The human spaceflight environment is notable for the unique factor of microgravity, which exerts numerous physiologic effects on macroscopic organisms, but how this environment may affect single-celled microbes is less clear. In an effort to understand how the microbial transcriptome responds to the unique environment of spaceflight, the model Gram-positive bacterium Bacillus subtilis was flown on two separate missions to the International Space Station in experiments dubbed BRIC-21 and BRIC-23. Cells were grown to late-exponential/early stationary phase, frozen, then returned to Earth for RNA-seq analysis in parallel with matched ground control samples. A total of 91 genes were significantly differentially expressed in both experiments; 55 exhibiting higher transcript levels in flight samples and 36 showing higher transcript levels in ground control samples. Genes upregulated in flight samples notably included those involved in biofilm formation, biotin and arginine biosynthesis, siderophores, manganese transport, toxin production and resistance, and sporulation inhibition. Genes preferentially upregulated in ground control samples notably included those responding to oxygen limitation, e.g., fermentation, anaerobic respiration, subtilosin biosynthesis, and anaerobic regulatory genes. The results indicated differences in oxygen availability between flight and ground control samples, likely due to differences in cell sedimentation and the toroidal shape assumed by the liquid cultures in microgravity.

No abstract

The International Space Station (ISS) is a uniquely enclosed environment that has been continuously occupied for the last two decades. Throughout its operation, protecting the health of the astronauts on-board has been a high priority. The human microbiome plays a significant role in maintaining human health, and disruptions in the microbiome have been linked to various diseases. To evaluate the effects of spaceflight on the human microbiome, body swabs and saliva samples were collected from four ISS astronauts on consecutive expeditions. Astronaut samples were analyzed using shotgun metagenomic sequencing and microarrays to characterize the microbial biodiversity before, during, and after the astronauts’ time onboard the ISS. Samples were evaluated at an individual and population level to identify changes in microbial diversity and abundance. No significant changes in the number or relative abundance of taxa were observed between collection time points when samples from all four astronauts were analyzed together. When the astronauts’ saliva samples were analyzed individually, the saliva samples of some astronauts showed significant changes in the relative abundance of taxa during and after spaceflight. The relative abundance of Prevotella in saliva samples increased during two astronauts’ time onboard the ISS while the relative abundance of other commensal taxa such as Neisseria, Rothia, and Haemophilus decreased. The abundance of some antimicrobial resistance genes within the saliva samples also showed significant changes. Most notably, elfamycin resistance gene significantly increased in all four astronauts post-flight and a CfxA6 beta-lactam marker significantly increased during spaceflight but returned to normal levels post-flight. The combination of both shotgun metagenomic sequencing and microarrays showed the benefit of both technologies in monitoring microbes on board the ISS. There were some changes in each astronaut’s microbiome during spaceflight, but these changes were not universal for all four astronauts. Two antimicrobial resistance gene markers did show a significant change in abundance in the saliva samples of all four astronauts across their collection times. These results provide insight for future ISS microbial monitoring studies and targets for antimicrobial resistance screenings.

Background The International Space Station (ISS) is a unique and complex built environment with the ISS surface microbiome originating from crew and cargo or from life support recirculation in an almost entirely closed system. The Microbial Tracking 1 (MT-1) project was the first ISS environmental surface study to report on the metagenome profiles without using whole-genome amplification. The study surveyed the microbial communities from eight surfaces over a 14-month period. The Microbial Tracking 2 (MT-2) project aimed to continue the work of MT-1, sampling an additional four flights from the same locations, over another 14 months. Methods Eight surfaces across the ISS were sampled with sterile wipes and processed upon return to Earth. DNA extracted from the processed samples (and controls) were treated with propidium monoazide (PMA) to detect intact/viable cells or left untreated and to detect the total DNA population (free DNA/compromised cells/intact cells/viable cells). DNA extracted from PMA-treated and untreated samples were analyzed using shotgun metagenomics. Samples were cultured for bacteria and fungi to supplement the above results. Results Staphylococcus sp. and Malassezia sp. were the most represented bacterial and fungal species, respectively, on the ISS. Overall, the ISS surface microbiome was dominated by organisms associated with the human skin. Multi-dimensional scaling and differential abundance analysis showed significant temporal changes in the microbial population but no spatial differences. The ISS antimicrobial resistance gene profiles were however more stable over time, with no differences over the 5-year span of the MT-1 and MT-2 studies. Twenty-nine antimicrobial resistance genes were detected across all samples, with macrolide/lincosamide/streptogramin resistance being the most widespread. Metagenomic assembled genomes were reconstructed from the dataset, resulting in 82 MAGs. Functional assessment of the collective MAGs showed a propensity for amino acid utilization over carbohydrate metabolism. Co-occurrence analyses showed strong associations between bacterial and fungal genera. Culture analysis showed the microbial load to be on average 3.0 × 105 cfu/m2 Conclusions Utilizing various metagenomics analyses and culture methods, we provided a comprehensive analysis of the ISS surface microbiome, showing microbial burden, bacterial and fungal species prevalence, changes in the microbiome, and resistome over time and space, as well as the functional capabilities and microbial interactions of this unique built microbiome. Data from this study may help to inform policies for future space missions to ensure an ISS surface microbiome that promotes astronaut health and spacecraft integrity.

Past results have suggested that bacterial antibiotic susceptibility is altered during space flight. To test this notion, cells were cultivated in matched hardware, medium, and environmental conditions either in spaceflight microgravity on the International Space Station, termed Flight (FL) samples, or at Earth-normal gravity, termed Ground Control (GC) samples. Susceptibility of FL and GC samples was compared to 72 antibiotics and growth-inhibitory compounds using the Omnilog Phenotype Microarray (PM) system. Only 9 compounds were identified by PM screening as exhibiting significant differences ( < 0.05, Student's t-test) in FL vs. GC samples: 6-mercaptopurine, cesium chloride, enoxacin, lomefloxacin, manganese (II) chloride, nalidixic acid, penimepicycline, rolitetracycline, and trifluoperazine. Testing of the same compounds by standard broth dilution assay did not reveal statistically significant differences in the IC values between FL and GC samples. The results indicate that the susceptibility of cells to a wide range of antibiotics and growth inhibitors is not dramatically altered by space flight. This study addresses a major concern of mission planners for human spaceflight, that bacteria accompanying astronauts on long-duration missions might develop a higher level of resistance to antibiotics due to exposure to the spaceflight environment. The results of this study do not support that notion.