Studying the effects of galactic cosmic radiation on astro- and microbiological model systems

In-depth knowledge regarding the biological effects of the radiation field in space is required for assessing the radiation risks in space. Within the last 50 years, space technology has provided tools for transporting terrestrial life beyond this protective magnetic field in order to study in situ responses to selected conditions of space (reviewed in Horneck et al., 2010). From a biological perspective applicable to simple and complex organisms (ranging from biomolecules and microorganisms to humans) various influential physical modifications such as increased radiation exposure were experienced onboard an orbiting spacecraft in low Earth orbit (LEO), out- and inside the International Space Station (ISS), orbiting Moon or on the way to other astrobiological-interesting targets (Mars or icy moons of Saturn or Jupiter). The majority of experiments on microorganisms in space were performed using Earth-orbiting robotic spacecraft, e.g., the Russian Foton satellites (FOTON) and the European Retrievable Carrier (EURECA), or human-tended spacecraft, such as space shuttles and space stations, e.g., MIR and ISS (reviewed in Nicholson, 2009; Nicholson et al., 2009; Horneck et al., 2010)

The microbial diversity of the Su Bentu cave, Italy and the influence of human exploration.

Introduction: The microbial diversity in the Su Bentu Cave (Sardinia, Italy) was investigated by means of Illumina MiSeq analysis. The hypogean environment is of great interest for astrobiological research as cave conditions may resemble those in extra-terrestrial regions. Furthermore, they hold high potential to identify novel, extremely adapted organisms to severely oligo-trophic habitats. However, the influence of human is not neglectable and in-depth investigations are needed to determine the impact of exploration on an otherwise mostly pristine ecosystem. The cave investigated in this study develops for several kilometres into the mountain, two hundred metres below the topographic surface and is characterized by a strong air circulation. Its structure is composed of huge passages carved in limestone where an ephemeral underground stream creates some lakes, close to which seven samples of visible calcite rafts, manganese deposits and moonmilk (a hydrated calcium carbonate speleothem), were sampled during an expedition in 2014. Other samples were re-trieved from a frequently used campsite and from some dry cave passages leading deeper into the cave

Carotenoid analysis of Halophilic Archaea by Resonance Raman spectroscopy

This is the publisher's version, also available electronically from "http://online.liebertpub.com".Recently, halite and sulfate evaporate rocks have been discovered on Mars by the NASA rovers, Spirit and Opportunity. It is reasonable to propose that halophilic microorganisms could have potentially flourished in these settings. If so, biomolecules found in microorganisms adapted to high salinity and basic pH environments on Earth may be reliable biomarkers for detecting life on Mars. Therefore, we investigated the potential of Resonance Raman (RR) spectroscopy to detect biomarkers derived from microorganisms adapted to hypersaline environments. RR spectra were acquired using 488.0 and 514.5 nm excitation from a variety of halophilic archaea, including Halobacterium salinarum NRC-1, Halococcus morrhuae, and Natrinema pallidum. It was clearly demonstrated that RR spectra enhance the chromophore carotenoid molecules in the cell membrane with respect to the various protein and lipid cellular components. RR spectra acquired from all halophilic archaea investigated contained major features at approximately 1000, 1152, and 1505 cm−1. The bands at 1505 cm−1 and 1152 cm−1 are due to in-phase C=C (ν1 ) and C–C stretching ( ν2 ) vibrations of the polyene chain in carotenoids. Additionally, in-plane rocking modes of CH3 groups attached to the polyene chain coupled with C–C bonds occur in the 1000 cm−1 region. We also investigated the RR spectral differences between bacterioruberin and bacteriorhodopsin as another potential biomarker for hypersaline environments. By comparison, the RR spectrum acquired from bacteriorhodopsin is much more complex and contains modes that can be divided into four groups: the C=C stretches (1600–1500 cm−1), the CCH in-plane rocks (1400–1250 cm−1), the C–C stretches (1250–1100 cm−1), and the hydrogen out-of-plane wags (1000–700 cm−1). RR spectroscopy was shown to be a useful tool for the analysis and remote in situ detection of carotenoids from halophilic archaea without the need for large sample sizes and complicated extractions, which are required by analytical techniques such as high performance liquid chromatography and mass spectrometry

