Metabolic adaptation and nutrient acquisition of Paracoccidioides spp. during host interactions: Search for targets and new therapeutic approaches

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The number of life-threatening fungal infections is steadily increasing. Due to limited diagnostic and therapeutic opportunities invasive fungal infections may cause lethality rates of more than 50%. Paracoccidioides brasiliensis is an ubiquitous fungus in Latin America. This fungus has a dimorphic life-style with a filamentous growth morphology at lower and a yeast morphology at elevated temperatures. In soil the filamentous form is the prevalent morphology and asexual conidia are formed from hyphae. These conidia are easily inhaled by farm workers. After inhalation condia may germinate and form persisting yeast cells in the lung.It has been assumed that in Latin America -and especially in Brazil- approximately 10 million people carry a latent infection with Paracoccidioides spp. and in 1 - 2% of cases the fungus is no longer controlled by the immune system, which leads to acute infections that, when untreated, are life-threatening.In order to persist and proliferate within host tissues the acquisition and utilisation of host-provided nutrients is indispensable for all pathogenic organisms. In terms of P. brasiliensis, previous studies have shown that regulation of metabolism and metabolic physiology among hyphae and yeasts substantially differs. Even more, at least four virulent P. brasiliensis subspecies (S1, PS2, PS3 and 01-like) have been described and first preliminary analyses indicate that these subspecies differ in their metabolic preferences when entering the yeast form. However, the impact on virulence, fungal proliferation and therapeutic options has not been studied yet.Therefore, this projects deals on the identification and characterisation of the metabolic physiology of the different P. brasiliensis subspecies to gain a deeper understanding on the impact and interplay of metabolic pathways during the infection process. Identification of essential metabolic processes provides new targets for antifungal therapies that may not only be suitable for combating P. brasiliensis, but also other invasive life threatening fungal infections.By an generating bioluminescence imaging system for Paracoccidioides species, it will be possible to establish an in vivo imaging approach that allows the real-time visualisation of disease progression in temporal and spatial resolution using murine model systems of Paracoccidiomycosis. This system will be of high value to investigate the niche-specific metabolic capacity of Paracoccidioides spp. that is required proliferate within the host. To assess the impact of distinct metabolic pathways key enzymes will be silenced by gene knock down approaches, which will be performed in combination with bioluminescence reporter expression. The resulting mutants will be tested in alternative and murine infection models to investigate the fungal proliferation rate and dissemination.To identify new compounds targeting key metabolic pathways, a library of natural products available at the Brazilian partners will be screened. This includes the direct inhibition of purified enzymes as well as inhibition analyses of fungal growth under in vitro conditions. Interesting candidates will subsequently become analysed for potential cytotoxic effects on cell lines and eventually in murine models.By performing these complex analyses the project not only aims in broadening the understanding of metabolic adaptations during the infection process, but also tries to refine the understanding of Paracoccidioides spp infection process and will identify new compounds that target essential metabolic pathways and could be suitable to prevent and cure invasive fungal infections.