Understanding responses to chemical and environmental perturbations in blood stage Babesia parasites

Abstract

Babesiosis is a disease caused by apicomplexan parasites related to the malaria parasite Plasmodium. Like Plasmodium, Babesia infects red blood cells (RBCs) in the host animal. Babesia is one of the most widespread blood parasites of vertebrates, second only to the trypanosomes. About 130 years since the discovery of Babesia in cattle with hemolytic febrile illness (1888), and despite rising morbidity and economic impact due to this infection, basic parasite biology remains poorly understood. Further, since the first reported case of human babesiosis in 1956, compounded by the emergence of Babesia microti as a major zoonotic threat, Babesia has steadily been gaining recognition as an important human parasitic disease. The disease can range from mild febrile illness to severe, life threatening disease. Yet, despite its growing importance, there continues to be no FDA approved treatment solely for babesiosis in either humans or animals. Further, understanding of the basic biology such as intraerythrocytic development cycle severely lags behind that of related parasites like Plasmodium and Toxoplasma. Finally, Babesia has recently been classified as a top threat to the blood supply for its ability to be effectively transmitted via blood transfusion. The goals of this dissertation work were to unravel the basic biology of the parasite in order to facilitate novel drug discovery and understand the dynamics of transfusion transmission, thus addressing the three major challenges in the control of babesiosis. In chapter one, I review the current state of knowledge in the Babesia field as it pertains to drug discovery, cellular development, and transfusion transmission. Additionally, I review mechanisms in other studied cellular systems pertinent to these challenges in Babesia, including mechanisms of quiescence. In chapter two, I aimed to address the major challenge of ineffective treatments for Babesia by developing and validating a comparative chemical genomics pipeline for rapid discovery of novel drug targets. Through this, I was able to identify a novel druggable target, the alkaline phosphatase phoD, in Babesia divergens. I used genetic tools to validate this target as the causative agent in resistance to a novel antibabesial compound, MMV019266, and localized phoD in the cell, as well as began to interrogate the mechanism of action. In chapter 2, we worked to describe the replicative cycle at the molecular level using single cell RNA sequencing. We were able to describe the replicative cycle of three species of Babesia: B. divergens, B. bovis, and B. bigemina. We revealed both conserved and divergent markers of cellular development, and were able to take advantage of B. divergens ability to grow in both human and bovine erythrocytes to interrogate differences in the parasites based on the host cell. In chapter 4, I explored the mechanisms underlying cold tolerance in B. divergens, the feature that allows the parasite to be successfully undergo transfusion transmission. Here, in collaboration, we identified transcriptional and epigenetic mechanisms involved in cold induced quiescence. We were also able to identify canonical mechanisms of quiescence, as well as possible divergent apicomplexan biology. In chapter 5, I discuss the implications of this work, detail some preliminary results based on the foundations built in this dissertation, and discuss vital future directions to be taken. Overall, this work has contributed important resources to the Babesia community and addressed the three essential challenges: lack of an effective treatment, limited information about basic molecular parasite biology, and transfusion transmission. Both the molecular tools and data sets generated and this work will greatly facilitate future investigations into parasite biology.