Graduation Term

Summer 2026

Degree Name

Master of Science (MS)

Department

School of Biological Sciences

Committee Chair

Jan-Ulrik Dahl

Committee Co-Chair

Brian J. Wilkinson

Committee Member

Benjamin Sadd

Committee Member

Kyle Floyd

Abstract

Staphylococcus aureus is a prominent pathogen that is known to incorporate straight-chain unsaturated fatty acids (SCUFAs) into its lipids in addition to the normally biosynthesized branched-chain fatty acids (BCFAs) and straight-chain fatty acids (SCFAs). Furthermore, it has recently been shown that the incorporation of oleic acid into S. aureus lipids significantly enhanced the growth of the organism at low temperatures due to the greater fluidity imparted to the membrane. Together with others in the Wilkinson and Dahl laboratory, I have shown that low-temperature growth is enhanced by various SCUFAs, including the human antimicrobial fatty acids sapienic acid, linoleic acid, and arachidonic acid, at low concentrations. In a second project, which is the foundation of my master’s thesis, I focused on the role of cholesterol for bacterial membrane physiology during bacterial exposure to SCUFAs, which can exhibit potent antimicrobial activity by fluidizing and disrupting the bacterial membrane. Here, I show that cholesterol, typically only associated with the eukaryotic plasma membrane, plays a crucial role in modulating membrane properties during SCUFA exposure, influencing bacterial survival and potentially adaptation and virulence. While bacteria do not synthesize cholesterol de novo, some species incorporate host-derived cholesterol, impacting membrane fluidity, permeability, and interactions with antimicrobial agents. However, the precise mechanisms by which host cholesterol modulate SCUFA’s toxicity and influences bacterial fitness remain poorly defined. Hence, in this study, I have investigated the protective role of exogenous cholesterol against SCUFA-induced stress in S. aureus. Here, the bacterial growth, survival, and membrane dynamics using checkerboard assays, Nile Red membrane fluidity tracking, and high-resolution confocal microscopy with fluorescent TopFluor-Cholesterol (TF-Chol) have been evaluated. I have shown that supplementation with physiological concentrations of cholesterol or its esterified counterparts (cholesteryl arachidonate and cholesteryl linoleate) completely rescued S. aureus from SCUFA-mediated growth inhibition and restored survival by up to 5 logs. Nile Red assays confirmed that exogenous cholesterol counteracts fatty acid-induced membrane disordering. Furthermore, I discovered that sapienic acid actively drives a robust, dose-dependent acceleration of host cholesterol internalization. Flow cytometry and confocal imaging revealed that with increasing concentration of sapienic acid the cholesterol acquisition in the cells increases. Together, these findings aim to reveal a lipid-remodeling strategy wherein S. aureus actively harnesses host cholesterol to stabilize its membrane, neutralize the unsaturated fatty acid toxicity, and adapt to lipid-rich host niches like blood and tissue.

Access Type

Thesis-Open Access

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