First Name:
Samuel
Last Name:
Pendergraft
Major Department:
Biology
Thesis Director:
Ece Karatan
Date of Thesis:
May 2010
Biofilms exist as coordinated enchained bacterial communities of self-excreted exopolysaccharide matrices. Biofilm formation is important for the survival of the pathogenic bacterium Vibrio cholerae both in its natural aquatic environments and the human host. Development of biofilms is partly regulated by small organic molecules known as polyamines. Previous work in the Karatan lab has also shown that overexpression of the gene (nspC) encoding the NspC enzyme, which is responsible for the synthesis of the polyamine norspermidine, leads to a large increase in biofilm formation by an unknown mechanism.
The objective of this project is to explore the role of nspC gene overexpression in V. cholerae biofilm phenotype. To do this I wanted to explore the interplay between polyamine transport pathways, which also regulate biofilm formation, and biofilm synthesis pathways. I constructed a bacterial mutant that was overexpressing the nspC gene but missing the PotD1 polyamine transporter. I was able to confirm through western blotting that the V. cholerae nspC overexpression strain indeed has significantly high levels of NspC protein within the bacterium. Biofilm assays conducted using this new strain showed that presence of the transporter is necessary for the increase in the biofilm seen as a result of nspC overexpression, contrasting hypothesized effects. In addition, motility assays, used to see if the swimming ability of bacteria is affected, revealed a significant decrease in motility of the nspC overexpression strain. As motility and biofilm formation are usually inversely regulated, experimental data contrary to this perception provides valuable information as to the mechanisms by which polyamine synthesis and transport pathways regulate biofilm formation.