Esa Caves: training astronauts for space exploration

The first spaceflight was several decades ago, and yet extraterrestrial exploration is only at the beginning and has mainly been carried out by robotic probes and rovers sent to extraterrestrial planets and deep space. In the future human extraterrestrial exploration will take place and to get ready for long periods of permanence in space, astronauts are trained during long duration missions on the International Space Station (ISS). To prepare for such endeavours, team training activities are performed in extreme environments on Earth, as isolated deserts, base camps on Antarctica, or stations built on the bottom of the sea, trying to simulate the conditions and operations of space. Space agencies are also particularly interested in the search of signs of life forms in past or present extreme natural environments, such as salt lakes in remote deserts, very deep ocean habitats, submarine volcanic areas, sulphuric acid caves, and lava tubes. One natural environment that very realistically mimics an extraterrestrial exploration habitat is the cave. Caves are dark, remote places, with constant temperature, many logistic problems and stressors (isolation, communication and supply difficulties, physical barriers), and their exploration requires discipline, teamwork, technical skills and a great deal of behavioural adaptation. For this reason, since 2008 the European Space Agency has carried out training activities in the subterranean environment and the CAVES project is one of those training courses, probably the most realistic one. CAVES stands for Cooperative Adventure for Valuing and Exercising human behaviour and performance Skills, and is meant as a multidisciplinary multicultural team exploration mission in a cave. It has been developed by ESA in the past few years (2008-2011) and is open for training of astronauts of the ISS Partner Space Agencies (USA, Russia, Japan, Canada, and Europe). Astronauts are first trained for 5 days to explore, document and survey a karst system, then take on a cave exploration mission for 6 days underground. A team of expert cave instructors, a Human Behaviour and Performance facilitator, scientists and video reporters, ensure that all tasks are performed in complete safety and guides all these astronauts\u27 activities. During the underground mission the astronauts\u27 technical competences are challenged (exploring, surveying, taking pictures), their human behaviour and decision-making skills are debriefed, and they are required to carry out an operational programme which entails performing scientific tasks and testing equipment, similarly to what they are required to do on the ISS. The science program includes environmental and air circulation monitoring, mineralogy, microbiology, chemical composition of waters, and search for life forms adapted to the cavern environment. The CAVES 2012 Course will be explained and the first interesting scientific results will be presented

Exploring Andean High-Altitude Lake Extremophiles through Advanced Proteotyping

Quickly identifying and characterizing isolates from extreme environments is currently challenging while very important to explore the Earth′s biodiversity. As these isolates may, in principle, be distantly related to known species, techniques are needed to reliably identify the branch of life to which they belong. Proteotyping these environmental isolates by tandem mass spectrometry offers a rapid and cost-effective option for their identification using their peptide profiles. In this study, we document the first high-throughput proteotyping approach for environmental extremophilic and halophilic isolates. Microorganisms were isolated from samples originating from high-altitude Andean lakes (3700–4300 m a.s.l.) in the Chilean Altiplano, which represent environments on Earth that resemble conditions on other planets. A total of 66 microorganisms were cultivated and identified by proteotyping and 16S rRNA gene amplicon sequencing. Both the approaches revealed the same genus identification for all isolates except for three isolates possibly representing not yet taxonomically characterized organisms based on their peptidomes. Proteotyping was able to indicate the presence of two potentially new genera from the families of Paracoccaceae and Chromatiaceae/Alteromonadaceae, which have been overlooked by 16S rRNA amplicon sequencing approach only. The paper highlights that proteotyping has the potential to discover undescribed microorganisms from extreme environments

Biological Contamination Prevention for Outer Solar System Moons of Astrobiological Interest: What Do We Need to Know?

To ensure that scientific investments in space exploration are not compromised by terrestrial contamination of celestial bodies, special care needs to be taken to preserve planetary conditions for future astrobiological exploration. Significant effort has been made and is being taken to address planetary protection in the context of inner Solar System exploration. In particular for missions to Mars, detailed internationally accepted guidelines have been established. For missions to the icy moons in the outer Solar System, Europa and Enceladus, the planetary protection requirements are so far based on a probabilistic approach and a conservative estimate of poorly known parameters. One objective of the European Commission-funded project, Planetary Protection of Outer Solar System, was to assess the existing planetary protection approach, to identify inherent knowledge gaps, and to recommend scientific investigations necessary to update the requirements for missions to the icy moons

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

MEXEM – Mars Exposed Extremophile Mixture – a space experiment to investigate the capability of anaerobic organisms to survive on Mars

