Characterization of Biofilm Formation, Growth and Gene Expression on Different Materials and Environmental Conditions in Microgravity

1. To use microbial strains that are relevant to human spaceflight, and nosocomial infections that have also occurred in spaceflight, and substrata materials that are relevant to (i) spacecraft structures and surfaces, (ii) life support systems, (iii) space biology research hardware, and (iv) medical instrumentation. We are meeting this objective by the use of the same fungal strain that was isolated from a bio-damaged Mir window (Penicillium rubens), and an uropathogenic bacterial strain (Pseudomonas aeruginosa). 2. To culture the selected microbial strains in a way that enables biofilm formation, during an up- to-6-month experiment. To achieve this goal, we used BioServe’s already spaceflight- proven hardware and conducted preliminary testing during the first year of the experiment. 3. To quantify potential changes on biofilm mass, thickness, and overall morphology (including the formation of the “column-and-canopy structures) on the spaceflight cultures with respect to matched ground controls. This was achieved by acquiring post-flight (samples fixed in paraformaldehyde) morphological data through confocal laser scanning microscopy. 4. To elucidate the molecular mechanisms behind the observed morphological changes. This was done with the bacterial samples by acquiring post-flight transcriptomic data (samples fixed with RNA Later II) and conducting differential gene expression analyses between spaceflight cultures and matched ground controls. 5. To assess differential expression of genes associated with conferring microbes with oxidative, acidity, and antimicrobial resistance. This was done through RNA Sequencing and comparative gene analysis. 6. To determine physical mechanisms of material/microorganisms interaction in biofilms and their associated molecular basis. The morphological and transcriptomic data was utilized to identify potential correlation in between the differential gene expression and altered material/microorganism interaction in space, with respect to matched ground controls. Different topographic structures were assessed to provide further insight into this aspect (topographical patters with periodicities, same size, as the microbial size).