During space flight bone mineral density is decreased by the influence of osteoclast activation which molecular mechanism is expectantly investigated. In the study of medaka bone development we investigated the system of vertebra formation and firstly identified the presence of osteoclasts in medaka. Moreover osteoclast rsorbing activity was affected by hypergravity indicating the possibility that we can investigate the effect of microgravity on osteoclasts in space. To find this effect we examine the alteration of osteoclast activity under microgravity with the histological analysis or the expression analysis by RNA in-situ hybridization. Furthermore since we have succeeded the establishment of the medaka osteoclast-specific transgenic lines we perform the in-vivo imaging analyses for gene expression and cell mobility. Finally to examine the gravity sensing system we employ tooth and bone as the high density organs which are highly sensitive to gravity and perform the histological analysis and the gene expression analysis of such gravity-sensitive tissues at surrounding pharyngeal teeth and supporting bone.

To understand how humans adapt to space environments many experiments can be conducted on astronauts while they work aboard the Space Shuttle or the International Space Station (ISS). We also need animal experiments that can apply to human models and help prevent or solve the physical issues we face in space travel. The medaka is a suitable model fish for studying space adaptation because in the second International Microgravity Laboratory mission in 1994 adult fish mated successfully in space during 15 days of flight. In 2012 another space experiment Medaka Osteoclast was performed. Male and female fish (6 weeks old at launching) were maintained in the Aquatic Habitat system for 2 months in the ISS. The RNA-seq analysis of tissues from these fish will revealed tissue-specific responsiveness and common stress responses during space adaptation.

Spaceflight is a unique environment with profound effects on biological systems including tissue redistribution and musculoskeletal stresses. However the more subtle biological effects of spaceflight on cells and organisms are difficult to measure in a systematic unbiased manner. Here we test the utility of the molecularly barcoded yeast deletion collection to provide a quantitative assessment of the effects of microgravity on a model organism. We developed robust hardware to screen in parallel the complete collection of ~4800 homozygous and ~5900 heterozygous (including ~1100 single-copy deletions of essential genes) yeast deletion strains each carrying unique DNA that acts as strain identifiers. We compared strain fitness for the homozygous and heterozygous yeast deletion collections grown in spaceflight and ground as well as plus and minus hyperosmolar sodium chloride providing a second additive stressor. The genome-wide sensitivity profiles obtained from these treatments were then queried for their similarity to a compendium of drugs whose effects on the yeast collection have been previously reported. We found that the effects of spaceflight have high concordance with the effects of DNA-damaging agents and changes in redox state suggesting mechanisms by which spaceflight may negatively affect cell fitness.

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. B.subtilis omics data is deposited in GLDS-138.

Astronauts have been previously shown to exhibit decreased salivary lysozyme and increased dental calculus and gingival inflammation in response to space flight host factors that could contribute to oral diseases such as caries and periodontitis. However the specific physiological response of caries-causing bacteria such as Streptococcus mutans to space flight and/or ground-based simulated microgravity has not been extensively investigated. In this study High Aspect Ratio Vessel (HARV) S. mutans simulated microgravity and normal gravity cultures were assessed for changes in metabolite and transcriptome profiles H2O2 resistance and competence in sucrose-containing biofilm media. Stationary phase S. mutans simulated microgravity cultures displayed increased killing by H2O2 compared to normal gravity control cultures but competence was not affected. RNA-seq analysis revealed that expression of 153 genes was up-regulated >= 2-fold and 94 genes down-regulated >= 2-fold during simulated microgravity HARV growth. These included a number of genes located on extrachromosomal elements as well as genes involved in carbohydrate metabolism translation and stress responses. Collectively these results suggest that growth under microgravity analog conditions promotes changes in S. mutans gene expression and physiology that may translate to an altered cariogenic potential of this organism during space flight missions. Overall design: Differential gene expression was compared between RNA from S. mutans grown in normal gravity HARVs (n=3 independent cultures) and RNA from S. mutans grown in simulated microgravity HARVs (n=3 independent cultures)

Here in this study we systematically examined the patterns of DNA methylation and hydroxy-methylation with its functional implications in gene regulation for the cultured TK6 lymphoblastoid cells upon exposure to micro-gravity conditions. The results reported here indicate that simulated microgravity alters methylation patterns in a limited way and subsequently the expression of genes involved in stress response like ATF3 FBXO17 MAP3K13 and VCL in TK6 cells. Overall design: Examination of 2 different DNA modifications with 2 replicates each for 1 cell type.

