Glycan Triggers of Life Cycle Development in the Apicomplexan Parasite Cryptosporidium

Cryptosporidium is an apicomplexan parasite that causes the diarrheal disease cryptosporidiosis, an infection that can become chronic and life threating in immunocompromised and malnourished individuals. Development of novel therapeutic interventions is critical as current treatments are entirely ineffective in treating cryptosporidiosis in populations at the greatest risk for disease. Repeated cycling of host cell invasion and replication by sporozoites results in the rapid amplification of parasite numbers and the pathology associated with the disease. Little is known regarding the factors that promote the switch from invasion to replication of Cryptosporidium, or the mechanisms underlying this change, but identification of replication triggers could provide potential targets for drugs designed to prevent cryptosporidiosis. The focus of this dissertation was to identify potential triggers and the mechanisms underlying the transition from invasive sporozoite to replicative trophozoites in Cryptosporidium. We demonstrate glycoproteins secreted by host cells promote the transition from invasion to replication in Cryptosporidium, and free Gal-GalNAc triggers nuclear division in linear and rounded sporozoites. The proportion of rounded and multinucleated cells in response to the host secretions differed between C. parvum and C. hominis, the two species of greatest concern to human health, with C. parvum sporozoites progressing towards rounding and replication faster. We demonstrated the use of glycomimetic polymers for studying Cryptosporidium biology. Slides imprinted with increasing densities of Gal and GalNAc had the greatest proportion of trophozoite development. Gal and GalNAc glycomimetic polymers were able to cause parasite replication; however the effect was less than what was observed in the presence of secreted host glycoproteins and free Gal-GalNAc. We also characterized a rhomboid protease (ROM) in Cryptosporidium which we name CpROM2, providing information regarding expression and localization of the protease during C. parvum development. Using inhibitors specific for ROM activity we show ROMs play a critical role in excystation, but are also required for the transition from invasion to replication in C. parvum. These findings have strengthened our understanding of how Cryptosporidium transitions from invasion to replication by identifying host glycoproteins and Gal-GalNAc as triggers for trophozoite development via a ROM driven mechanism.NIH 1R15AI122152-01A1North Dakota State University. Graduate Schoo