The Cryptococcus gattii species complex comprises some of the aetiological agents of cryptococcosis, a severe fungal disease that affects a wide variety of hosts and is acquired from the environment by inhalation. Koalas (Phascolarctos cinereus) appear to be particularly susceptible to cryptococcosis. In Australia, eucalypt tree hollows are the classic ecological niche for C. gattii molecular type VGI and therefore are also a potential source of infection. Aspects of the tree hollow microenvironment that may allow for the growth and dispersal of C. gattii VGI remain poorly understood. The C. gattii species complex has been associated with outbreaks and case clusters, and animals are often considered useful sentinels for the disease in these scenarios. The prevalence of cryptococcosis in Australian wildlife remains unknown. Given the koala’s propensity towards developing cryptococcosis, and its regular contact with a common ecological niche for the C. gattii species complex (eucalypts), it is an ideal sentinel species. The host-pathogen-environment interactions of cryptococcosis caused by the C. gattii species complex, particularly progression from exposure to colonisation of the respiratory mucosa to eventual tissue invasion, remain poorly understood. This thesis uses amplicon-based next generation sequencing to characterise the fungal microbiome of Australian tree hollows, focusing on the role that the C. gattii species complex may play in this microenvironment. The prevalence of cryptococcosis in a population of free-ranging koalas is systematically characterised, while the pathogenesis, treatment and diagnosis of the disease in this host species are also explored. Finally, fine-scale molecular epidemiology tools (multi-locus sequence typing and whole genome sequencing) are used to determine sources of infection and examine disease caused by the C. gattii species complex in Australia, using primarily the koala as a model for naturally-occurring cryptococcosis.