Date of Award:

5-2009

Document Type:

Dissertation

Degree Name:

Doctor of Philosophy (PhD)

Department:

Biology

Committee Chair(s)

Jon Y. Takemoto

Committee

Jon Y. Takemoto

Committee

Anne Anderson

Committee

Michelle Grilley

Committee

Brad Kropp

Committee

Marie K. Walsh

Abstract

The need for new antimicrobial agents has become important in the last decade due to emerging resistance to a number of conventional antimicrobial agents. New approaches and sources are needed to generate novel and effective antimicrobials. For example, synergistic combinations between two or more agents may lead to new antimicrobial therapies. Furthermore, the increase in health problems caused by the exposure to agricultural crop pesticides and synthetic fungicides and the emerging development of organic farming has increased the necessity to develop natural products than can be used safely in controlling crop diseases.

In this work, I present the first studies on the bioactive properties, particularly fungicidal activities, of mixtures of SRE and rhamnolipids. The in vitro results clearly showed strong synergism between SRE and rhamnolipids against phytopathogenic fungi and yeast. However, no activity was observed against bacteria. The hemolytic activities and cytotoxicities of SRE and SYRA were dose dependent.

SRE acts on yeast and plant plasma membranes to cause numerous cellular effects. The effects are consistent with SRE's ability to form ion-conducting voltage sensitive channels in membrane bilayers. In addition, studies with yeast have revealed that sphingolipids and sterols modulate the fungicidal activity of SRE. Saccharomyces cerevisiae sphingolipid and sterol biosynthetic mutants were used to investigate the mechanism of action of SYRA against fungi. These results suggest that similar to SRE, SYRA antifungal action is promoted by sphingolipids and sterols of the plasma membrane and involves pore formation.

I further explored the antimicrobial spectrum of syringopeptin SP25A and show that it specifically inhibits Gram-positive bacteria and yeast. I also investigated its mechanism of action against yeast and bacteria. The results revealed the role for D-alanylation of teichoic acids in modulating the susceptibility of B. subtilis to SP25A and other syringopeptins. This is consistent with the charged nature of the cyclic peptide portions of the syringopeptins, and it provides an explanation for SP25A's higher degree of specificity for Gram-positive bacteria. In addition and similar to SRE, SP25A antifungal action is promoted by sphingolipids and sterols of the plasma membrane and involves pore formation.

Overall, the research shows that SRE and rhamnolipids are synergistically active against yeast and fungi and that the syringopeptins have antimicrobial activities against yeast and Gram-positive bacteria. Insights into the mechanisms of action of the SRE and rhamnolipid mixtures and the syringopeptins and their potential as novel antimicrobial agents are revealed.

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