Phenotypic Changes Occurring in Escherichia coli Cells Previously Exposed to Simulated Microgravity

As humans explore space, bacteria will not only be present but will also be adapting to the extreme environment of space. During space missions astronauts have been shown to be immunocompromised which makes it imperative to understand the effects of space stressors (i.e., increased background radiation and microgravity(μg)) on bacteria. Current research has demonstrated an increase in virulence, biofilm formation, and antibiotic resistance. This research aims to identify multiple phenotypical changes that occur in E. coli cells exposed to simulated μg after 24 hours, 4 days, and 22 days. Specifically, this work observes changes in antibiotic resistance, osmotic tolerance, acidic tolerance, and oxidative stress. Preliminary experiments with antibiotic resistance utilizing the Kirby Bauer disc diffusion technique and the Minimum Inhibitory Concentration (MIC) method have demonstrated that exposure to simulated μg has a direct correlation of increased antibiotic resistance to some antibiotics with the increased duration of exposure. Additionally, increasing osmotic concentration inhibits the growth until four percent of sodium chloride and further increases have completely inhibited growth. These results can potentially be utilized for controlling growth of pathogenic bacteria in space. Additional studies using these two experiments as well as oxidative and pH stress will be explored

Space Microbial Ecology

With the expansion of human space exploration, there is a growing demand to better understand the impacts of space stressors. These stressors include microgravity (µG), space radiation, extreme temperatures, and extreme isolation. Ongoing research has demonstrated that the space environment alters the physiology of bacteria. The changes observed have included increases in biofilm formation, antibiotic resistance, and growth rate. Understanding the effects on bacteria in these conditions is vital as they can affect astronaut health, spacecraft life support systems, and space crops used for food. The ERAU Space Microbiology Lab (SML) is working to identify how microbial communities are impacted by simulated µG. In natural microbial communities (e.g., human gut microbiome), bacteria can develop antagonistic or synergistic relationships between different species. Based on what we know about the response of individual species to space conditions, their interaction with other species and the host can change as well. By observing community development in simulated µG, we can gain insight on how microbial communities naturally adapt to the space environment. Our research is focused on observing the changes of a mixed culture of two bacteria subjected to simulated µG using the EagleStat, a microgravity analog developed by the SML. The mixed culture consists of Escherichia coli and Staphylococcus epidermidis bacteria due to the ability to separate the bacteria visually and physically after simulated µG exposure. Bacterial response will be evaluated by colony composition, biofilm development, antibiotic resistance, and differential gene expression of biofilm and virulence related genes

Determination of free amino acids in plants by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS)

A robust and sensitive method for identification (quantification and confirmation) of 19 free amino acids in the plant matrix, Stellaria media, based on liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS), with a triple quadrupole analyser, has been developed. Regarding MS optimization, flow injection analysis (FIA) was used in scan and selected reaction monitoring (SRM) mode. The collision energies optimized varied from −12 to −39 eV. The acquisition of three MS/MS transitions for most of the compounds allowed the accurate confirmation of these analytes, which was supported by the accomplishment of ion intensity ratios and retention time as compared with the corresponding standards. The use of a Phenomenex EZ:faast™ Free (Physiological) Amino Acid kit speeds up the sample preparation immeasurably. Nineteen amino acids were separated within 18 minutes on a reverse-phase column under a gradient stepwise programme using 10 mM ammonium formate both in water and methanol. The detection limit (LOD) for free amino acids varied from 0.4 to 9.1 pmol mL−1, except for asparagine amounting to 3000 pmol mL−1. The quantification precision (RSD) of free amino acids for intra- and interday assays was 0.05–19% and 0.2–19%, respectively, but for most of the compounds, it did not exceed 5%. The optimized and validated method was subsequently utilized for free amino acid identification in weed collected from field locations in Poland.National Science Centre: UMO-2013/09/N/NZ9/0196