Taxonomic and functional analyses of intact microbial communities thriving in extreme, astrobiology-relevant, anoxic sites.

BACKGROUND: Extreme terrestrial, analogue environments are widely used models to study the limits of life and to infer habitability of extraterrestrial settings. In contrast to Earth's ecosystems, potential extraterrestrial biotopes are usually characterized by a lack of oxygen. METHODS: In the MASE project (Mars Analogues for Space Exploration), we selected representative anoxic analogue environments (permafrost, salt-mine, acidic lake and river, sulfur springs) for the comprehensive analysis of their microbial communities. We assessed the microbiome profile of intact cells by propidium monoazide-based amplicon and shotgun metagenome sequencing, supplemented with an extensive cultivation effort. RESULTS: The information retrieved from microbiome analyses on the intact microbial community thriving in the MASE sites, together with the isolation of 31 model microorganisms and successful binning of 15 high-quality genomes allowed us to observe principle pathways, which pinpoint specific microbial functions in the MASE sites compared to moderate environments. The microorganisms were characterized by an impressive machinery to withstand physical and chemical pressures. All levels of our analyses revealed the strong and omnipresent dependency of the microbial communities on complex organic matter. Moreover, we identified an extremotolerant cosmopolitan group of 34 poly-extremophiles thriving in all sites. CONCLUSIONS: Our results reveal the presence of a core microbiome and microbial taxonomic similarities between saline and acidic anoxic environments. Our work further emphasizes the importance of the environmental, terrestrial parameters for the functionality of a microbial community, but also reveals a high proportion of living microorganisms in extreme environments with a high adaptation potential within habitability borders. Video abstract.

Microbiome dynamics during the HI-SEAS IV mission, and implications for future crewed missions beyond Earth.

BACKGROUND: Human health is closely interconnected with its microbiome. Resilient microbiomes in, on, and around the human body will be key for safe and successful long-term space travel. However, longitudinal dynamics of microbiomes inside confined built environments are still poorly understood. Herein, we used the Hawaii Space Exploration Analog and Simulation IV (HI-SEAS IV) mission, a 1 year-long isolation study, to investigate microbial transfer between crew and habitat, in order to understand adverse developments which may occur in a future outpost on the Moon or Mars. RESULTS: Longitudinal 16S rRNA gene profiles, as well as quantitative observations, revealed significant differences in microbial diversity, abundance, and composition between samples of the built environment and its crew. The microbiome composition and diversity associated with abiotic surfaces was found to be rather stable, whereas the microbial skin profiles of individual crew members were highly dynamic, resulting in an increased microbiome diversity at the end of the isolation period. The skin microbiome dynamics were especially pronounced by a regular transfer of the indicator species Methanobrevibacter between crew members within the first 200 days. Quantitative information was used to track the propagation of antimicrobial resistance in the habitat. Together with functional and phenotypic predictions, quantitative and qualitative data supported the observation of a delayed longitudinal microbial homogenization between crew and habitat surfaces which was mainly caused by a malfunctioning sanitary facility. CONCLUSIONS: This study highlights main routes of microbial transfer, interaction of the crew, and origins of microbial dynamics in an isolated environment. We identify key targets of microbial monitoring, and emphasize the need for defined baselines of microbiome diversity and abundance on surfaces and crew skin. Targeted manipulation to counteract adverse developments of the microbiome could be a highly important strategy to ensure safety during future space endeavors. Video abstract.

Space Station conditions are selective but do not alter microbial characteristics relevant to human health.

The International Space Station (ISS) is a unique habitat for humans and microorganisms. Here, we report the results of the ISS experiment EXTREMOPHILES, including the analysis of microbial communities from several areas aboard at three time points. We assess microbial diversity, distribution, functional capacity and resistance profile using a combination of cultivation-independent analyses (amplicon and shot-gun sequencing) and cultivation-dependent analyses (physiological and genetic characterization of microbial isolates, antibiotic resistance tests, co-incubation experiments). We show that the ISS microbial communities are highly similar to those present in ground-based confined indoor environments and are subject to fluctuations, although a core microbiome persists over time and locations. The genomic and physiological features selected by ISS conditions do not appear to be directly relevant to human health, although adaptations towards biofilm formation and surface interactions were observed. Our results do not raise direct reason for concern with respect to crew health, but indicate a potential threat towards material integrity in moist areas.

