Neurospora crassa Δccg8 compromises cell surface integrity and antifungal tolerance: Insights from in vitro and Galleria mellonella studies

When studying filamentous fungi in terms of adaptation to environmental conditions, a reference organism is generally missing. This is mainly obstructing in the research of pathogens (well defined model organisms are non-pathogenic). Here, we investigated the mechanism underlying the ability of N. crassa to respond to various environmental stimuli, including the presence of azole and echinocandin antifungals. Our molecular analysis revealed that the Δccg8 deletion impacts the fungus on a broad physiological level, including dysregulation of ribosome biogenesis and fatty acid β-oxidation (as demonstrated by proteomic profiling), and supporting changes in glycosylated proteins and fatty acid and ergosterol composition. To bridge the gap between in vitro and in vivo conditions, we successfully applied Galleria mellonella larval model. Despite being non-pathogenic, when injected with conidia of tested filamentous fungi, the immune system cleared conidia of Δccg8 mutant strain more effectively than wild-type strain, mirroring in vitro observations and demonstrating a reproducible immune reaction. This work not only provides molecular insights into the fungal stress adaptation but also establishes N. crassa as a viable non-pathogenic organism for in vivo analysis, substantially broadening its application, reaching into dominion of research of pathogenic filamentous fungi.