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This research focuses on the development of new techniques for studying the physical and molecular basis of behavior in swarming strains of bacteria and their connection to biofilm formation. Swarming is a phenotype that was first observed in bacteria over a century ago, and is recognized to play a role in the pathogenesis of bacteria in animals and humans, and yet, very little is known about the mechanisms that regulate coordinated cell growth behavior and their emergence into biofilms. <P>We propose to carry out a systematic study of the influence of the chemical and physical properties of the microenvironment--that is, the immediate extracellular environment--on the differentiation and behavior of swarming cells. <P>A central component in these studies will be the development of microtools: microfabricated polymer structures that are transparent to ultraviolet and visible light, with dimensions on the order of the intrinsic length scale of cells. These polymer systems will make it possible to study the interactions of individual cells with other cells, and with surfaces, by confining swarming colonies of bacteria into two-dimensional cellular structures that can be imaged directly using time-resolved microscopy.<P> The interdisciplinary approaches described in this proposal will improve our understanding of swarming, and may suggest new mechanisms for inhibiting the growth movement of bacteria on surfaces and the formation of biofilms and other multicellular structures.
Non-Technical Summary: We still understand very little about how populations of bacteria coordinate their growth and behavior, and yet, these phenotypes play a critical role in plant and human infections and pathogenesis. This research will uncover fundamental mechanisms that bacteria use to become pathogenic and will help us design and implement new mechanisms for controlling bacteria-based infections. <P> Approach: We are creating microfluidic systems for controlling the microenvironment around swarming cells. We have developed microfluidic systems embossed in the surface of the transparent polymer poly(dimethylsiloxane) (PDMS) that confine swarming colonies of several genera of bacteria into two-dimensional structures and control the microenvironment around these cells. Confining cells in swarming colonies into a single focal plane makes it possible to track and analyze the movement of individual, and small groups of cells; this characteristic also makes it possible to measure swarming quantitatively. By controlling the microenvironment around cells, we will manipulate the chemical and physical interactions between cells and their environment and understand what factors are important for coordinating the growth and motility of bacteria on surfaces. We are also interested in the connection between surface motility and biofilm formation. To study the connection between these two phenotypes, we are developing a technique for patterning individual or small groups of cells on surfaces and allowing them to grow, spread, and mature into biofilms. The technique involves using thin elastomeric stencils with patterns of holes that define where cells can and cannot adsorb to a surface. Using this technique we have demonstrated that this approach makes it possible to create biofilm arrays in which the biofilms are structurally identical. The structures created using this technique are reproducible, making it an important tool for systematically studying the stimuli that regulate biofilm formation.