Design and Implementation of Microfluidic Systems for Bacterial Biofilm Monitoring and Manipulation

Abstract

Bacterial biofilms - pathogenic matrices formed through bacterial communication and subsequent extracellular matrix secretion - characterize the majority of clinical bacterial infections. Biofilms exhibit increased resistance to conventional antibiotics, necessitating development of alternative treatments. Standard microbiological methods for studying biofilms often rely on in vitro systems with involved instrumentation for biofilm quantification, or destroy the biofilm in the process of characterization. Additionally, biofilm formation is sensitive to many growth parameters, and can exhibit a large degree of variability between repeated experiments. This dissertation presents the development of systems designed to address these challenges through integration of continuous biofilm monitoring in a microfluidic platform, and through creation of a microfluidic platform for multiple assays performed on one biofilm formed in a single channel. The microsystems developed in this work provide building blocks for developing controlled, high throughput testbeds for development and evaluation of drugs targeting bacterial biofilms. The first platform developed relied on optical density monitoring as a means for evaluating biofilm formation. This method was noninvasive, as it used an external light source and array of photodiodes to evaluate biofilms by the amount of light transmitted through the microfluidic channel where they were grown. The optical density biofilm measurement method and microfluidic platform were used to evaluate the dependence of biofilm formation on quorum sensing, an autoinducer-mediated intercellular communication process. This system was also used in the first demonstration of biofilm inhibition and reduction by two different autoinducer-2 analogs. The second microfluidic system developed addressed the challenge of variability in biofilm formation. Biofilms formed in a single microfluidic channel were partitioned by hydraulically actuated valves into three separate segments, which were then treated as representatives of the original biofilm in further experiments. A novel photoresist passivation process was developed in order to create the multi-depth channels needed to accommodate both valve actuation and biofilm formation. Biofilms grown in the device were uniform throughout, providing reliable experimental controls within the system. Biofilm partitioning was demonstrated by exposing three segments of one biofilm to varying detergent concentrations.