Graduation Date

Summer 8-17-2018

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Programs

Pathology & Microbiology

First Advisor

Paul Fey, Ph.D

Abstract

Biofilm formation is the primary virulence factor in Staphylococcus epidermidis. Polysaccharide intercellular adhesin (PIA) is an adhesive molecule and a significant component of the biofilm matrix. It is synthesized by the products of the icaADBC operon whose regulation has been shown to involve environmental factors as well as many transcriptional regulators. Of these regulators, we explored the function of the repressors IcaR and TcaR and their roles in directly influencing icaADBC transcription and PIA synthesis. Based on previous observations that icaADBC positive clinical isolates of S. epidermidis are highly variable in PIA synthesis and biofilm formation, our goal was to further investigate why this may be. We generated icaR and tcaR mutations in S. epidermidis strain 1457, a high PIA producing strain, and CSF41498, a low and inducible PIA producing strain. We observed that icaADBC is primarily regulated by TcaR in 1457 and by IcaR in CSF41498 and this may be due to icaR being expressed at lower levels in 1457, leading to de-repression of icaADBC. DNase I footprinting results confirmed that TcaR binds to multiple sequences in the icaR-icaA intergenic region, including the ica and icaR promoters, providing evidence that TcaR can repress both icaADBC and icaR transcription. Finally, we generated mutants in CSF41498 exhibiting high PIA synthesis as well as mutants in 1457 that were no longer able to synthesize high levels of PIA. Sequencing of these mutants provided insight into genes with potential functions in regulating icaADBC. Overall, our data demonstrate the complexity with which icaADBC is regulated, especially in regards to IcaR and TcaR, and that icaADBC regulation is strain specific.

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