Understanding How Bacteria Sense Mechanics Upon Attaching to Surfaces

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Biofilms are communities of microbes that are bound to each other and to a surface by a matrix of polymers and proteins. They foul and corrode infrastructure and cause chronic infections. At present, approaches to making surfaces that prevent infection have met with only limited success. The goal of this project is to determine how bacteria sense the mechanics of the surface upon attachment and respond to surface mechanics by starting to develop a biofilm. This work will establish basic knowledge that can be used to design surfaces that resist the development of biofilms by not giving bacteria the mechanical cue(s) needed to start forming a biofilm. Thus, the scientific aims will improve biofilm prevention and thereby benefit public health and infrastructure such as water treatment, shipping, and oil transport. Mentoring and education in ethics and mindset will improve the undergraduate environment and pipeline for STEM majors. A study of classroom education methods will improve undergraduate STEM education. The research objective of this project is to develop a predictive framework for understanding how bacteria use proteins in their cell envelopes to sense and respond to the mechanics of the surface to which they attach. The working hypothesis is that bacteria do this by transducing stresses and deformations in their envelopes that depend on the elasticity of both the bacterium and the substrate as well as the energy of adhesion with the surface. This study will extend studies of P. aeruginosa and generalize to include another rod-shaped Gram-negative bacterium (Escherichia coli), a rod-shaped Gram-positive bacterium (Bacillus subtilis), and a spherical Gram-positive bacterium (Staphylococcus aureus). Atomic force microscopy (AFM) will be used to measure the adhesion forces between bacteria and a wide range of test substrates with different surface chemistries and elasticities; AFM will also characterize the elasticity and topography of substrates. High-throughput culturing in well plates will be used to measure bacterial accumulation and growth on substrates, as well as the increase in antibiotic resistance that is a biofilm phenotype. Finite-element modeling will characterize the stresses and deformations that arise when different types of bacteria attach to different substrates. Trends shown by modeling will be validated by measuring the activity of mechanosensitive ion channels that are more open when the membrane stress is greater. Quantitative confocal microscopy and image analysis will measure the intracellular signaling that arises as a result of bacteria attaching to surfaces and the intercellular signaling that arises as a result of bacterial growth on the surface. Genetic manipulation (reporter strains and isogenic knockouts) will be used to measure signaling and to elucidate the role of specific gene products, such as candidate mechanosensory proteins.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.