active, microbial community in one of the driest places on Earth. We base this observation on four major lines of evidence: (i) a physico-chemical characterization of the soil habitability after an exceptional rain event, (ii) identified biomolecules indicative of potentially active cells [e.g., presence of ATP, phospholipid fatty acids (PLFAs), metabolites, and enzymatic activity], (iii) measurements of in situ replication rates of genomes of uncultivated bacteria reconstructed from selected samples, and (iv) microbial community patterns specific to soil parameters and depths. We infer that the microbial populations have undergone selection and adaptation in response to their specific soil microenvironment and in particular to the degree of aridity. Collectively, our results highlight that even the hyperarid Atacama Desert can provide a habitable environment for microorganisms that allows them to become metabolically active following an episodic increase in moisture and that once it decreases, so does the activity of the microbiota. These results have implications for the prospect of life on other planets such as Mars, which has transitioned from an earlier wetter environment to today's extreme hyperaridity.

Deliquescence-induced wetting and RSL-like darkening of a Mars analogue soil containing various perchlorate and chloride salts.

Recurring slope lineae (RSL) are flow-like features on Mars characterized by a local darkening of the soil thought to be generated by the formation and flow of liquid brines. One possible mechanism responsible for forming these brines could be the deliquescence of salts present in the Martian soil. We show that the JSC Mars-1a analogue soil undergoes a darkening process when salts dispersed in the soil deliquesce, but forming continuous liquid films and larger droplets takes much longer than previously assumed. Thus, RSL may not necessarily require concurrent flowing liquid water/brine or a salt-recharge mechanism, and their association with gullies may be the result of previously flowing water and deposited salts during an earlier warmer and wetter period. In addition, our results show that electrical conductivity measurements correlate well with the deliquescence rates and provide better overall characterization than either Raman spectroscopy or estimates based on deliquescence relative humidity.

Mechanisms of Evolutionary Innovation Point to Genetic Control Logic as the Key Difference Between Prokaryotes and Eukaryotes.

The evolution of life from the simplest, original form to complex, intelligent animal life occurred through a number of key innovations. Here we present a new tool to analyze these key innovations by proposing that the process of evolutionary innovation may follow one of three underlying processes, namely a Random Walk, a Critical Path, or a Many Paths process, and in some instances may also constitute a "Pull-up the Ladder" event. Our analysis is based on the occurrence of function in modern biology, rather than specific structure or mechanism. A function in modern biology may be classified in this way either on the basis of its evolution or the basis of its modern mechanism. Characterizing key innovations in this way helps identify the likelihood that an innovation could arise. In this paper, we describe the classification, and methods to classify functional features of modern organisms into these three classes based on the analysis of how a function is implemented in modern biology. We present the application of our categorization to the evolution of eukaryotic gene control. We use this approach to support the argument that there are few, and possibly no basic chemical differences between the functional constituents of the machinery of gene control between eukaryotes, bacteria and archaea. This suggests that the difference between eukaryotes and prokaryotes that allows the former to develop the complex genetic architecture seen in animals and plants is something other than their chemistry. We tentatively identify the difference as a difference in control logic, that prokaryotic genes are by default 'on' and eukaryotic genes are by default 'off.' The Many Paths evolutionary process suggests that, from a 'default off' starting point, the evolution of the genetic complexity of higher eukaryotes is a high probability event.

How Many Biochemistries Are Available To Build a Cell?

The manual experimental evolution of the bacterium Escherichia coli allowed the design of a noncanonical genetic code in which complete replacement of the endogenous building block tryptophan (left) by an exogenous one based on a thienylpyrrole (right) was achieved after 506 days of continuous culturing.

Proposed Missions to Collect Samples for Analyzing Evidence of Life in the Venusian Atmosphere.

The recent and still controversial claim of phosphine detection in the venusian atmosphere has reignited consideration of whether microbial life might reside in its cloud layers. If microbial life were to exist within Venus' cloud deck, these microorganisms would have to be multi-extremophiles enclosed within the cloud aerosol particles. The most straightforward approach for resolving the question of their existence is to obtain samples of the cloud particles and analyze their interior. While developing technology has made sophisticated in situ analysis possible, more detailed information could be obtained by examining samples with instrumentation in dedicated ground-based facilities. Ultimately, therefore, Venus Cloud-level Sample Return Missions will likely be required to resolve the question of whether living organisms exist in the clouds of Venus. Two multiphase mission concepts are currently under development for combining in situ analyses with a sample return component. The Venus Life Finder architecture proposes collection of cloud particles in a compartment suspended from a balloon that floats for weeks at the desired altitude, while the Novel solUtion for Venus explOration and Lunar Exploitation (NUVOLE) concept involves a glider that cruises within the cloud deck for 1200 km collecting cloud aerosol particles through the key regions of interest. Both architectures propose a rocket-driven ascent with the acquired samples transported to a high venusian orbit as a prelude to returning to Earth or the Moon. Both future conceptual missions with their combined phases will contribute valuable information relative to the habitability of the clouds at Venus, but their fulfillment is decades away. We suggest that, in the meantime, a simplification of a glider cloud-level sample collection scenario could be accomplished in a shorter development time at a lower cost. Even if the cloud particles are not organic and show no evidence of living organisms, they would reveal critical insights about the natural history and evolution of Venus.

