Function-morphological investigations of fish inner ear otoliths as basis for interpretation of human space sickness

In man, altered gravity may lead to a vestibular dysfunction causing space motion sickness. A hypothesis was developed, according to which asymmetric inner ear statoliths might be the morphological basis of space sickness. The animal model, fish, revealed further information: inner ear "stone" (otolith) growth is dependent on the amplitude and the direction of gravity, regulated by a negative feedback mechanism. The present study was focused on the question, where the regulation centre of adaptive otolith growth may be situated. Therefore, the vestibular nerve was unilaterally transected in neonate swordtail fish (Xiphophorus helleri). As growth marker, the calcium tracer alizarin-complexone was used. It was found that otolith growth had ceased on the operated head sides indicating that the brain is significantly involved in regulating otolith growth. About 2 weeks after nerve transection, otoliths had regained normal growth, probably due to nerve regeneration. Concerning fish, it has now to be tested, if this regeneration is affected by altered gravity, e.g. in a long-term experiment on the International Space Station. Regarding mammals, it has to be proved if asymmetric statoliths are the basis of kinetosis and whether or not the mammalian brain has an effect on statolith growth in the course of compensating altered gravity.     

Survival of Bacillus pumilus spores for a prolonged period of time in real space conditions

To prevent forward contamination and maintain the scientific integrity of future life-detection missions, it is important to characterize and attempt to eliminate terrestrial microorganisms associated with exploratory spacecraft and landing vehicles. Among the organisms isolated from spacecraft-associated surfaces, spores of Bacillus pumilus SAFR-032 exhibited unusually high resistance to decontamination techniques such as UV radiation and peroxide treatment. Subsequently, B. pumilus SAFR-032 was flown to the International Space Station (ISS) and exposed to a variety of space conditions via the European Technology Exposure Facility (EuTEF). After 18 months of exposure in the EXPOSE facility of the European Space Agency (ESA) on EuTEF under dark space conditions, SAFR-032 spores showed 10-40% survivability, whereas a survival rate of 85-100% was observed when these spores were kept aboard the ISS under dark simulated martian atmospheric conditions. In contrast, when UV (>110?nm) was applied on SAFR-032 spores for the same time period and under the same conditions used in EXPOSE, a ～7-log reduction in viability was observed. A parallel experiment was conducted on Earth with identical samples under simulated space conditions. Spores exposed to ground simulations showed less of a reduction in viability when compared with the "real space" exposed spores (～3-log reduction in viability for "UV-Mars," and ～4-log reduction in viability for "UV-Space"). A comparative proteomics analysis indicated that proteins conferring resistant traits (superoxide dismutase) were present in higher concentration in space-exposed spores when compared to controls. Also, the first-generation cells and spores derived from space-exposed samples exhibited elevated UVC resistance when compared with their ground control counterparts. The data generated are important for calculating the probability and mechanisms of microbial survival in space conditions and assessing microbial contaminants as risks for forward contamination and in situ life detection.     

Survival of rock-colonizing organisms after 1.5 years in outer space

Cryptoendolithic microbial communities and epilithic lichens have been considered as appropriate candidates for the scenario of lithopanspermia, which proposes a natural interplanetary exchange of organisms by means of rocks that have been impact ejected from their planet of origin. So far, the hardiness of these terrestrial organisms in the severe and hostile conditions of space has not been tested over extended periods of time. A first long-term (1.5 years) exposure experiment in space was performed with a variety of rock-colonizing eukaryotic organisms at the International Space Station on board the European EXPOSE-E facility. Organisms were selected that are especially adapted to cope with the environmental extremes of their natural habitats. It was found that some-but not all-of those most robust microbial communities from extremely hostile regions on Earth are also partially resistant to the even more hostile environment of outer space, including high vacuum, temperature fluctuation, the full spectrum of extraterrestrial solar electromagnetic radiation, and cosmic ionizing radiation. Although the reported experimental period of 1.5 years in space is not comparable with the time spans of thousands or millions of years believed to be required for lithopanspermia, our data provide first evidence of the differential hardiness of cryptoendolithic communities in space.     

Trojans in habitable zones

With the aid of numerical experiments we examined the dynamical stability of fictitious terrestrial planets in 1:1 mean motion resonance with Jovian-like planets of extrasolar planetary systems. In our stability study of the so-called "Trojan" planets in the habitable zone, we used the restricted three-body problem with different mass ratios of the primary bodies. The application of the three-body problem showed that even massive Trojan planets can be stable in the 1:1 mean motion resonance. From the 117 extrasolar planetary systems only 11 systems were found with one giant planet in the habitable zone. Out of this sample set we chose four planetary systems--HD17051, HD27442, HD28185, and HD108874--for further investigation. To study the orbital behavior of the stable zone in the different systems, we used direct numerical computations (Lie Integration Method) that allowed us to determine the escape times and the maximum eccentricity of the fictitious "Trojan planets."     

