Macrophage – cryptococcus interactions during cryptococcosis

The human fungal pathogens Cryptococcus neoformans and C. gattii cause life-threatening infections of the central nervous system. One of the major characteristics of cryptococcal disease is the ability of the pathogen to parasitise upon phagocytic effector cells. Cryptococcus can survive and proliferate within macrophages, and is also capable of escaping into the intracellular environment via a non-lytic mechanism (‘expulsion’) and can be transferred directly from one cell to another (lateral transfer). In the first part of this thesis, I demonstrate that enhanced Th2, but not Th1, cytokine levels lead to increased intracellular cryptococcal proliferation but lower levels of cryptococcal expulsion. In the second part, I describe the generation and characterisation of GFP-expressing derivates of two widely used cryptococcal strains: C. neoformans serotype A type strain H99 and C. gattii serotype B type strain R265. Furthermore, I have developed a method to effectively and rapidly investigate macrophage parasitism by flow cytometry that preserves the accuracy of current approaches but offers a four-fold improvement in speed. The final part dissects the regulation and induction of mitochondrial tubularisation in hypervirulent C. gattii strains and describes the first steps towards a comparative mitochondrial genome sequencing approach to identify the underlying molecular mechanisms