Date of Award

Summer 6-8-2018

Level of Access Assigned by Author

Open-Access Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Microbiology

Advisor

Robert Wheeler

Second Committee Member

Julie Gosse

Third Committee Member

John Singer

Additional Committee Members

Roger Sher

Rebecca Van Beneden

Abstract

Candida albicans is a common hospital-acquired fungal infection, and disseminated disease claims up to one-half of those afflicted. C. albicans has a unique ability to differentiate its shape during infection, and this differentiation is thought to be a major virulence factor during invasive infections. Each shape is proposed to have a specialized role: filaments drive tissue invasion and yeast mediate dissemination to the bloodstream. However, it has been difficult to test these hypotheses for two reasons. First, rigorous testing of shape-specific roles requires diverse strategies of shape modulation that restrict the possibility of manipulation-specific artifacts. Second, although connecting shape to function requires simultaneous visualization of shape and location of fungi during infection, this is not feasible in opaque mammalian infection models. Thus, we aimed to more comprehensively analyze how C. albicans utilizes shape to disseminate and invade during infection by using a larval zebrafish infection model. This model permitted the use of diverse, complementary strategies to manipulate shape, subsequently allowing us to monitor dissemination, invasion, and overall pathogenesis using intravital imaging of individual fungal cells throughout the host. To control cell shape, we employed three different strategies: filament-specific gene deletion, overexpression of yeast or filament master regulators, and alteration of infection temperature. The effects of these orthogonal manipulations were consistent, support proposed specialized roles of yeast in dissemination and filaments in tissue invasion and pathogenesis, and indicate conserved mechanisms in zebrafish. Based on the conservation of cell type-specific functions in zebrafish infection, we tested whether either morphotype changes the effectiveness of the other cell type. By infecting fish with a known mix of shape-locked strains, we surprisingly found that mixed infections were associated with additive, but not synergistic, filament invasion and yeast dissemination. Taken together, these findings provide rigorous confirmation of previous hypotheses linking shape and pathogenesis in a new infection model, but also reveal the independent roles of yeast and filaments during disseminated candidiasis. Overall, we hope that characterizing cell type-specific roles during infection can tailor novel therapies to limit both dissemination and pathogenesis, and efficiently target C. albicans in patients who would otherwise succumb to such deadly infections.

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