Detecting biochemical evidence for life with the signs of life detector (solid) in an anaerobic microorganism under fossilization conditions

The definitive detection of biosignatures in the context of astrobiological missions to Mars is not without difficulty. Could it be possible to detect biomarkers from an extinct form of life in a very ancient material? The traces of some microorganisms can be well preserved thanks to rapid mineralization of the organisms and cementation of the sediments in which they occur [1]. Thus biosignatures could be indicators of either extant or extinct life, the search for which is one of the main objectives of Mars exploration [1]. The central motivation of the MASE project (Mars Analogues for Space Exploration) is to gain knowledge about the habitability of Mars by the study of the adaptation of anaerobic life forms to extreme environments, their environmental context, and the methods used to detect their biosignatures. Within this background a fundamental target of MASE project is to improve and optimize methods for biosignature detection in samples with low biomass from certain Mars analogue sites. In this context we applied antibody multiarray competitive immunoassay to follow the evolution of specific biochemical signatures from a culture under fossilization conditions. An antibody multiarray competitive immunoassay for the simultaneous detection of compounds of a wide range of molecular sizes or whole spores and cells [2] [3] has revealed as suitable option to achieve this MASE purpose. It consists in a rapid strategy to detect a huge set of different epitopes in extracted samples by a sandwich multiarray immunoassay in a slide or LDChip (Life Detector Chip) where huge range of different antibodies are coated. In this report, we present the results from an experiment in which we followed the biochemical signatures from a growing culture of an isolate of Yersinia sp. in fresh media and in a culture growing under fossilization conditions in silica and gypsum. A decrease in the signal of relative fluorescence of antibody-antigen binding (biomarkers detected) is observed when comparing an untreated Yersinia sp. culture and those induced to mineralization at different time points

Mars Analogues for space exploration - from anaerobic field site to culture collection

Astrobiology seeks to understand the limits of life and to determine the physiology of organisms in order to be able to better assess the potential habitability of other worlds and improve our ability to assay them for the presence of life. To successfully achieve this we require representative microorganisms from environments on Earth that in physical and/or chemical conditions approximate to extraterrestrial environments. The most challenging of these environments with respect to the sample collection and follow on isolation and cultivation of microorganisms are anaerobic environments. Here we describe a systematic approach to this challenge and aim to provide a guideline for future fieldwork and sampling campaigns. We selected a number of anaerobic environments based on characteristics that make them analogous to past and present locations on Mars (Icelandic lakes, sulfidic springs, deep hypersaline environments, acidic iron-rich environments, and permafrost). We implemented a culturing approach to enrich organisms from these environments under anaerobic conditions using a defined medium that would allow for all organisms to be grown under identical culturing conditions m future physiological comparisons. We then isolated anaerobic microorganisms, carried out a study of their basic physiology and deposited these organisms in the DSMZ (Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH) culture collection to make them available to astrobiologists and microbiologists. This project represents the first attempt to implement a coordinated effort from the selection of extraterrestrial analog sites through to the isolation and the characterisation of organisms and their deposition within a culture collection

Potential for fossilization of an extremotolerant bacterium isolated from a past mars analog environment

In the context of astrobiological missions to Mars, the key question is what biosignatures to search for and how? lndigenous Martian organisms, if they existed or still exist, can be classified as extremophile per se. Following this precept the FP7-funded European MASE project (Mars Analogues for Space Exploration} is investigating various aspects of anaerobic life under Mars' extreme envrionmental conditions, including the potential for preservation over long geological time periods of certain strains. In this contribution, we report on the mineralisation and preservation of Yersinia sp. in silica and gypsum, two minerals that have been reported on Mars, in cold and anaerobic conditions, similar to Martian conditions. The organism, polyextremotolerant bacterium Yersinia sp. MASE-LG-1 (hereafter named Yersinia. sp.) was isolated from the lcelandic Graenavatn Lake, an acidic (pH3), cold and oligotrophic volcanic crater lake. These organisms have a strong tolerance to diverse Mars-like stresses (Rettberg et al., 2015). We also studied the effect of physiological status on mineralisation by exposing Yersinia to two common stresses thought to have increased du ring Mars history, desiccation and radiation. The mineralisation process has been studied using microbiological (microbial viability), morphological (scanning and transmission electron microscopy), biochemical (GC-MS, Rock-Eval) and spectroscopic (FTIR and RAMAN spectroscopy) methodologies. Based on these approaches, the potential of mineralised Yersinia sp. cells to be preserved over geological time scales is also discussed. Salient results include the fact that fossilisation in gypsum solutions is slower than in silica; not all cells were mineralised, even after 6-months in the fossilising solutions, although the FTIR, Raman and SOLID biomarker signatures were lost by this time period; Rock-Eval analysis suggests that the kerogen in the fossilised strain may not survive preservation over long geological periods, although carbon molecules preserved in fossil microbial traces up to ~3.45 Ga have been detected in the rock record

Metabolie response of Yersinia MASE-LGl to osmotic stress and ionizing radiation

The MASE (Mars Analogues for space exploration) project intends to gain deeper insights into the habitability of Mars by searching for anaerobic extremophiles in Mars analogue environments on Earth like the cold sulfidic springs in Germany, the deep-subsurface salt mine in UK, the iron-rich Rio Tinto and the cold acidic lake Graenavatn in lceland. From the latter, the MASE team isolated a Yersinia sp. strain. The surface of Mars is known to host deposits of magnesium and iron sulfates, suggesting that liquid water on that planet might contain high concentrations of sulfates. Halites have also been identified. Therefore, of significance to astrobiology and understanding the habitability of Mars is to understand the microbial response to sulfate and chloride salt exposure in combination with the ubiquitous ionizing radiation in the near-surface of Mars

Defining the boarders of extreme life: The study of microbial communities and their settings gives insights into Mars analogue life within the MASE project

The search for life beyond Earth is challenging and requires, as a prerequisite, intensive research on microbial life in similar, extreme environments on Earth. Mars analogue sites are characterised by e.g. anoxic conditions, organic compound limitation, low temperatures, high salinity or presence of oxidising compounds, and consequently represent the chemical and physical borders of life as we know it. The analysis of microorganisms withstanding such conditions is embedded in the European Commission-funded MASE (Mars Analogues for Space Exploration; (http://mase.esf.org/) project. Combining a broad spectrum of interdisciplinary expertise, the European project members aim at a better understanding of habitability, microbial lifestyles and biomarker preservation in Mars analogues. For the first time, the selected sites (e.g. salt mine, sulfidic springs) have undergone a profound analysis of their microbial communities on various levels, including vast cultivation of anaerobic microorganisms and molecular screening. In this work, we applied propidium monoazide in order to distinguish between cells with intact membrane (considered as viable) and dead cells on molecular stage, followed by DNA extraction, and amplicon-sequencing of the archaeal and bacterial 16S rRNA genes. The geochemistry of the sites was comprehensively investigated (i.e. elemental analysis, amino acid chirality, minerology), to determine triggers for microbial community composition. We aim to set up a model of potential metabolism reactions based on the different setting conditions and compare it with microbiome data. Consequently, we will obtain insights into the prerequisites of possible extra-terrestrial life forms and into their lifestyles, which may enable them to thrive under most extreme conditions