Defining the cell and molecular basis of Toxoplasma recrudescence

Project Summary Reactivation of toxoplasmosis is a significant health threat to people chronically infected with this parasite and is life-threatening to infected individuals that are or become immunocompromised. Millions of people face this threat as it is estimated one third of human populations are infected with this pathogen. Recrudescence of the Toxoplasma bradyzoite tissue cyst is the cause of toxoplasmosis reactivation, which can not be prevented as there is no current treatment that eliminates the dormant tissue cyst in chronically infected individuals. Approaches to find therapeutic solutions to treat and prevent chronic toxoplasmosis have suffered from limited accessibility to the relevant Toxoplasma stages and a lack of accurate in vitro developmental models. Our goal in this proposal is to breakthrough these impasses. We have developed a new innovative ex vivo model of bradyzoite recrudescence that we will utilize to define the host cell specificity (Aim 1a), whole-cell gene expression (Aim 1b) and metabolic changes (Aim 2) that unfold when a bradyzoite converts back to the tachyzoite and also in a newly discovered alternate pathway where bradyzoites directly replicate to reform the tissue cyst. This information is critically needed in order to understand how we might prevent toxoplasmosis reactivation. The loss of developmental competency in vitro that is exacerbated in current protocols producing transgenic strains is also a major impediment to understanding the molecular basis of tissue cyst reactivation. In this proposal, we will implement and optimize an innovative approach to generate developmentally competent transgenic strains (Aim 3a), and use this new protocol to define cyclin and other protein mechanisms (Aim 3b) that have critical roles in regulating bradyzoite recrudescence and tissue cyst re-formation