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Long-term human missions to space require methods for sustaining the health and safety of the crew. Therefore, microbiological monitoring and reduction of contamination is necessary. Microbial biofilms are of special interest because they can cause damage to spaceflight equipment and are difficult to eliminate due to their increased resistance to antibiotics and disinfectants. The introduction of antimicrobial surfaces for medical, pharmaceutical, and industrial purposes has shown a unique potential for reducing and preventing biofilm formation. The European Space Agency (ESA) launched the BIOFILMS-project, which is evaluating the effect of microgravity on biofilm formation on non-inhibitory surfaces, such as steel, and on antimicrobial metal surfaces. These surfaces are composed out of different metals with and without nanostructures. For the project, human-relevant bacterial strains were selected. One of the selected strains is Staphylococcus capitis subsp. capitis K1-2-2-23, which was isolated from the International Space Station (ISS) on V2A steel and is forming strong biofilms. A comparison of this strain to the DSMZ type strain revealed several differences that include colony pigmentation, increased growth, and tolerance to desiccation. Furthermore, tests with S. capitis subsp. capitis strain K1-2-2-23 within the BIOFILMS hardware revealed that the hardware is biocompatible and allowed biofilm formation on steel surfaces. Additionally, the use of pure copper and brass surfaces inside the hardware led to a significant reduction in growth and biofilm formation, whereby nanostructured copper surfaces were observed to be more effective. The obtained results lay the foundation for the BIOFILMS spaceflight project that will be conducted aboard the International Space Station and may provide suitable antimicrobial surfaces for spaceflight purposes.

Gravity is the most consistent physical parameter to which all life has so far been subordinated and has not changed from the beginning of evolution to the present day. Since Apollo 16 the Gram-positive bacterium Bacillus subtilis has been used to study the influence of gravity on vegetative cells, spores and biofilms and still it remains unclear to what extent biofilm formation is influenced by the loss of gravity. As a model, B. subtilis can be used to investigate the intrinsic resistance of biofilms under space conditions. Biofilms are a challenge in medicine and in industry and with increasing space activities and manned long-term missions or even moon bases, new insights into the behavior of biofilms under altered gravity conditions are becoming increasingly important. We used B. subtilis NCIB 3610 as spore and biofilm-forming model organism as well as biofilm deficient mutants to investigate the influence of simulated microgravity (sim-µg) on structural and resistance properties. For the simulation, a fast-rotating 2-D clinostat was used in combination with a standardized method for growing and harvesting biofilms on filter membranes to achieve highly reproducible results. By comparing biofilm growth under normal gravitational conditions (1g) and sim-µg, the results of profilometry showed no significant changes in biofilm topography. Scanning electron microscopic images of biofilm cross-sections and transmission microscopic analyses showed differences in the internal biofilm structure. Typical biofilm properties such as matrix development in the basal regions of biofilms were increased in sim-µg. Growth and resistance properties of biofilms grown under sim-µg showed small differences, but hydrophobicity values were significantly increased under simulated microgravity. Spores isolated after sim-µg treatment showed homogeneous germination behavior in contrast to 1g spores and tended to germinate more spontaneously.