Damage escape and repair in dried Chroococcidiopsis spp. from hot and cold deserts exposed to simulated space and martian conditions

The cyanobacterium Chroococcidiopsis, overlain by 3?mm of Antarctic sandstone, was exposed as dried multilayers to simulated space and martian conditions. Ground-based experiments were conducted in the context of Lichens and Fungi Experiments (EXPOSE-E mission, European Space Agency), which were performed to evaluate, after 1.5 years on the International Space Station, the survival of cyanobacteria (Chroococcidiopsis), lichens, and fungi colonized on Antarctic rock. The survival potential and the role played by protection and repair mechanisms in the response of dried Chroococcidiopsis cells to ground-based experiments were both investigated. Different methods were employed, including evaluation of the colony-forming ability, single-cell analysis of subcellular integrities based on membrane integrity molecular and redox probes, evaluation of the photosynthetic pigment autofluorescence, and assessment of the genomic DNA integrity with a PCR-based assay. Desiccation survivors of strain CCMEE 123 (coastal desert, Chile) were better suited than CCMEE 134 (Beacon Valley, Antarctica) to withstand cellular damage imposed by simulated space and martian conditions. Exposed dried cells of strain CCMEE 123 formed colonies, maintained subcellular integrities, and, depending on the exposure conditions, also escaped DNA damage or repaired the induced damage upon rewetting.     

The SOS-LUX-LAC-FLUORO-Toxicity-test on the International Space Station (ISS).

In the 21st century, an increasing number of astronauts will visit the International Space Station (ISS) for prolonged times. Therefore it is of utmost importance to provide necessary basic knowledge concerning risks to their health and their ability to work on the station and during extravehicular activities (EVA) in free space. It is the aim of one experiment of the German project TRIPLE-LUX (to be flown on the ISS) to provide an estimation of health risk resulting from exposure of the astronauts to the radiation in space inside the station as well as during extravehicular activities on one hand, and of exposure of astronauts to unavoidable or as yet unknown ISS-environmental genotoxic substances on the other. The project will (i) provide increased knowledge of the biological action of space radiation and enzymatic repair of DNA damage, (ii) uncover cellular mechanisms of synergistic interaction of microgravity and space radiation and (iii) examine the space craft milieu with highly specific biosensors. For these investigations, the bacterial biosensor SOS-LUX-LAC-FLUORO-Toxicity-test will be used, combining the SOS-LUX-Test invented at DLR Germany (Patent) with the commercially available LAC-FLUORO-Test. The SOS-LUX-Test comprises genetically modified bacteria transformed with the pBR322-derived plasmid pPLS-1. This plasmid carries the promoterless lux operon of Photobacterium leiognathi as a reporter element under control of the DNA-damage dependent SOS promoter of ColD as sensor element. This system reacts to radiation and other agents that induce DNA damages with a dose dependent measurable emission of bioluminescence of the transformed bacteria. The analogous LAC-FLUORO-Test has been developed for the detection of cellular responses to cytotoxins. It is based on the constitutive expression of green fluorescent protein (GFP) mediated by the bacterial protein expression vector pGFPuv (Clontech, Palo Alto, USA). In response to cytotoxic agents, this system reacts with a dose-dependent reduction of GFP-fluorescence. Currently, a fully automated miniaturized hardware system for the bacterial set up, which includes measurements of luminescence and fluorescence or absorption and the image analysis based evaluation is under development. During the first mission of the SOS-LUX-LAC-FLUORO-Toxicity-Test on the ISS, a standardized, DNA-damaging radiation source still to be determined will be used as a genotoxic inducer. A panel of recombinant Salmonella typhimurium strains carrying either the SOS-LUX plasmid or the fluorescence-mediating lac-GFPuv plasmid will be used to determine in parallel on one microplate the genotoxic and the cytotoxic action of the applied radiation in combination with microgravity. Either in addition to or in place of the fluorometric measurements of the cytotoxic agents, photometric measurements will simultaneously monitor cell growth, giving additional data on survival of the cells. The obtained data will be available on line during the TRIPLE-LUX mission time. Though it is the main goal during the TRIPLE-LUX mission to measure the radiation effect in microgravity, the SOS-LUX-LAC-FLUORO-Toxicity-test in principle is also applicable as a biomonitor for the detection and measurement of genotoxic substances in air or in the (recycled) water system on the ISS or on earth in general.     

Survival of spores of the UV-resistant Bacillus subtilis strain MW01 after exposure to low-earth orbit and simulated martian conditions: data from the space experiment ADAPT on EXPOSE-E

In the space experiment "Molecular adaptation strategies of microorganisms to different space and planetary UV climate conditions" (ADAPT), bacterial endospores of the highly UV-resistant Bacillus subtilis strain MW01 were exposed to low-Earth orbit (LEO) and simulated martian surface conditions for 559 days on board the European Space Agency's exposure facility EXPOSE-E, mounted outside the International Space Station. The survival of B. subtilis MW01 spores from both assays (LEO and simulated martian conditions) was determined by a colony-formation assay after retrieval. It was clearly shown that solar extraterrestrial UV radiation (λ≥110?nm) as well as the martian UV spectrum (λ≥200?nm) was the most deleterious factor applied; in some samples only a few spore survivors were recovered from B. subtilis MW01 spores exposed in monolayers. However, if shielded from solar irradiation, about 8% of MW01 spores survived in LEO conditions, and 100% survived in simulated martian conditions, compared to the laboratory controls. The results demonstrate the effect of shielding against the high inactivation potential of extraterrestrial solar UV radiation, which limits the chances of survival of even the highly UV-resistant strain of B. subtilis MW01 in the harsh environments of outer space and the martian surface.     

