Presented here is the environmental microbiome study of the International Space Station surfaces. The environmental samples were collected with the polyester wipes from eight different locations in the ISS during two consecutive sampling sessions (three months apart). The specific objective was to unveil the pool of genes for each location during two separate sessions to learn of functional and metabolic diversity of microorganisms in the ISS. The International Space Station (ISS) as a closed built environment has its own environmental microbiome which is shaped by microgravity radiation and limited human presence. The microbial diversity associated with ISS environmental surfaces was investigated during this study. Polyester wipes and contact slides were used for sampling of eight various surface locations on the ISS at different time periods. The samples were retrieved and analyzed immediately upon the return to the Earth (via Soyuz TMA-14M or Dragon capsule from SpaceX). After surface sample collection contact slides containing nutrient media for the growth of bacteria and fungi were incubated at 25 xcb x9aC. The polyester wipes were processed to measure microbial burden (R2A Blood Agar and Potato Dextrose Agar) and recover cultivable bacteria as well as fungi. Subsequently viable microbial burden was assessed using Adenosine Triphosphate (ATP) assay and quantitative polymerase chain reaction (PCR) methods after propidium monoazide (PMA) treatment. The 16S-tag and metagenome analyses were used to elucidate viable microbial diversity. The cultivable bacterial population yield from the polyester wipes was very high (5 to 7-logs) when compared with the contact slides (102 to 103 CFU/m2). The PMA-qPCR analysis showed considerable variation of viable bacterial population (105 to 109 16S rDNA gene copies/m2) among locations sampled. Unlike contact slides polyester wipes cover much larger sample surface (~1 m2) and produce much more reliable results of the microbial diversity of the ISS covering both cultivable and non-cultivable species. The cultivable total and viable microbial diversity was determined utilizing state-of-the art molecular techniques. The implementation of the PMA assay before DNA extraction allowed distinguishing viable microorganisms which is crucial for determining their role to the crew health the ISS maintenance and the general knowledge of the closed environmentally controlled built systems.

The Rodent Research-3 (RR-3) mission was sponsored by the pharmaceutical company Eli Lilly and Co. and the Center for the Advancement of Science in Space to study the effectiveness of a potential countermeasure for the loss of muscle and bone mass that occurs during spaceflight. Twenty BALB/c 18-weeks old female mice (ten controls and ten treated) were flown to the ISS and housed in the AEM-X habitat for 39-42 days. Twenty mice of similar age sex and strain were used for ground controls housed in identical hardware and matching ISS environmental conditions. Basal controls were housed in standard vivarium cages. Spaceflight ground controls and basal groups had blood collected then were euthanized had one hind limb removed and finally whole carcasses were stored at -80 C until dissection. All mice in this data set received only the control/sham injection.

NASA s Rodent Research (RR) project is playing a critical role in advancing biomedical research on the physiological effects of space environments. Due to the limited resources for conducting biological experiments aboard the International Space Station (ISS) it is imperative to use crew time efficiently while maximizing high-quality science return. NASA s GeneLab project has as its primary objectives to 1) further increase the value of these experiments using a multi-omics systems biology-based approach and 2) disseminate these data without restrictions to the scientific community. The current investigation assessed viability of RNA DNA and protein extracted from archived RR-1 tissue samples for epigenomic transcriptomic and proteomic assays. During the first RR spaceflight experiment a variety of tissue types were harvested from subjects snap-frozen or RNAlater-preserved and then stored at least a year at -80C after return to Earth. They were then prioritized for this investigation based on likelihood of significant scientific value for spaceflight research. All tissues were made available to GeneLab through the bio-specimen sharing program managed by the Ames Life Science Data Archive and included mouse adrenal glands quadriceps gastrocnemius tibialis anterior extensor digitorum longus soleus eye and kidney. We report here protocols for and results of these tissue extractions and thus the feasibility and value of these kinds of omics analyses. In addition to providing additional opportunities for investigation of spaceflight effects on the mouse transcriptome and proteome in new kinds of tissues our results may also be of value to program managers for the prioritization of ISS crew time for rodent research activities.

Space travel is associated with continuous low-dose-rate exposure to high Linear Energy Transfer (LET) radiation. Pathophysiological manifestations after low-dose radiation exposure are strongly influenced by non-cytocidal radiation effects including microbiome and cellular gene expression. Using a mouse model for exposure to high LET radiation we observed substantial changes in the composition and functional potential of the microbiome. These were paralleled by changes in the abundance of multiple metabolites which were related to the enzymatic activity of the altered metagenome by means of metabolic network modeling. There was a complex dynamic in microbial and metabolic composition at different radiation doses suggestive of transient dose-dependent interactions between microbial ecology and signals from the host s cellular damage repair processes. Functional shifts included features associated with dysbiosis at the onset of chronic inflammatory responses which could prMouse fecal microbiome after exposure to high LET radiatione-dispose space travelers to systemic long-term health risks.

Proper interpretation of RNA sequencing data requires an understanding of assay sensitivity and sources of variability. To this end the External RNA Control Consortium (ERCC) developed a standard set of 92 poly-adenylated RNA transcripts that are orthogonal to mammalian RNA that can be added to RNA extracts before library generation and sequencing. The presence of these RNA standards at known ratios improves interpretation of RNA sequencing datasets. To test the utility of the ERCC RNA controls total RNA extracted from mouse livers from the Rodent Research 1 (flight and ground groups) and Rodent Research 3 (flight and ground groups) missions was sequenced with and without the ERCC control RNA. To allow comparison within and between groups ERCC Mix 1 or Mix 2 were added to half of the samples from each group respectively.

