Date of Award
Doctor of Philosophy (PhD)
The parasitic protist Toxoplasma gondii is a common pathogen of rodents and felines that also infects humans. The most severe clinical manifestations of toxoplasmosis in humans derive from the systemic dissemination of T. gondii, during which the parasite penetrates biological barriers and accesses protected host compartments such as the central nervous system. T. gondii dissemination is enabled by the intrinsic gliding motility of extracellular parasites, which allows for travel to new host cells and tissues, and also powers the invasion of diverse host cells including migratory leukocytes. Dissemination is further advanced when migrating infected leukocytes shuttle intracellular parasites to new locations as they traffic throughout the host.
All T. gondii gliding motility and host cell invasion was long presumed to be powered by the work of a parasite actin-myosin motor. The possibility of alternative gliding and invasion mechanisms was suggested by the development of inducible Cre-Lox technology that facilitated inducible disruption of genes thought to encode critical components of the T. gondii invasion machinery, including the parasite actin gene ACT1. To determine whether ACT1-independent invasion was likely, inducible Δact1 parasites were examined for uniformity of ACT1 protein depletion. Individual parasites with residual ACT1 protein persisted long after inducible ACT1 excision. Suggesting the residual ACT1 content of these parasites was functionally relevant, the invasion of Δact1 parasites was highly sensitive to an actin polymerization inhibitor. Parasite invasive ability was also found to negatively correlate with the length of time parasites were subjected to ACT1 depletion. Although the existence of ACT1-independent invasion mechanisms cannot be formally excluded, they do not appear to comprise robust alternatives to actin-dependent gliding and invasion in T. gondii.
As the most abundantly infected circulating leukocyte during murine toxoplasmosis, monocytes have been theorized to be poised to deliver intracellular T. gondii across the blood-brain barrier and into the central nervous system. However, in vivo evidence supporting this theory was scarce. In vitro models had demonstrated that infection could alter the motility of monocytes when interacting with endothelial vasculature. However, whether infected monocytes could efficiently traverse the specialized endothelium that comprises the blood-brain barrier had not been tested, nor had the ability of infected monocytes to migrate through the tissue environments where T. gondii is first encountered. Models of peripheral and blood-brain barrier endothelium were used to show that infection markedly inhibited monocyte transendothelial migration. In contrast, infected monocytes and macrophages migrated through three-dimensional matrices in vitro and collagen-rich tissues in vivo with enhanced efficiency. Enhanced tissue migration relied on host Rho/ROCK and formin signaling, and the secreted T. gondii kinase ROP17. In a murine model, infection with Δrop17 parasites that fail to enhance tissue migration resulted in delayed dissemination and prolonged mouse survival. These results implicate monocytes in advancing the tissue spread of T. gondii during in vivo dissemination.
Chair and Committee
L D. Sibley
John A. Cooper, Tamara L. Doering, Daniel E. Goldberg, Robyn S. Klein,
Drewry, Lisa L., "Dissemination of the apicomplexan parasite, Toxoplasma gondii" (2019). Arts & Sciences Electronic Theses and Dissertations. 1784.