Identifying the mechanism of intracellular parasitism by Cryptococcus

Cryptococcus is a free-living fungus that is able to cause a fatal infection in humans and other animals. Our current understanding of this disease suggests that fungal spores are inhaled into the lung and then ?hide? from the immune system, sometimes for several years, by living inside a particular type of host cell called a macrophage. Although this cell would kill most other pathogens, Cryptococcus has a remarkable ability to survive and indeed grow inside macrophages. In addition, we have recently discovered that the fungus is also able to ?escape? from inside of this host cell by a novel process called ?reverse phagocytosis?. This process is likely to be advantageous to the pathogen since it does not damage the host cell and is therefore unlikely to trigger a strong inflammatory response.

In order to develop better treatments for this lethal disease, we need to understand how the fungus can manipulate the host cell in this way. Our application is therefore directed at learning more about the molecular process that underlies this behaviour. In particular, we hope to answer two specific questions:
1) can we identify new ways in which human macrophages can be induced to destroy intracellular cryptococci and thereby eradicate latent infections?
2) how great is the risk that particular strains of Cryptococcus will become more dangerous to patients by evolving an improved ability to manipulate host cells in this way?

Cryptococcosis is a life-threatening disease of the central nervous system caused by the fungus Cryptococcus neoformans. The disease is uniformly fatal without rapid clinical intervention and even with the best available anti-fungal treatment, patient mortality exceeds 10%, rising to 88% in areas of the world with poor healthcare infrastructure. Cryptococcosis is primarily a disease of immunocompromised hosts (especially AIDS patients), although some strains of the closely related species C. gattii have the capacity to infect and kill immunocompetent individuals.

Pathogenic cryptococci show a remarkable ability to survive and proliferate within host macrophages following phagocytosis and are also capable of exiting the host cell via a unique, non-lytic, mechanism. Together these behaviours are thought to underlie the capacity of the pathogen to remain latent within the human host for long periods and, similarly, to cause relapsing episodes of cryptococcosis in patients after initially successful antifungal treatment. Importantly, this aspect of the disease means that infected patients are currently advised to remain on lifelong antifungal medication, despite the considerable cost and toxicity issues associated with this treatment regime. Thus an improved understanding of the cryptococcal-macrophage interaction may lead to significant improvements in our management of latent cryptococcosis.

We therefore propose to build on recent preliminary work from our group and use high-throughput timelapse microscopy to investigate how the normal cell biology of a macrophage is manipulated by Cryptococcus. In particular, we hypothesise that two factors may influence the outcome of this interaction. Firstly, it is known that changes in host inflammatory signalling (for example, in the levels of T-cell derived cytokines) alter cryptococcal disease progression. We therefore propose to investigate whether this may, in part, result from an effect on infected macrophages. Secondly, there is a wealth of data showing that variability in Cryptococcus genotype strongly influences virulence in mammals. We therefore propose to investigate what effect pathogen genotype and phenotype have on intracellular parasitism by this pathogen and thus better understand what risk novel cryptococcal strains may present to human health.

In summary, this proposal is intended to address an important aspect of the host-pathogen interaction in this fatal disease. By understanding the molecular basis of this interaction, we hope to identify possible targets for therapeutic intervention in the future and thereby improve long-term prognosis for infected patients.