Assessing the habitability of Mars and detecting life, if it ever existed there, depends on knowledge of whether the combined environmental stresses experienced on Mars are compatible with life as we know it and whether a record of that life could ever be detected. So far, only few investigations were performed to understand the combined effect of different environmental stresses on survival and growth of anaerobic and extremophilic organisms. In the space experiment MEXEM (formerly known as MASE-in-SPACE) the hypothesis will be tested that selected terrestrial organisms, enrichment cultures and original samples from extreme Mars-analogue environments on Earth are able to withstand the Martian environ- mental stress factors due to their highly effective cellular and molecular adaptation and repair mechanisms. In addition, artificially fossilized and aged isolates from Mars-analogue environments on Earth will be examined and assessed with respect to their suitability for biosigna- ture identification. MEXEM samples will be (i) oxygen-depleted natural sediment samples, (ii) natural sediments spiked with selected, defined strains representative for the respective analogue site, (iii) individual (facultative) anaerobic / micro-aerophilic species including ciliates and viruses, (iv) defined mixtures of these biological entities, (v) isolated strains from samples collected inside the ISS and (vi) artificially fossilized isolates from the natural environments. Most of these samples and isolates were obtained from Mars-analogue envi- ronments on Earth in the frame of the EC funded project MASE (Grant Agreement 607297) and from the space experiment EXTREMOPHILES (PI C. Moissl-Eichinger). MEXEM will be flown outside on the ISS in the new exobiology facility ESA is building now. It offers the possibility to simulate of the martian environment, in particular the martian UV cl e, which cannot be done in the lab, but also martian atmosphere and pressure in LEO

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

ANALOGUE SAMPLES IN AN EUROPEAN SAMPLE CURATION FACILITY - THE EURO-CARES PROJECT.

The objective of the H2020-funded EURO-CARES project (grant agreement n° 640190) was to create a roadmap for the implementation of a European Extraterrestrial Sample Curation Facility (ESCF) that would be suitable for the curation of samples from all possible return missions likely over the next few decades, i.e. from the Moon, asteroids and Mars. The return of extraterrestrial samples brought to Earth will require specific storage conditions and handling procedures, in particular for those coming from Mars. For practical reasons and sterility concerns it might be necessary for such a facility to have its own collection of analogue samples permitting the testing of storage conditions, and to develop protocols for sample prepartion and analyses. Within the framework of the EURO-CARES project, we havecreated a list of the different types of samples that would be relevant for such a curation facility. The facility will be used for receiving and opening of the returned sample canisters, as well as for handling and preparation of the returned samples. Furthermore, it will provide some analysis of the returned samples, i.e. early sample characterisation, and is expected to provide longterm storage of the returned samples. Each of these basic functions requires special equipment. Equipment, handling protocols and long-term storage conditions will strongly depend on the characteristics of the materials, and on whether returned samples are from the Moon, Mars or an asteroidal body. Therefore the different types and aspects of analogue samples one need to be considered, i.e. the nature of the materials, which analogues are needed for what purpose, what mass is needed, and how should the analogue samples be stored within the facility. We distinguished five different types of anologue samples: analogue (s.s.), witness plate, voucher specimen, reference sample, and standard. Analogues are materials that have one or more physical or chemical properties similar to Earth-returned extraterrestrial samples. Reference samples are well-characterised materials with known physical and chemical properties used for testing. They may not necessarily be the same materials as the analogues defined above. Standards are internationally recognised, homogeneous materials with known physical and chemical properties that are used for calibration. They can also be used as reference samples in certain circumstances. They may be made of natural materials but are often produced artificially. A voucher specimen is a duplicate of materials used at any stage during sample acquisition, storage, transport, treatment etc., e.g. spacecraft materials (including solar panels), lubricants, glues, gloves, saws, drills, and others. In addition, Earth landing site samples (from the touch down site) would be necessary in case of doubtful analysis, even if normally this type of contamination is not expected. Finally, a witness plate is defined as material left in an area where work is being done to detect any biological, particulate, chemical, and/or organic contamination. It is a spatial and temporal document of what happens in the work area. Analogue materials could be solids (including ices), liquids or gases. These could contain biological (extant and/or exinct) and/or organic components. They could be natural materials, e.g. rocks or minerals, or could be manufactured, such as mixtures of different components, which may be biologically and/or organically doped. Analogues with appropriate sample size and nature will be well-suited for testing and training of sample handling procedures, and for transport protocols. The training of science and curation teams also requires reference samples and standards. Long-term storage needs special witness plates and voucher specimes. Developing and testing sample preparation protocols needs all sample types