F. Gomez

Development and Structure of Eukaryotic Biofilms in an Extreme Acidic Environment, Rio Tinto (SW, Spain)

An in situ colonization assay was performed to study the early stages of biofilm formation in Río Tinto (SW, Spain), an extremely acidic environment (pH ca. 2). Eukaryotic assemblages were monitored at monthly intervals for 1 year. Diversity, colonization rates, and seasonal variations were analyzed. Structural features of naturally grown biofilms were explored by light and scanning electron microscopy in backscattered electron mode. A total of 14 taxa were recognized as constituents of the eukaryotic assemblages. The eukaryotic communities were dissimilar at the different sampling sites. The lowest diversity was found at the most extreme locations, in terms of pH and heavy metal concentrations. The biofilms were mainly formed by species from the genera Dunaliella and Cyanidium. Two genera of filamentous algae, Zygnemopsis and Klebsormidium, were principally responsible for the variability in the cell number throughout the year. These species appear in June to decrease almost completely between October and November. In contrast, the number of heterotrophic flagellates and ciliates remained constant throughout the year. The microcolonization sequence showed an initial accumulation of amorphous particles composed of bacteria and inorganic grains of minerals. By the end of the second month, the organic matrix was also populated by fungi, bacteria, and a few eukaryotic heterotrophs such as amoebae and small flagellates. Diatoms only showed significant colonization in regions where mycelial matrices were first established. Flagellated green algae such as Dunaliella or Chlamydomonas as well as Euglena were also present at the very beginning of the biofilm development, although in low numbers (<100 cells cm–2). After the flagellated cells, sessile species of algae such Chlorella or Cyanidium appeared. Filamentous algae were the last species to colonize the biofilms. Most of the naturally grown biofilms were found to be structures composed of different species organized in different layers separated, probably by extracellular polymeric substances, although more analysis should be done in this regard. The possible implications of the biofilm structure in the adaptation to this extreme habitat are discussed.

Beyond Chloride Brines: Variable Metabolomic Responses in the Anaerobic Organism Yersinia intermedia MASE-LG-1 to NaCl and MgSO4 at Identical Water Activity

Growth in sodium chloride (NaCl) is known to induce stress in non-halophilic microorganisms leading to effects on the microbial metabolism and cell structure. Microorganisms have evolved a number of adaptations, both structural and metabolic, to counteract osmotic stress. These strategies are well-understood for organisms in NaCl-rich brines such as the accumulation of certain organic solutes (known as either compatible solutes or osmolytes). Less well studied are responses to ionic environments such as sulfate-rich brines which are prevalent on Earth but can also be found on Mars. In this paper, we investigated the global metabolic response of the anaerobic bacterium Yersinia intermedia MASE-LG-1 to osmotic salt stress induced by either magnesium sulfate (MgSO4) or NaCl at the same water activity (0.975). Using a non-targeted mass spectrometry approach, the intensity of hundreds of metabolites was measured. The compatible solutes L-asparagine and sucrose were found to be increased in both MgSO4 and NaCl compared to the control sample, suggesting a similar osmotic response to different ionic environments. We were able to demonstrate that Yersinia intermedia MASE-LG-1 accumulated a range of other compatible solutes. However, we also found the global metabolic responses, especially with regard to amino acid metabolism and carbohydrate metabolism, to be salt-specific, thus, suggesting ion-specific regulation of specific metabolic pathways.

Mineralization and Preservation of an extremotolerant Bacterium Isolated from an Early Mars Analog Environment

The artificial mineralization of a polyresistant bacterial strain isolated from an acidic, oligotrophic lake was carried out to better understand microbial (i) early mineralization and (ii) potential for further fossilisation. Mineralization was conducted in mineral matrixes commonly found on Mars and Early-Earth, silica and gypsum, for 6 months. Samples were analyzed using microbiological (survival rates), morphological (electron microscopy), biochemical (GC-MS, Microarray immunoassay, Rock-Eval) and spectroscopic (EDX, FTIR, RAMAN spectroscopy) methods. We also investigated the impact of physiological status on mineralization and long-term fossilisation by exposing cells or not to Mars-related stresses (desiccation and radiation). Bacterial populations remained viable after 6 months although the kinetics of mineralization and cell-mineral interactions depended on the nature of minerals. Detection of biosignatures strongly depended on analytical methods, successful with FTIR and EDX but not with RAMAN and immunoassays. Neither influence of stress exposure, nor qualitative and quantitative changes of detected molecules were observed as a function of mineralization time and matrix. Rock-Eval analysis suggests that potential for preservation on geological times may be possible only with moderate diagenetic and metamorphic conditions. The implications of our results for microfossil preservation in the geological record of Earth as well as on Mars are discussed.

Lack of correlation of desiccation and radiation tolerance in microorganisms from diverse extreme environments tested under anoxic conditions

Abstract Four facultative anaerobic and two obligate anaerobic bacteria were isolated from extreme environments (deep subsurface halite mine, sulfidic anoxic spring, mineral-rich river) in the frame MASE (Mars Analogues for Space Exploration) project. The isolates were investigated under anoxic conditions for their survivability after desiccation up to 6 months and their tolerance to ionizing radiation up to 3000 Gy. The results indicated that tolerances to both stresses are strain-specific features. Yersinia intermedia MASE-LG-1 showed a high desiccation tolerance but its radiation tolerance was very low. The most radiation-tolerant strains were Buttiauxella sp. MASE-IM-9 and Halanaerobium sp. MASE-BB-1. In both cases, cultivable cells were detectable after an exposure to 3 kGy of ionizing radiation, but cells only survived desiccation for 90 and 30 days, respectively. Although a correlation between desiccation and ionizing radiation resistance has been hypothesized for some aerobic microorganisms, our data showed that there was no correlation between tolerance to desiccation and ionizing radiation, suggesting that the physiological basis of both forms of tolerances is not necessarily linked. In addition, these results indicated that facultative and obligate anaerobic organisms living in extreme environments possess varied species-specific tolerances to extremes.