Microbial biodiversity assessment of the European Space Agency's ExoMars 2016 mission.

BACKGROUND: The ExoMars 2016 mission, consisting of the Trace Gas Orbiter and the Schiaparelli lander, was launched on March 14 2016 from Baikonur, Kazakhstan and reached its destination in October 2016. The Schiaparelli lander was subject to strict requirements for microbial cleanliness according to the obligatory planetary protection policy. To reach the required cleanliness, the ExoMars 2016 flight hardware was assembled in a newly built, biocontrolled cleanroom complex at Thales Alenia Space in Turin, Italy. In this study, we performed microbiological surveys of the cleanroom facilities and the spacecraft hardware before and during the assembly, integration and testing (AIT) activities. METHODS: Besides the European Space Agency (ESA) standard bioburden assay, that served as a proxy for the microbiological contamination in general, we performed various alternative cultivation assays and utilised molecular techniques, including quantitative PCR and next generation sequencing, to assess the absolute and relative abundance and broadest diversity of microorganisms and their signatures in the cleanroom and on the spacecraft hardware. RESULTS: Our results show that the bioburden, detected microbial contamination and microbial diversity decreased continuously after the cleanroom was decontaminated with more effective cleaning agents and during the ongoing AIT. The studied cleanrooms and change room were occupied by very distinct microbial communities: Overall, the change room harboured a higher number and diversity of microorganisms, including Propionibacterium, which was found to be significantly increased in the change room. In particular, the so called alternative cultivation assays proved important in detecting a broader cultivable diversity than covered by the standard bioburden assay and thus completed the picture on the cleanroom microbiota. CONCLUSION: During the whole project, the bioburden stayed at acceptable level and did not raise any concern for the ExoMars 2016 mission. The cleanroom complex at Thales Alenia Space in Turin is an excellent example of how efficient microbiological control is performed.

Resilient microorganisms in dust samples of the International Space Station-survival of the adaptation specialists.

BACKGROUND: The International Space Station (ISS) represents a unique biotope for the human crew but also for introduced microorganisms. Microbes experience selective pressures such as microgravity, desiccation, poor nutrient-availability due to cleaning, and an increased radiation level. We hypothesized that the microbial community inside the ISS is modified by adapting to these stresses. For this reason, we analyzed 8-12 years old dust samples from Russian ISS modules with major focus on the long-time surviving portion of the microbial community. We consequently assessed the cultivable microbiota of these samples in order to analyze their extremotolerant potential against desiccation, heat-shock, and clinically relevant antibiotics. In addition, we studied the bacterial and archaeal communities from the stored Russian dust samples via molecular methods (next-generation sequencing, NGS) and compared our new data with previously derived information from the US American ISS dust microbiome. RESULTS: We cultivated and identified in total 85 bacterial, non-pathogenic isolates (17 different species) and 1 fungal isolate from the 8-12 year old dust samples collected in the Russian segment of the ISS. Most of these isolates exhibited robust resistance against heat-shock and clinically relevant antibiotics. Microbial 16S rRNA gene and archaeal 16S rRNA gene targeting Next Generation Sequencing showed signatures of human-associated microorganisms (Corynebacterium, Staphylococcus, Coprococcus etc.), but also specifically adapted extremotolerant microorganisms. Besides bacteria, the detection of archaeal signatures in higher abundance was striking. CONCLUSIONS: Our findings reveal (i) the occurrence of living, hardy microorganisms in archived Russian ISS dust samples, (ii) a profound resistance capacity of ISS microorganisms against environmental stresses, and (iii) the presence of archaeal signatures on board. In addition, we found indications that the microbial community in the Russian segment dust samples was different to recently reported US American ISS microbiota.