Physical forces greatly influence the growth and function of an organism. Altered gravity can perturb normal development and induce corresponding changes in gene expression. Understanding this relationship between the physical and biological realms is important for NASA s space travel goals. We use combined RNA-Seq and qRT-PCR to profile changes in early Drosophila melanogaster pupae exposed to chronic hypergravity (3 g three times Earth s gravity) to highlight gravity-dependent pathways and gene products. Robust transcriptional response was evident among the pupae developed in a hypergravity environment compared to control. 1,513 genes showed significantly (p < 0.05) altered gene expression in the 3 g samples. These findings were supported with qRT-PCR data. Major biological processes affected include ion transport redox homeostasis immune and humoral stress response proteolysis and cuticle development.

The environmental samples were collected with the polyester wipes from eight different locations in the International Space Station (ISS) during two consecutive sampling sessions (three months apart) within the ISS Microbial Observatory Experiment. DNA extracted from each of the samples was used to create amplicon libraries based on customized panel of 500 antimicrobial resistance genes followed by next-generation sequencing. This is the first study of that shows the reservoir of antimicrobial genes in the ISS. The International Space Station (ISS) as a closed built environment has its own environmental microbiome which is shaped by microgravity radiation and limited human presence. The microbial diversity associated with ISS environmental surfaces was investigated during this study. Polyester wipes and contact slides were used for sampling of eight various surface locations on the ISS at different time periods. The samples were retrieved and analyzed immediately upon the return to the Earth (via Soyuz TMA-14M or Dragon capsule from SpaceX). After surface sample collection contact slides containing nutrient media for the growth of bacteria and fungi were incubated at 25 xcb x9aC. The polyester wipes were processed to measure microbial burden (R2A Blood Agar and Potato Dextrose Agar) and recover cultivable bacteria as well as fungi. Subsequently viable microbial burden was assessed using Adenosine Triphosphate (ATP) assay and quantitative polymerase chain reaction (PCR) methods after propidium monoazide (PMA) treatment. The 16S-tag and metagenome analyses were used to elucidate viable microbial diversity. The cultivable bacterial population yield from the polyester wipes was very high (5 to 7-logs) when compared with the contact slides (102 to 103 CFU/m2). The PMA-qPCR analysis showed considerable variation of viable bacterial population (105 to 109 16S rDNA gene copies/m2) among locations sampled. Unlike contact slides polyester wipes cover much larger sample surface (~1 m2) and produce much more reliable results of the microbial diversity of the ISS covering both cultivable and non-cultivable species. The cultivable total and viable microbial diversity was determined utilizing state-of-the art molecular techniques. The implementation of the PMA assay before DNA extraction allowed distinguishing viable microorganisms which is crucial for determining their role to the crew health the ISS maintenance and the general knowledge of the closed environmentally controlled built systems.

Microbes interact with humans in complex ways and understanding how they respond to the spaceflight environment is important to the success of future manned spaceflight missions. The BRIC-23 mission was designed to measure the response of Bacillus subtilis and Staphylococcus aureus to the spaceflight environment. This experiment aimed to produce high quality omics data from B. subtilis and S. aureus grown aboard the International Space Station (ISS) to allow comparison to matched ground controls. There were two primary objectives for this experiment: (1) Demonstrate all post-flight processes and operations required for successful completion of GeneLab Reference Missions conducted on ISS and (2) Generate high quality GeneLab Reference Mission omics data sets for two prokaryotic model organisms Bacillus subtilis and Staphylococcus aureus. Freezing Control Experiment: The BRIC hardware has significant thermal inertia thus the freezing rate of samples placed at -80 C is quite slow. This could affect RNA-sequencing proteomic and metabolic data sets. In an effort to understand how slow freezing could affect these data sets a control experiment was designed in which B. subtilis and S. aureus were grown in petri plates and either slow frozen to -80 C at a rate matching the BRIC-23 spaceflight samples or processed immediately to harvest RNA and protein.

NASA s Rodent Research (RR) project is playing a critical role in advancing biomedical research on the physiological effects of space environments. Due to the limited resources for conducting biological experiments aboard the International Space Station (ISS) it is imperative to use crew time efficiently while maximizing high-quality science return. NASA s GeneLab project has as its primary objectives to 1) further increase the value of these experiments using a multi-omics systems biology-based approach and 2) disseminate these data without restrictions to the scientific community. The current investigation assessed viability of RNA DNA and protein extracted from archived RR-1 tissue samples for epigenomic transcriptomic and proteomic assays. During the first RR spaceflight experiment a variety of tissue types were harvested from subjects snap-frozen or RNAlater-preserved and then stored at least a year at -80C after return to Earth. They were then prioritized for this investigation based on likelihood of significant scientific value for spaceflight research. All tissues were made available to GeneLab through the bio-specimen sharing program managed by the Ames Life Science Data Archive and included mouse adrenal glands quadriceps gastrocnemius tibialis anterior extensor digitorum longus soleus eye and kidney. We report here protocols for and results of these tissue extractions and thus the feasibility and value of these kinds of omics analyses. In addition to providing additional opportunities for investigation of spaceflight effects on the mouse transcriptome and proteome in new kinds of tissues our results may also be of value to program managers for the prioritization of ISS crew time for rodent research activities.

The whole-genome sequences of eight fungal strains that were selected for exposure to microgravity at the International Space Station are presented here. These baseline sequences will help to understand the observed production of novel bioactive compounds.