Development and characterisation of a zebrafish larval model to investigate mechanisms for pathophysiology of intracranial hypertension in cryptococcal meningitis

Cryptococcosis is a fungal infection caused by members of the genus Cryptococcus. Worldwide, the most prevalent pathogen of this genus is the encapsulated saprophyte species Cryptococcus neoformans. Cryptococcosis most commonly occurs as an opportunistic airborne lung infection which can disseminate to most organ systems. The central nervous system appears particularly susceptible to developing a pathology from the infection, with more than half of cryptococcosis patients diagnosed with cryptococcal meningitis despite infection in multiple organs. Cryptococcal meningitis (CM) is a meningoencephalitis (infection of the brain parenchyma and meninges) which globally accounts for 19% (13-24) of AIDS-related mortality (Rajansingham et al., 2022). In 2020, reports show annual incidence of 152 000 cases of cryptococcal meningitis, resulting in 112 000 cryptococcal-related deaths, almost half of which are in eastern and southern Africa (Rajansingham et al., 2022). 50-70% of CM cases present with a pathologically elevated intracranial pressure (intracranial hypertension) (Graybill et al., 2000; Jarvis et al., 2014; Kagimu et al., 2022;).
This thesis aims to improve our understanding of intracranial hypertension in CM to help identify potential targets for treatment, by developing and testing new models of intracranial hypertension in CM. Three different approaches were used – theoretical, in vitro rheology and in vivo in zebrafish. Zebrafish was chosen as the core experimental system in which to develop new models due to its physiology, tractability for live imaging and susceptibility to cryptococcosis. In zebrafish larvae, the dynamic nature of cranial vasculature compartments and the CSF during infection was examined using wide field and light sheet microscopy techniques. The physical properties of tissues and fluids when interacting with cryptococcal yeast cells was modelled with theoretical and in vitro rheological measurements. In vitro it was found that viscosity of fluids may increase in the presence of heat killed C. neoformans, but whether this change is pathologically significant requires further investigation. Using a model of cryptococcal infection in zebrafish larvae, a “pulsation” phenomenon was identified, consisting of vasodilation and constriction in the cranial vasculature with an impact on vessel wall permeability. The findings in this work, are reflective of the CM pathology as seen in human patients and suggest impaired CSF and blood flow homeostasis may contribute to intracranial hypertension in CM.