Species Capable of Growth at Pressures Ranging Over 5 Orders of Magnitude, from the Surface of Mars (10 Pa) to Deep Oceans (10 Pa) in the Solar System.

Several permanently cold solar system bodies are being investigated with regard to their potential habitability, including Mars and icy moons. In such locations, microbial life would have to cope with low temperatures and both high and low pressures, ranging from ∼10 to 10 Pa on the surface of Mars to upward of ∼10-10 Pa in the subsurface oceans of icy moons. The bacterial genus consists of species that were previously shown to be capable of growth in the absence of oxygen at low temperatures and at either low pressure or high pressure, but to date the entire pressure range of the genus has not been explored.

Environmental Dependence of Competitive Fitness in Rifampin-Resistant Mutants of Bacillus subtilis.

RNA polymerase (RNAP) is a highly conserved macromolecular machine that contributes to the flow of genetic information from genotype to phenotype. In Bacillus subtilis, mutations in the gene encoding the β-subunit of RNAP have been shown to alter a number of global phenotypes, including growth, utilization of unusual nutrient sources, sporulation, germination, and production of secondary metabolites. In addition, the spectrum of mutations in leading to rifampin resistance (Rif) can change dramatically depending upon the environment to which B.

Shelf Life and Simulated Gastrointestinal Tract Survival of Selected Commercial Probiotics During a Simulated Round-Trip Journey to Mars.

To enhance the gastrointestinal health of astronauts, probiotic microorganisms are being considered for inclusion on long-duration human missions to the Moon and Mars. Here we tested three commercial probiotics- strain BB536, strain DDS-1, and spores of strain HU58-for their survival to some of the conditions expected to be encountered during a 3-year, round trip voyage to Mars. All probiotics were supplied as freeze-dried cells in capsules at a titer of >10 colony forming units per capsule.

The Spectrum of Spontaneous Rifampin Resistance Mutations in the Bacillus subtilis Gene Depends on the Growth Environment.

Results from previous investigations into spontaneous rifampin resistance (Rif) mutations in the Bacillus subtilis gene suggested that the spectrum of mutations depends on the growth environment. However, these studies were limited by low sample numbers, allowing for the potential distortion of the data by the presence of "jackpot" mutations that may have arisen early in the growth of a population. Here, we addressed this issue by performing fluctuation analyses to assess both the rate and spectrum of Rif mutations in two distinct media: LB, a complete laboratory medium, and SMM, a minimal medium utilizing l-asparagine as the sole carbon source.

Growth at 5 kPa Causes Differential Expression of a Number of Signals in a Strain Adapted to Enhanced Growth at Low Pressure.

To determine microbial evolutionary strategies to low-pressure (LP; 5 kPa) growth, an environmental condition not experienced on Earth until ∼20 km in altitude, a previously described evolutionary experiment was conducted. The resulting LP evolved strain WN1106, isolated from the terminus of the experiment, was shown to have several genomic mutations absent in the ancestral strain, WN624. Three of the mutations were in regulatory genes: , , and .

Mechanotransduction in Prokaryotes: A Possible Mechanism of Spaceflight Adaptation.

Our understanding of the mechanisms of microgravity perception and response in prokaryotes (Bacteria and Archaea) lag behind those which have been elucidated in eukaryotic organisms. In this hypothesis paper, we: (i) review how eukaryotic cells sense and respond to microgravity using various pathways responsive to unloading of mechanical stress; (ii) we observe that prokaryotic cells possess many structures analogous to mechanosensitive structures in eukaryotes; (iii) we review current evidence indicating that prokaryotes also possess active mechanosensing and mechanotransduction mechanisms; and (iv) we propose a complete mechanotransduction model including mechanisms by which mechanical signals may be transduced to the gene expression apparatus through alterations in bacterial nucleoid architecture, DNA supercoiling, and epigenetic pathways.

Comparisons of Transcriptome Profiles from Cells Grown in Space versus High Aspect Ratio Vessel (HARV) Clinostats Reveal a Low Degree of Concordance.

Although clinostats have long been used in space microbiology studies as ground-based analogs of spaceflight, few studies to date have systematically compared -omics data from clinostats versus spaceflight. This study compared the transcriptomic response of the Gram-positive bacterium flown in space with corresponding transcriptomes derived from 2-D clinostat (High Aspect Ratio Vessel: HARV) experiments performed under the same conditions of bacterial strain, growth medium, temperature, and incubation time. High-quality total RNA (RNA Integrity Number >9.