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.

Persistent microbial communities in hyperarid subsurface habitats of the Atacama Desert: Insights from intracellular DNA analysis.

Desert environments constitute one of the largest and yet most fragile ecosystems on Earth. Under the absence of regular precipitation, microorganisms are the main ecological component mediating nutrient fluxes by using soil components, like minerals and salts, and atmospheric gases as a source for energy and water. While most of the previous studies on microbial ecology of desert environments have focused on surface environments, little is known about microbial life in deeper sediment layers. Our study is extending the limited knowledge about microbial communities within the deeper subsurface of the hyperarid core of the Atacama Desert. By employing intracellular DNA extraction and subsequent 16S rRNA sequencing of samples collected from a soil pit in the Yungay region of the Atacama Desert, we unveiled a potentially viable microbial subsurface community residing at depths down to 4.20 m. In the upper 80 cm of the playa sediments, microbial communities were dominated by Firmicutes taxa showing a depth-related decrease in biomass correlating with increasing amounts of soluble salts. High salt concentrations are possibly causing microbial colonization to cease in the lower part of the playa sediments between 80 and 200 cm depth. In the underlying alluvial fan deposits, microbial communities reemerge, possibly due to gypsum providing an alternative water source. The discovery of this deeper subsurface community is reshaping our understanding of desert soils, emphasizing the need to consider subsurface environments in future explorations of arid ecosystems.

Noachian and more recent phyllosilicates in impact craters on Mars.

Hundreds of impact craters on Mars contain diverse phyllosilicates, interpreted as excavation products of preexisting subsurface deposits following impact and crater formation. This has been used to argue that the conditions conducive to phyllosilicate synthesis, which require the presence of abundant and long-lasting liquid water, were only met early in the history of the planet, during the Noachian period (> 3.6 Gy ago), and that aqueous environments were widespread then. Here we test this hypothesis by examining the excavation process of hydrated minerals by impact events on Mars and analyzing the stability of phyllosilicates against the impact-induced thermal shock. To do so, we first compare the infrared spectra of thermally altered phyllosilicates with those of hydrated minerals known to occur in craters on Mars and then analyze the postshock temperatures reached during impact crater excavation. Our results show that phyllosilicates can resist the postshock temperatures almost everywhere in the crater, except under particular conditions in a central area in and near the point of impact. We conclude that most phyllosilicates detected inside impact craters on Mars are consistent with excavated preexisting sediments, supporting the hypothesis of a primeval and long-lasting global aqueous environment. When our analyses are applied to specific impact craters on Mars, we are able to identify both pre- and postimpact phyllosilicates, therefore extending the time of local phyllosilicate synthesis to post-Noachian times.

Life Unknown: Preliminary Scheme for a Magnetotrophic Organism.

No magnetotrophic organism on Earth is known to use magnetic fields as an energy source or the storage of information. However, a broad diversity of life forms is sensitive to magnetic fields and employs them for orientation and navigation, among other purposes. If the magnetic field strength were much larger, such as that on planets around neutron stars or magnetars, metabolic energy could be obtained from these magnetic fields in principle. Here, we introduce three hypothetical models of magnetotrophic organisms that obtain energy via the Lorentz force. Even if an organism uses magnetic fields only as an energy source, but otherwise is relying on biochemistry, this organism would be by definition a magnetotrophic form of life as we do not know it.

The Hypothesis of a "Living Pulse" in Cells.

Motility is a great biosignature and its pattern is characteristic for specific microbes. However, motion does also occur within the cell by the myriads of ongoing processes within the cell and the exchange of gases and nutrients with the outside environment. Here, we propose that the sum of these processes in a microbial cell is equivalent to a pulse in complex organisms and suggest a first approach to measure the "living pulse" in microorganisms. We emphasize that if a "living pulse" can be shown to exist, it would have far-reaching applications, such as for finding life in extreme environments on Earth and in extraterrestrial locations, as well as making sure that life is not present where it should not be, such as during medical procedures and in the food processing industry.

Variations in climate habitability parameters and their effect on Earth's biosphere during the Phanerozoic Eon.