The exposure to the space radiation environment remains a major limiting factor for human long-duration space missions and permanent presence in space habitats due to its high biological effectiveness and the difficulties to effectively shield the radiation. The next decade in human spaceflight will be characterized by a continuous presence of human beings in Low Earth Orbit (LEO), by their return to the Moon and by longer stays in a Moon orbit on the Lunar Orbital Platform - Gateway (LOP-G). These endeavors are also performed to prepare for a human Mars mission. Also, plans for permanent presence of humans on the Moon are assuming shape. To support and enable these missions, ongoing improvement of radiation dosimetry for accurate and continuous monitoring and the development of shielding approaches including personal shielding equipment are necessary. In a cooperative project, a personal protection vest will be tested during the first Orion mission using female phantoms. This will be the first determination of depth dose distribution beyond LEO. The organ doses which are essential for space radiation risk assessment will be derived from the dose distribution in the phantoms. For protection from high-dose exposure during solar flares, a warning system and a suitable radiation shelter are required. In the last decades, the cancer risk induced by exposure to galactic cosmic rays was in the focus of attention. Numerous animal experiments and mechanistic studies were performed to derive the relative biological effectiveness of heavy ions to induce cancer and to elucidate the mechanisms and patterns specific to heavy ions. E.g., heavy ions activate the Nuclear Factor κB (NF-κB) pathway which is involved in inflammatory responses with very high efficiency [1-3]. Radiation quality factors and dose-rate modifying factors based on these results were integrated into a model to estimate space radiation cancer risk (NASA Space Cancer Risk (NSCR) model) [4]. Now, target organs for degenerative effects induced by galactic cosmic rays, especially the brain, the cardiovascular system and the eye lens, are in the focus of the radiobiological research. A suspected cognitive decline by chronic exposure to galactic cosmic rays has achieved some celebrity as “Space Brain”. The roles of neurons, glia cells including astrocytes, oligodendrocytes and microglia remain to be elucidated – even sex-specific differences in the involvement of microglia have to be considered [5]. Experiments with accelerated heavy ions will help to elucidate the mechanisms of the degenerative effects of space radiation and will set the foundation to develop countermeasures. Here, besides the combination of beams to simulate better the radiation field in space, low dose rate experiments are of high interest. Besides the chronic space radiation exposure, astronauts experience a quite unique combination of possibly health-deteriorating environmental factors such as microgravity, noise, smell, disturbed circadian rhythm, increased carbon dioxide concentrations and decreased sleep quality. The interaction of radiation exposure with these space environmental factors such as microgravity or changes in the atmospheric conditions might influence the cells’ capability to cope with radiation damage. Also, the fluid shift towards the head might modulate radiation effects on the brain and eye. Recently, it was observed that the body temperature of astronauts on ISS is increased. In this context, it has to be considered that in some cancer therapy regimens, hyperthermia is combined with radiotherapy in order to augment the tumor cell killing effect. Finally, it has to be considered that humans are not only composed of their body cells that can be affected by heavy ion hits, but they also carry a microbiome inside and at the surface that quickly colonizes the surroundings, also spacecraft. The knowledge on how the human-microbiome and microbiome-environment interactions change under chronic space radiation exposure is very scarce. In conclusion, health risks by space radiation exposure have to be taken into account for an integrated design concept of space habitats, spacesuits and spacecraft. Despite some promising results on dietary measures (berries, dried plums) modulating the deteriorating effects of space radiation, a “magic pill” that can erase all radiation damage will most probably not be available.

The exposure to the space radiation environment remains a major limiting factor for human long-duration space missions due to its high biological effectiveness and the difficulties to effectively shield the radiation. The next decade in human spaceflight will be characterized by a continuous presence of human beings in Low Earth Orbit (LEO), by their return to the Moon and by longer stays in a Moon orbit on the Lunar Orbital Platform - Gateway (LOP-G). These endeavours are also performed to prepare for a human Mars mission.