Nanosatellites for Biology in Space: In Situ Measurement of Spore Germination and Growth after 6 Months in Low Earth Orbit on the Mission.

We report here complete 6-month results from the orbiting Space Environment Survivability of Living Organisms (SESLO) experiment. The world's first and only long-duration live-biology cubesat experiment, SESLO was executed by one of two 10-cm cube-format payloads aboard the 5.5-kg (Organism/Organic Exposure to Orbital Stresses) free-flying nanosatellite, which launched to a 72°-inclination, 650-km Earth orbit in 2010.

A Bumpy Pathway to Stationary-Phase Survival in Bacillus subtilis.

cells can mount a number of responses to nutritional deprivation but ultimately either form dormant spores or enter a metabolically quiescent state. In a recent article (mBio 10:e01414-19, https://doi.org/10.

Spore Resistance to Simulated Mars Surface Conditions.

In a Mars exploration scenario, knowing if and how highly resistant spores would survive on the Martian surface is crucial to design planetary protection measures and avoid false positives in life-detection experiments. Therefore, in this study a systematic screening was performed to determine whether spores could survive an average day on Mars. For that, spores from two comprehensive sets of isogenic mutant strains, defective in DNA protection or repair genes, were exposed to 24 h of simulated Martian atmospheric environment with or without 8 h of Martian UV radiation [M(+)UV and M(-)UV, respectively].

Comparison of transcriptome profiles from two separate missions to the International Space Station.

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 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.

Transcriptomic responses of Serratia liquefaciens cells grown under simulated Martian conditions of low temperature, low pressure, and CO-enriched anoxic atmosphere.

Results from previous experiments indicated that the Gram-negative α-proteobacterium Serratia liquefaciens strain ATCC 27592 was capable of growth under low temperature (0 °C), low pressure (0.7 kPa), and anoxic, CO-dominated atmosphere-conditions intended to simulate the near-subsurface environment of Mars. To probe the response of its transcriptome to this extreme environment, S.

Meta-analysis of data from spaceflight transcriptome experiments does not support the idea of a common bacterial "spaceflight response".

Several studies have been undertaken with the goal of understanding how bacterial transcriptomes respond to the human spaceflight environment. However, these experiments have been conducted using a variety of organisms, media, culture conditions, and spaceflight hardware, and to date no cross-experiment analyses have been performed to uncover possible commonalities in their responses. In this study, eight bacterial transcriptome datasets deposited in NASA's GeneLab Data System were standardized through a common bioinformatics pipeline then subjected to meta-analysis to identify among the datasets (i) individual genes which might be significantly differentially expressed, or (ii) gene sets which might be significantly enriched.

The Photochemistry of Unprotected DNA and DNA inside Bacillus subtilis Spores Exposed to Simulated Martian Surface Conditions of Atmospheric Composition, Temperature, Pressure, and Solar Radiation.

DNA is considered a potential biomarker for life-detection experiments destined for Mars. Experiments were conducted to examine the photochemistry of bacterial DNA, either unprotected or within Bacillus subtilis spores, in response to exposure to simulated martian surface conditions consisting of the following: temperature (-10°C), pressure (0.7 kPa), atmospheric composition [CO (95.

Alterations in the Spectrum of Spontaneous Rifampicin-Resistance Mutations in the Gene after Cultivation in the Human Spaceflight Environment.

The effect of exposure to the human spaceflight environment on growth, mutagenic frequency, and spectrum of mutations to rifampicin resistance (Rif) was investigated. cells were cultivated in Biological Research in Canister-Petri Dish Fixation Units (BRIC-PDFUs) on two separate missions to the International Space Station (ISS), dubbed BRIC-18 and BRIC-21, with matching asynchronous ground controls. No statistically significant difference in either growth or in the frequency of mutation to Rif was found in either experiment.

Cultivation in Space Flight Produces Minimal Alterations in the Susceptibility of Bacillus subtilis Cells to 72 Different Antibiotics and Growth-Inhibiting Compounds.

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 space flight microgravity on the International Space Station, termed flight (FL) samples, or at Earth-normal gravity, termed ground control (GC) samples. The susceptibility of FL and GC samples was compared to 72 antibiotics and growth-inhibitory compounds using the Omnilog phenotype microarray (PM) system.

Experimental evolution of Bacillus subtilis.

The endospore-forming bacteria have persisted on earth perhaps 3Ga, leveraging the flexibility of their distinctive lifestyle to adapt to a remarkably wide range of environments. This process of adaptation can be investigated through the simple but powerful technique of laboratory evolution. Evolved strains can be analyzed by whole genome sequencing and an array of omics technologies.