Essential insights on the characterization and quality of a detectable biosphere are gained by analyzing the effects of its environmental parameters. We compiled environmental and biological properties of the Phanerozoic Eon from various published data sets and conducted a correlation analysis to assess variations in parameters relevant to the habitability of Earth's biosphere. We showed that environmental parameters such as oxygen, global average surface temperatures, runoff rates and carbon dioxide are interrelated and play a key role in the changes of biomass and biodiversity. We showed that there were several periods with a highly thriving biosphere, with one even surpassing present day biodiversity and biomass. Those periods were characterized by increased oxygen levels and global runoff rates, as well as moderate global average surface temperatures, as long as no large or rapid positive and/or negative temperature excursions occurred. High oxygen contents are diagnostic of biomass production by continental plant life. We find that exceptionally high oxygen levels can at least in one instance compensate for decreased relative humidities, providing an even more habitable environment compared to today. Beyond Earth, these results will help us to understand how environmental parameters affect biospheres on extrasolar planets and guide us in our search for extraterrestrial life.

Response of cyanobacterial mats to ambient phosphate fluctuations: phosphorus cycling, polyphosphate accumulation and stoichiometric flexibility.

Cyanobacterial mats inhabit a variety of aquatic habitats, including the most extreme environments on Earth. They can thrive in a wide range of phosphorus (P) levels and are thus important players for ecosystem primary production and P cycling at the sediment-water interface. Polyphosphate (polyP), the major microbial P storage molecule, is assigned a critical role in compensating for phosphate fluctuations in planktonic cyanobacteria, but little is known about potentially analogous mechanisms of mat-forming cyanobacteria. To investigate acclimation strategies of cyanobacterial mats to fluctuating phosphate concentrations, laboratory batch experiments were conducted, in which the cosmopolitan mat-forming, marine cyanobacterium Sodalinema stali was exposed to low dissolved P concentrations, followed by a P pulse. Our results show that the cyanobacteria dynamically adjusted cellular P content to ambient phosphate concentrations and that they had accumulated polyP during periods of high phosphate availability, which was subsequently recycled to sustain growth during phosphate scarcity. However, following the depletion of dispensable cellular P sources, including polyP, we observed a reallocation of P contained in DNA into polyP, accompanied by increasing alkaline phosphatase activity. This suggests a change of the metabolic focus from growth towards maintenance and the attempt to acquire organic P, which would be naturally contained in the sediment. P overplus uptake following a simulated P pulse further suggests that Sodalinema-dominated mats exhibit elaborated mechanisms to cope with severe P fluctuations to overcome unfavourable environmental conditions, and potentially modulate critical P fluxes in the aquatic cycle.

Eolian erosion of polygons in the Atacama Desert as a proxy for hyper-arid environments on Earth and beyond.

Polygonal networks occur on various terrestrial and extraterrestrial surfaces holding valuable information on the pedological and climatological conditions under which they develop. However, unlike periglacial polygons that are commonly used as an environmental proxy, the information that polygons in the hyper-arid Atacama Desert can provide is little understood. To promote their use as a proxy, we investigated a polygonal network within an inactive channel that exhibits uncommonly diverse surface morphologies and mineral compositions, using geochemical and remote sensing techniques. Our findings show that the polygons belong to a continuous network of the same genetic origin. Their differences result from post-formational differential eolian erosion up to 50 cm depth, exposing indurated subsurface horizons rich in sulfate or nitrate and chloride. Their location in an ancient channel could lead to the misinterpretation of fluvial polygon erosion, however, we find no such signs but evidence for aqueous resurfacing of microtopography by fog and minimal rainwater infiltration. Our findings extend the use of polygons as proxies in the Atacama Desert, indicating saline soils and hyper-arid conditions. We conclude that this example of polygon erosion can guide future polygon research, especially regarding the use of erosional surfaces on Earth and beyond to gain valuable subsurface insights.

Whole genome sequencing of cyanobacterium Nostoc sp. CCCryo 231-06 using microfluidic single cell technology.

The Nostoc sp. strain CCCryo 231-06 is a cyanobacterial strain capable of surviving under extreme conditions and thus is of great interest for the astrobiology community. The knowledge of its complete genome sequence would serve as a guide for further studies. However, a major concern has been placed on the effects of contamination on the quality of sequencing data without a reference genome. Here, we report the use of microfluidic technology combined with single cell sequencing and de novo assembly to minimize the contamination and recover the complete genome of the Nostoc strain CCCryo 231-06 with high quality. 100% of the whole genome was recovered with all contaminants removed and a strongly supported phylogenetic tree. The data reported can be useful for comparative genomics for phylogenetic and taxonomic studies. The method used in this work can be applied to studies that require high-quality assemblies of genomes of unknown microorganisms.

Non-random genetic alterations in the cyanobacterium Nostoc sp. exposed to space conditions.