We report how high and low linear energy transfer (LET) radiation microgravity and elevated dietary iron affect colon microbiota (determined by 16S rDNA pyrosequencing) and colon function. Three independent experiments were conducted: 1) fractionated low LET gamma radiation (137Cs 3 Gy RAD) high Fe diet (IRON) (650 mg/kg diet) and a combination of low LET gamma radiation and high Fe diet (IRON+RAD) in male Sprague-Dawley rats; 2) high LET 38Si particle exposure (0.050 Gy) 1/6 G partial weight bearing (PWB) and a combination of high LET38Si particle exposure and PWB in female BalbC/ByJ mice; and 3) 13 d spaceflight in female C57BL/6 mice. For each experiment the colon was resected and feces removed for microbial sequencing analysis on a Roche 454 Genome Sequencer FLX Titanium instrument (Microbiome Core Facility Chapel Hill NC) using the GS FLX Titanium XLR70 sequencing reagents and protocols. Analysis of amplicon sequencing data was carried out using the QIIME pipeline. Low LET radiation high iron diet and spaceflight increased Bacteroidetes and decreased Firmicutes. Low LET radiation high Fe diet and spaceflight did not significantly affect diversity or richness or elevate pathogenic genera. Spaceflight increased Clostridiales and decreased Lactobacillales and similar trends were observed in the experiment using a ground-based model of microgravity suggesting altered gravity may affect colonic microbiota. Microbiota characteriztion in these models is a first step in understanding the impact of the space environment on intestinal health.

Aspergillus fumigatus is a saprophytic filamentous fungus that is ubiquitous outdoors (soil decaying vegetation) and indoors (hospitals simulated closed habitats etc.). A. fumigatus can adapt to various environmental conditions and form airborne conidia that are the inoculum for a variety of diseases (e.g. non- and invasive pulmonary infections allergic bronchopulmonary aspergillosis etc.) in immunocompromised hosts. In an on-going Microbial Observatory Experiments on the International Space Station (ISS) molecular phylogeny of several fungal isolates were characterized. Two strains ISSF 21 and IF1SW-F4 were isolated from the HEPA filter and the surface of the Cupola of the ISS respectively. Using primers targeting the internal transcribed spacers ITS1 and 2 both isolates were identified as A. fumigatus. The whole genome sequence analysis of ISSF 21 revealed increased number of single nucleotide polymorphisms (SNPs) when compared to the reference A. fumigatus 293. Knowing that A. fumigatus is an opportunistic pathogen and microgravity highly influences the antibiotic susceptibility and pathogenicity of microorganisms we examined pathogenicity of both ISS isolates using the zebrafish larval model. The space station isolates (ISSF-021 and IF1SW-F4) were more virulent than two clinical strains (Af293 and CEA10) whose pathogenicity was highly characterized. Here the whole genome sequences of ISSF-021 strain are being deposited.

The vestibular system receives a permanent influence from gravity and reflexively controls equilibrium. If we assume gravity has remained constant during the species evolution will its sensory system adapt to abrupt loss of that force? We address this question in the land snail Helix lucorum exposed to 30 days of near weightlessness aboard the Bion-M1 satellite and studied geotactic behavior of postflight snails differential gene expressions in statocyst transcriptome and electrophysiological responses of mechanoreceptors to applied tilts. Each approach revealed plastic changes in the snail s vestibular system assumed in response to spaceflight. Absence of light during the mission also affected statocyst physiology as revealed by comparison to dark-conditioned control groups. Readaptation to normal tilt responses occurred at ~20 h following return to Earth. Despite the permanence of gravity the snail responded in a compensatory manner to its loss and readapted once gravity was restored.

Our study focuses on the hardiest microorganisms inhabiting the ISS in order to assess their diversity and capabilities to resist certain stresses. We specifically selected dust samples from the Russian modules that were obtained 8-10 years ago and stored since then under sealed conditions on Earth. Targeting long-time survivors and spore-forming microorganisms we assessed consequently the cultivable microbial community of these samples in order to obtain model microbial strains that could help to analyze specific adaptation towards environmental stresses such as desiccation and lack of nutrients. In this study we analyzed these microorganisms with respect to their resistance towards thermal stress and exposure to clinically relevant antibiotics. In addition we assessed the bacterial and archaeal community via molecular methods (NGS sequencing) and compared our new data with the previously derived information from the ISS microbiome.