Twenty Species of Hypobarophilic Bacteria Recovered from Diverse Soils Exhibit Growth under Simulated Martian Conditions at 0.7 kPa.

Bacterial growth at low pressure is a new research area with implications for predicting microbial activity in clouds and the bulk atmosphere on Earth and for modeling the forward contamination of planetary surfaces like Mars. Here, we describe experiments on the recovery and identification of 20 species of bacterial hypobarophiles (def., growth under hypobaric conditions of approximately 1-2 kPa) in 10 genera capable of growth at 0.

An improved high-quality draft genome sequence of Carnobacterium inhibens subsp. inhibens strain K1(T).

Despite their ubiquity and their involvement in food spoilage, the genus Carnobacterium remains rather sparsely characterized at the genome level. Carnobacterium inhibens K1(T) is a member of the Carnobacteriaceae family within the class Bacilli. This strain is a Gram-positive, rod-shaped bacterium isolated from the intestine of an Atlantic salmon.

Cultivation of Staphylococcus epidermidis in the Human Spaceflight Environment Leads to Alterations in the Frequency and Spectrum of Spontaneous Rifampicin-Resistance Mutations in the rpoB Gene.

Bacteria of the genus Staphylococcus are persistent inhabitants of human spaceflight habitats and represent potential opportunistic pathogens. The effect of the human spaceflight environment on the growth and the frequency of mutations to antibiotic resistance in the model organism Staphylococcus epidermidis strain ATCC12228 was investigated. Six cultures of the test organism were cultivated in biological research in canisters-Petri dish fixation units for 122 h on orbit in the International Space Station (ISS) as part of the SpaceX-3 resupply mission.

Twenty-Three Species of Hypobarophilic Bacteria Recovered from Diverse Ecosystems Exhibit Growth under Simulated Martian Conditions at 0.7 kPa.

Unlabelled: Bacterial growth at low pressure is a new research area with implications for predicting microbial activity in clouds and the bulk atmosphere on Earth, and for modeling the forward contamination of planetary surfaces like Mars. Here, we describe experiments on the recovery and identification of 23 species of bacterial hypobarophiles (def., growth under hypobaric conditions of approximately 1-2 kPa) in 11 genera capable of growth at 0.

Anaerobic growth of Bacillus subtilis alters the spectrum of spontaneous mutations in the rpoB gene leading to rifampicin resistance.

Spontaneous rifampicin-resistant (RFM(R)) mutants were isolated from Bacillus subtilis 168 cultivated in the presence or absence of oxygen. By DNA sequencing, the mutations were located within Cluster I of the rpoB gene encoding the β subunit of RNA polymerase. The spectrum of RFM(R) rpoB mutations isolated from B.

Synthetic operon for (R,R)-2,3-butanediol production in Bacillus subtilis and Escherichia coli.

To reduce dependence on petroleum, an alternative route to production of the chemical feedstock 2,3-butanediol (2,3-BD) from renewable lignocellulosic sources is desirable. In this communication, the genes encoding the pathway from pyruvate to 2,3-BD (alsS, alsD, and bdhA encoding acetolactate synthase, acetolactate decarboxylase, and butanediol dehydrogenase, respectively) from Bacillus subtilis were engineered into a single tricistronic operon under control of the isopropyl β-D-1-thiogalactopyranoside (IPTG)-inducible Pspac promoter in a shuttle plasmid capable of replication and expression in either B. subtilis or Escherichia coli.

Experimental evolution of enhanced growth by Bacillus subtilis at low atmospheric pressure: genomic changes revealed by whole-genome sequencing.

Knowledge of how microorganisms respond and adapt to low-pressure (LP) environments is limited. Previously, Bacillus subtilis strain WN624 was grown at the near-inhibitory LP of 5 kPa for 1,000 generations and strain WN1106, which exhibited increased relative fitness at 5 kPa, was isolated. Genomic sequence differences between ancestral strain WN624 and LP-evolved strain WN1106 were identified using whole-genome sequencing.

Evolution in the Bacillaceae.

The family Bacillaceae constitutes a phenotypically diverse and globally ubiquitous assemblage of bacteria. Investigation into how evolution has shaped, and continues to shape, this family has relied on several widely ranging approaches from classical taxonomy, ecological field studies, and evolution in soil microcosms to genomic-scale phylogenetics, laboratory, and directed evolution experiments. One unifying characteristic of the Bacillaceae, the endospore, poses unique challenges to answering questions regarding both the calculation of evolutionary rates and claims of extreme longevity in ancient environmental samples.