EXPOSE-E: an ESA astrobiology mission 1.5 years in space

The multi-user facility EXPOSE-E was designed by the European Space Agency to enable astrobiology research in space (low-Earth orbit). On 7 February 2008, EXPOSE-E was carried to the International Space Station (ISS) on the European Technology Exposure Facility (EuTEF) platform in the cargo bay of Space Shuttle STS-122 Atlantis. The facility was installed at the starboard cone of the Columbus module by extravehicular activity, where it remained in space for 1.5 years. EXPOSE-E was returned to Earth with STS-128 Discovery on 12 September 2009 for subsequent sample analysis. EXPOSE-E provided accommodation in three exposure trays for a variety of astrobiological test samples that were exposed to selected space conditions: either to space vacuum, solar electromagnetic radiation at >110?nm and cosmic radiation (trays 1 and 3) or to simulated martian surface conditions (tray 2). Data on UV radiation, cosmic radiation, and temperature were measured every 10?s and downlinked by telemetry. A parallel mission ground reference (MGR) experiment was performed on ground with a parallel set of hardware and samples under simulated space conditions. EXPOSE-E performed a successful 1.5-year mission in space.     

Biological space experiments for the simulation of Martian conditions: UV radiation and Martian soil analogues.

The survivability of resistant terrestrial microbes, bacterial spores of Bacillus subtilis, was investigated in the BIOPAN facility of the European Space Agency onboard of Russian Earth-orbiting FOTON satellites (BIOPAN I -III missions). The spores were exposed to different subsets of the extreme environmental parameters in space (vacuum, extraterrestrial solar UV, shielding by protecting materials like artificial meteorites). The results of the three space experiments confirmed the deleterious effects of extraterrestrial solar UV radiation which, in contrast to the UV radiation reaching the surface of the Earth, also contains the very energy-rich, short wavelength UVB and UVC radiation. Thin layers of clay, rock or meteorite material were shown to be only successful in UV-shielding, if they are in direct contact with the spores. On Mars the UV radiation climate is similar to that of the early Earth before the development of a protective ozone layer in the atmosphere by the appearance of the first aerobic photosynthetic bacteria. The interference of Martian soil components and the intense and nearly unfiltered Martian solar UV radiation with spores of B. subtilis will be tested with a new BIOPAN experiment, MARSTOX. Different types of Mars soil analogues will be used to determine on one hand their potential toxicity alone or in combination with solar UV (phototoxicity) and on the other hand their UV protection capability. Two sets of samples will be placed under different cut-off filters used to simulate the UV radiation climate of Mars and Earth. After exposure in space the survival of and mutation induction in the spores will be analyzed at the DLR, together with parallel samples from the corresponding ground control experiment performed in the laboratory. This experiment will provide new insights into the principal limits of life and its adaptation to environmental extremes on Earth or other planets which and will also have implications for the potential for the evolution and distribution of life.     

Resistance of bacterial endospores to outer space for planetary protection purposes--experiment PROTECT of the EXPOSE-E mission

Spore-forming bacteria are of particular concern in the context of planetary protection because their tough endospores may withstand certain sterilization procedures as well as the harsh environments of outer space or planetary surfaces. To test their hardiness on a hypothetical mission to Mars, spores of Bacillus subtilis 168 and Bacillus pumilus SAFR-032 were exposed for 1.5 years to selected parameters of space in the experiment PROTECT during the EXPOSE-E mission on board the International Space Station. Mounted as dry layers on spacecraft-qualified aluminum coupons, the "trip to Mars" spores experienced space vacuum, cosmic and extraterrestrial solar radiation, and temperature fluctuations, whereas the "stay on Mars" spores were subjected to a simulated martian environment that included atmospheric pressure and composition, and UV and cosmic radiation. The survival of spores from both assays was determined after retrieval. It was clearly shown that solar extraterrestrial UV radiation (λ≥110?nm) as well as the martian UV spectrum (λ≥200?nm) was the most deleterious factor applied; in some samples only a few survivors were recovered from spores exposed in monolayers. Spores in multilayers survived better by several orders of magnitude. All other environmental parameters encountered by the "trip to Mars" or "stay on Mars" spores did little harm to the spores, which showed about 50% survival or more. The data demonstrate the high chance of survival of spores on a Mars mission, if protected against solar irradiation. These results will have implications for planetary protection considerations.