Type 1 fimbrial structure and regulation in Salmonella enterica serovar Typhimurium

has been approved by the Examining Committee for the thesis requirement for the Master of Science degree in Microbiology at the May 2012 graduation. ii ACKNOWLEDGEMENTS I would like to thank Dr. Steve Clegg for his support and guidance during my time in his lab. He has helped me learn a lot about how to do science and how to think about science. He has been supportive of the decisions I have made and I have enjoyed my time in the lab. I would also like to thank my committee members Dr. Craig Ellermeier and Dr. Brad Jones for their time and helpful advice. The members of the Clegg Lab have been sources of guidance, advice, and friendship during my time here. Jean Sippy have made the lab an interesting place to work and I will always be grateful to them for that. I must also thank my family, who has always been very supportive of me. They are always interested in my work and have helped me through all my years of education. iii ABSTRACT Salmonella enterica serovar Typhimurium is a common cause of bacterial food poisoning, and S. Typhimurium expresses type 1 fimbriae that enable the bacteria to bind to eukaryotic cells. Fimbrial proteins are encoded by the fim gene cluster (fimAICDHFZYW). The structural components of the fimbriae are FimA (major subunit), FimI, FimH (adhesin), and FimF (adaptor). In order to determine the genes required for fimbrial assembly in S. Typhimurium SL1344, mutations in fimA, fimI, fimH, and fimF were constructed and examined for their ability to produce fimbriae. While SL1344∆fimI was able to assemble fimbriae, SL1344∆fimA, ∆fimH, and ∆fimF were afimbriate, indicating that fimA, fimH, and fimF are each required for fimbrial formation in S. Typhimurium. These results suggest differences in the genetic requirements when comparing S. Typhimurium type 1 fimbrial and E. coli type 1 and Pap fimbrial systems. S. Typhimurium fim gene regulation was also examined. FimZ and FimY are positive regulators of fimbrial gene expression, and FimW is a negative regulator. FimZ is closely related to the family of response regulators of two-component systems. The response regulator activity of FimZ was examined by substituting the conserved aspartate-56 residue, the putative site of phosphorylation, with alanine, to generate an inactive phosphorylation site, or glutamate, to mimic a phosphorylated protein. Resulting strains were examined for fimbrial production and gene expression. It was observed that when the …