All systems are glow: in-situ cellular fluorescence during space exposure

Given the remarkable ability of microorganisms to both survive and prosper in extreme environments on Earth, several species have the potential to survive the environmental conditions of space1, 2. With the rapidly growing number of interplanetary space exploration missions, this evokes several interesting questions regarding forward planetary contamination and fundamental radiation-biology relating to the origins and extremes of life. It is therefore critically important to better understand microorganisms and their composite biomolecules under space conditions. However, experimental simulation of space and planetary environments is particularly challenging; while some conditions such as temperature or planetary atmospheres can be replicated in the laboratory, an accurate replication of microgravity3 or high-energy solar and cosmic radiation4 (and their interaction) is an ongoing challenge. As such, the continued development of space-based experimental platforms, particularly those incorporating in-situ measurements, is a crucial tool for ongoing astrobiology and astrochemistry research. Historical space-exposure experiments were limited to low-Earth orbit-based platforms, such as on the outside of the space shuttle5, 6, descent capsules7, or the International Space Station (ISS) 8, 9, and required sample return for post-flight analyses. In contrast, several recent satellite-based astrobiology experiments have integrated fluidic-hydration systems and compact optical detectors, allowing measurements to be performed in situ. Specifically, several space experiments (O/OREOS10, PharmaSat11, EcAMSat12, and BioSentinel13) utilized a colorimetric redox-indicator dye to measure metabolic responses from actively growing space-exposed organisms. This concept can be expanded by integrating fluorescence detection into the next generation of astrobiology exposure technology. Due to the wide functional variety of fluorescent compounds, fluorometry has the potential to greatly broaden the investigation of metabolic parameters under different space conditions. In-situ fluorometry is a goal highlighted by both the NASA Astrobiology Strategy14 and the ESA Astrobiology Roadmap15. The new astrobiology exposure platform, Exocube, aims to address questions regarding biomolecular stability and microbial responses to space conditions through use of in-situ cellular fluorescence detection. Exocube has been selected for implementation by the European Space Agency as part of the new European Space Exposure Platform, and as biological response to space radiation is a major focus, Exocube will be installed on the Bartolomeo platform, outside of the ISS. Here, a variety of samples ranging from biomarker molecules to live microorganisms isolated from extreme environments on Earth (chiefly those associated with high levels of radiation, elevated ultraviolet light, or extreme desiccation) will be exposed to the dramatically elevated levels of broad-range, high-energy, ionizing radiation beyond Earths atmosphere. Exocube will be capable of performing real-timein-situobservations of both microbial growth and survival, as well as an assessment of various metabolic function via fluorescence detection. Additionally, Exocube will be the first astrobiology exposure platform to combine both the strengths of real-time in-situ measurements, with the capacity for sample return and the subsequent strengths of detailed post-flight sample analyses. Here we present the current developments of Exocube, specifically focusing on the selection of sample species through preliminary biocompatibility testing, and the integration and optimization of cellular fluorescence using prototype space-experiment hardware. While the fluorescent staining of microorganisms is commonplace in laboratory settings, incorporating such compounds into space-flight hardware is unconventional and poses several challenges. However, this novel fluorescence detection has the potential to provide new, highly specific information pertaining to growth, cellular metabolism, membrane integrity and reactive oxygen species accumulation in live organisms exposed to space conditions. Exocube represents the newest generation of exobiological exposure platforms, aiming to address questions of early-life chemistry, and further our understanding of the limits of life in space. 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