Understanding the impact of long-term exposure of microorganisms to space is critical in understanding how these exposures impact the evolution and adaptation of microbial life under space conditions. In this work we subjected Nostoc sp. CCCryo 231-06, a cyanobacterium capable of living under many different ecological conditions, and also surviving in extreme ones, to a 23-month stay at the International Space Station (the Biology and Mars Experiment, BIOMEX, on the EXPOSE-R2 platform) and returned it to Earth for single-cell genome analysis. We used microfluidic technology and single cell sequencing to identify the changes that occurred in the whole genome of single Nostoc cells. The variant profile showed that biofilm and photosystem associated loci were the most altered, with an increased variant rate of synonymous base pair substitutions. The cause(s) of these non-random alterations and their implications to the evolutionary potential of single bacterial cells under long-term cosmic exposure warrants further investigation.

Biosignature stability in space enables their use for life detection on Mars.

Two rover missions to Mars aim to detect biomolecules as a sign of extinct or extant life with, among other instruments, Raman spectrometers. However, there are many unknowns about the stability of Raman-detectable biomolecules in the martian environment, clouding the interpretation of the results. To quantify Raman-detectable biomolecule stability, we exposed seven biomolecules for 469 days to a simulated martian environment outside the International Space Station. Ultraviolet radiation (UVR) strongly changed the Raman spectra signals, but only minor change was observed when samples were shielded from UVR. These findings provide support for Mars mission operations searching for biosignatures in the subsurface. This experiment demonstrates the detectability of biomolecules by Raman spectroscopy in Mars regolith analogs after space exposure and lays the groundwork for a consolidated space-proven database of spectroscopy biosignatures in targeted environments.

Fundamental Science and Engineering Questions in Planetary Cave Exploration.

Nearly half a century ago, two papers postulated the likelihood of lunar lava tube caves using mathematical models. Today, armed with an array of orbiting and fly-by satellites and survey instrumentation, we have now acquired cave data across our solar system-including the identification of potential cave entrances on the Moon, Mars, and at least nine other planetary bodies. These discoveries gave rise to the study of planetary caves. To help advance this field, we leveraged the expertise of an interdisciplinary group to identify a strategy to explore caves beyond Earth. Focusing primarily on astrobiology, the cave environment, geology, robotics, instrumentation, and human exploration, our goal was to produce a framework to guide this subdiscipline through at least the next decade. To do this, we first assembled a list of 198 science and engineering questions. Then, through a series of social surveys, 114 scientists and engineers winnowed down the list to the top 53 highest priority questions. This exercise resulted in identifying emerging and crucial research areas that require robust development to ultimately support a robotic mission to a planetary cave-principally the Moon and/or Mars. With the necessary financial investment and institutional support, the research and technological development required to achieve these necessary advancements over the next decade are attainable. Subsequently, we will be positioned to robotically examine lunar caves and search for evidence of life within Martian caves; in turn, this will set the stage for human exploration and potential habitation of both the lunar and Martian subsurface.

The Case (or Not) for Life in the Venusian Clouds.

The possible detection of the biomarker of phosphine as reported by Greaves et al. in the Venusian atmosphere stirred much excitement in the astrobiology community. While many in the community are adamant that the environmental conditions in the Venusian atmosphere are too extreme for life to exist, others point to the claimed detection of a convincing biomarker, the conjecture that early Venus was doubtlessly habitable, and any Venusian life might have adapted by natural selection to the harsh conditions in the Venusian clouds after the surface became uninhabitable. Here, I first briefly characterize the environmental conditions in the lower Venusian atmosphere and outline what challenges a biosphere would face to thrive there, and how some of these obstacles for life could possibly have been overcome. Then, I discuss the significance of the possible detection of phosphine and what it means (and does not mean) and provide an assessment on whether life may exist in the temperate cloud layer of the Venusian atmosphere or not.

Evolution of default genetic control mechanisms.

We present a model of the evolution of control systems in a genome under environmental constraints. The model conceptually follows the Jacob and Monod model of gene control. Genes contain control elements which respond to the internal state of the cell as well as the environment to control expression of a coding region. Control and coding regions evolve to maximize a fitness function between expressed coding sequences and the environment. The model was run 118 times to an average of 1.4∙106 'generations' each with a range of starting parameters probed the conditions under which genomes evolved a 'default style' of control. Unexpectedly, the control logic that evolved was not significantly correlated to the complexity of the environment. Genetic logic was strongly correlated with genome complexity and with the fraction of genes active in the cell at any one time. More complex genomes correlated with the evolution of genetic controls in which genes were active ('default on'), and a low fraction of genes being expressed correlated with a genetic logic in which genes were biased to being inactive unless positively activated ('default off' logic). We discuss how this might relate to the evolution of the complex eukaryotic genome, which operates in a 'default off' mode.