Survival of NASA-cleanroom microbial isolates under simulated space and Martian conditions

Planetary protection hinges on understanding microbial survival following reduction procedures, the stressors of space travel, and exposure to extraterrestrial environmental conditions. This study identified 23 fungal strains isolated from NASA spacecraft assembly cleanrooms, capable of surviving ultraviolet radiation exposure. Using experimental simulation facilities, we conducted a comprehensive assessment of microbial survivability and morphology on the most resilient spacecraft-associated microorganisms. Aspergillus calidoustus demonstrated remarkable survival under simulated Martian conditions, withstanding up to 1,440 min of Martian solar irradiation, Mars atmospheric pressure and composition, and the presence of Martian regolith. Lethality only occurred under combined irradiation and cooling to -60°C (the mean Mars surface temperature), emphasizing the synergistic effect of these conditions. Furthermore, A. calidoustus survived long-duration neutron radiation exposure (replicating ionizing space radiation doses) and dry-heat microbial reduction technique (typically used for spacecraft components). This is the first study to perform an end-to-end evaluation of eukaryotic microbial survival across conditions that occur during preparation for, travel to, and robotic exploration of Mars. The experimental facilities and chronic exposure methods utilized offer a biologically meaningful model for understanding microbial risks during long-duration space missions. The capacity for fungal conidia to survive multiple space-relevant conditions suggests their potential as forward contaminants, capable of being transported to and persisting on Mars. As current spacecraft microbial reduction protocols prioritize bacterial spores, this research highlights a critical gap in planetary protection strategies. In addition to offering novel insights into microbial survival, these findings have broader implications for biocontamination within the food, pharmaceutical, and medical sectors.IMPORTANCEThis study reveals that conidia of the fungus Aspergillus calidoustus, which was isolated from spacecraft assembly cleanrooms, can survive simulated space-relevant stressors like ultraviolet irradiation, Martian cold atmospheric pressure, regolith exposure, ionizing radiation, and specific doses of recommended dry-heat microbial reduction method for spacecraft. Such fungal resistance demonstrates that the species can survive certain space and Mars conditions previously thought to be sterilizing, highlighting a need to revise current spacecraft decontamination standards that focus mainly on bacterial spores. This study also emphasizes the need for continued microbial monitoring of spacecraft during transit from Earth to other planets, not only to achieve goals of planetary protection but also to maintain healthy closed systems for human missions. Moreover, fungal species are highlighted as biocontamination risks for food, medical, and pharmaceutical industries, which may require the need for new standards of sterilization approaches transferable to space exploration.