An enduring human presence in space is required to achieve many goals of ESA´s and NASA´s space programs. During long-term missions, astronauts are exposed to various conditions such as microgravity, radiation, sleep-disruption, insufficient nutrition and also microbial contamination. Since astronauts´ immune function is compromised, all these factors increase the health risk for the crew. Therefore, improved spaceflight-suitable methods for microbiological monitoring, hygiene and decontamination are important. Microbial biofilms are of particular concern in spaceflight, because they cannot only cause damage to equipment but also be a risk for human health. Certain metals have antimicrobial properties such as copper and its respective alloys and therefore, prevent the formation of biofilms. However, precise evaluation of suitable antimicrobial materials for space flight is essential. Within the ESA project BIOFILMS (“Biofilm Inhibition on Flight equipment and on board the ISS using microbiologically Lethal Metal Surfaces"), effects of microgravity on biofilm formation on tailor-made nanostructured metallic surfaces are going to be examined. For that, human-associated microorganisms and antimicrobial surfaces with different chemical compositions and geometric nanostructures are selected. Experiments will be conducted aboard the ISS, where microbial growth will occur under optimal biofilm inducing conditions inside a particular designed hardware. The selected antimicrobial surfaces are supposed to be tested under different spaceflight-relevant gravitational regimes. Here, preliminary experiments with Staphylococcus capitis subsp. capitis for the BIOFILMS project will be presented. S. capitis forms biofilms and is related to infections such as endocarditis, urinary tract infections and catheter induced bacteremia. The reduction of growth and biofilm formation under the influence of copper-based, nanostructured surfaces was investigated.

Bacillus subtilis is one of the most studied Gram positive model organisms. Since mission Apollo 16, B. subtilis has been used for a multitude of space experiments. Investigating the influence of extreme conditions like those in space, non-domesticated strains, such as NCIB 3610 are of special interest regarding their ability to form biofilms. Since it is known that planktonic life is the exception, biofilms are considered as predominant way of living (Moons et al., 2009). Biofilms are organized in a complex self-produced extracellular polymeric matrix commonly composed of polysaccharides, proteins and nucleic acids. Building a biofilm protects the individual cell against shear forces, chemicals (e.g. antibiotics or disinfectants), temperature changes and water as well as nutrient depletion (Vlamakis et al., 2013, Cairns et al., 2014). The intrinsic resistance of biofilms is a problem, not only in industry and medicine, but it can be problematic under spaceflight conditions. Especially the loss of gravity coupled with changed levels of radiation might influence the resistance and therefore the virulence of bacterial biofilms. This can possibly evoke problems for the crew as well as for the spacecraft. In particular, long term missions with complex cooling systems, water supply and heat pipes may be vulnerable to biofilm colonisation. In our work, we used the biofilm-forming wildtype strain NCIB 3610 and a biofilm-matrix deficient mutant (deletion of 15-gene exopolysaccharide operon, epsA-O) to study the impact of reduced gravity on maturated biofilms. Our major research goal is to compare biofilm formation in simulated microgravity (using a 2D clinostat) to terrestrial gravity (1g) conditions by using different microscopic techniques. White light profilometry, scanning and transmission electron microscopy (SEM, TEM) and confocal laser scanning microscopy (CLSM) were used to analyse biofilms regarding their topology and inner structure, respectively. First results show qualitative architectural differences between simulated microgravity and 1g in cross-sections, but no significant qualitative variations in biofilm surface topography.

Artificial gravity through centrifugation is currently seen as countermeasure with potential against negative effects of spaceflight. However, side effects like increased motion sickness need to be investigated. Habituation protocols abate conflicts by reducing response to rotation with prolonged exposure to mismatches. Visually reinforced habituation may induce less reduction of oculomotor response to rotation than classical habituation.

Summary Gravitactic orientation was investigated in the unicellular photosynthetic flagellate, Euglena gracilis , under different accelerations between 0 and 1.5 × g during a recent space flight on board the American shuttle Columbia. The threshold for gravitaxis was found at ≤ 0.16 ≤ g. Above the threshold the precision of orientation increased with acceleration in a sigmoidal fashion and reached saturation at about 0.32 ≤ g, a behavior typical for physiological receptors. At accelerations above the saturation point the cells were closely aligned with the gravity vector (negative gravitaxis) and deviated more and more as the acceleration decreased. Obviously the gravireceptor responds to an error signal that elicits a course correction, again indicating the involvement of an active physiological gravireceptor. No adaptation of the cells to the conditions of weightlessness could be observed over the duration of the space mission (12 days). After landing, the cells showed a normal gravitactic behavior at 1 ≤ g.