Aspergillus fumigatus is an opportunistic fungal pathogen that secretes an array of immune-modulatory molecules, including secondary metabolites (SMs), which contribute to enhancing fungal fitness and growth within the mammalian host. Gliotoxin (GT) is a SM that interferes with the function and recruitment of innate immune cells, which are essential for eliminating A. fumigatus during invasive infections. We identified a C6 Zn cluster-type transcription factor (TF), subsequently named RglT, important for A. fumigatus oxidative stress resistance, GT biosynthesis and self-protection. RglT regulates the expression of several gli genes of the GT biosynthetic gene cluster, including the oxidoreductase-encoding gene gliT, by directly binding to their respective promoter regions. Subsequently, RglT was shown to be important for virulence in a chemotherapeutic murine model of invasive pulmonary aspergillosis (IPA). Homologues of RglT and GliT are present in eurotiomycete and sordariomycete fungi, including the non-GT-producing fungus A. nidulans, where a conservation of function was described. Phylogenetically informed model testing led to an evolutionary scenario in which the GliT-based resistance mechanism is ancestral and RglT-mediated regulation of GliT occurred subsequently. In conclusion, this work describes the function of a previously uncharacterised TF in oxidative stress resistance, GT biosynthesis and self-protection in both GT-producing and non-producing Aspergillus species. A. fumigatus is a prevalent opportunistic fungal pathogen that causes a wide range of diseases, in both immunocompetent and immunocompromised subjects. Contributing to A. fumigatus virulence, is the in vivo secretion of gliotoxin, a mycotoxin with both immunosuppressive and immunomodulatory properties. Regulation of the gliotoxin-encoding biosynthetic gene cluster has proven to be a multifactorial and hierarchical process. In this work, we identify a transcription factor, named RglT (regulator of gliotoxin), that is crucial for gliotoxin biosynthesis and self-protection, a process that mainly occurs through the direct regulation of the oxidoreductase-encoding gene gliT. The absence of rglT resulted in a strain that was highly sensitive to oxidative stress, to exogenous gliotoxin and hypovirulent in a chemotherapeutic murine model of IPA. Furthermore, we show that homologues of both RglT and GliT co-exist within the class of eurotiomycetes and appear to be functionally conserved in non gliotoxin-producing fungal species. Our study therefore highlights the importance of mycotoxin self-protection in pathogenic and non-pathogenic fungal species with consequences for fungal virulence and survival.

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous group of immune cells that play a central role in regulating host immune responses during fungal infections. Their recruitment is mediated by pathogen recognition receptors, particularly Dectin-1 and CARD9 signaling, which promote the production of reactive oxygen species (ROS) and IL-1β. Once activated, MDSCs suppress T-cell and natural killer cell functions through immunosuppressive cytokines like IL-10 and TGF-β, as well as enzymes such as arginase-1 and indoleamine 2,3-dioxygenase 1 (IDO-1). This review explores the role of MDSCs in fungal infections caused by Candida spp., Paracoccidioides brasiliensis, Aspergillus spp., and Cryptococcus neoformans, emphasizing their impact on immune modulation and disease progression. The emerging evidence suggests that fungal bioactive compounds, such as polysaccharides, can influence MDSC activity and restore immune balance. Notably, therapies targeting MDSCs have demonstrated promise in both fungal infections. In particular, infections with P. brasiliensis and C. neoformans show improved T-cell responses following MDSC-targeted interventions. Additionally, polysaccharides from Grifola frondosa and exposure to Aspergillus sydowii affect MDSC behavior, supporting the potential of modulating these cells therapeutically. Together, these findings highlight the relevance of MDSCs in fungal pathogenesis and underscore their potential as targets for immunotherapeutic strategies in infectious diseases.

Cryptococcal infection commonly begins as an opportunistic infection in humans, however, this can escalate to a systemic or life-threatening form in immunocompromised individuals. Here, we aim to identify novel antifungal molecules from plants resources. Sclareolide, a phytochemical classified as a sesquiterpene lactone, was assessed against Cryptococcus neoformans H99. Sclareolide exhibited promising antifungal properties with a minimum inhibitory concentration (MIC) of 16 µg/mL. Additionally, the C. neoformans growth rate was significantly affected by sclareolide treatment in a concentration-dependent manner, as observed through a time killing assay, with a significant reduction at MIC × 8 compared to the control by 48 h. To elucidate the underlying mechanisms of sclareolide antifungal activity, fluorescence-based methods were employed. Propidium iodide (PI) accumulation assay indicated a reduction in C. neoformans membrane integrity, with values as low as 6.62 ± 0.18% after treatment. Moreover, sclareolide at MIC × 4 and MIC × 8 significantly increased the production of reactive oxygen species (ROS) and reduced the mitochondrial membrane potential (MMP), suggesting oxidative stress and mitochondrial dysfunction in C. neoformans. Sclareolide did not induce caspase-dependent apoptosis, suggesting a non-apoptotic mechanism. Further, a checkerboard experiment was performed to assess potential synergistic interaction with Amphotericin B, however, no synergism was observed. Moving on, sclareolide at 128 µg/mL did not exhibit toxicity in Galleria mellonella, further supporting its potential as a safe antifungal agent. These findings suggest that the antifungal activity of sclareolide against C. neoformans is mediated by oxidative stress. Further in vivo and pharmacokinetic studies are recommended to explore the potential of sclareolide as a prototype for the development of novel anti-cryptococcal therapies.

Paracoccidioidomycosis (PCM) is regulated by suppressive mechanisms mediated by plasmacytoid-dendritic cells, regulatory T cells and myeloid-derived suppressor cells (MDSCs). MDSC suppressive activity on Th1/Th17 immunity was shown to be mediated by inhibitory effect of IL-10, IDO-1, and PD-L1. Studies revealed the 5-fluorouracil (5-FU) as a selective MDSC apoptosis-inducing agent, but its in vivo effect on infectious processes remains poorly investigated. MDSCs and other leukocytes were evaluated in the lungs of 5-FU–treated mice after 4, 6, and 8 weeks of Paracoccidioides brasiliensis infection. Disease severity and immunological response were evaluated in MDSCs-depleted mice. 5-FU treatment caused a reduction of pulmonary MDSCs and fungal loads. The specific depletion of MDSCs reduced all pulmonary CD4⁺ T-cell populations resulting in improved tissue pathology and increased survival. This reduction was concomitant with increased frequencies of Th1/Th17 cells and the increased levels of Th1/Th2/Th17 cytokines in the lungs and liver of treated mice, suggesting an early and efficient protective effect of these cells. Furthermore, the immune protection conferred by the 5-FU treatment could be reversed by the MDSC-adoptive transfer. 5-FU depletes MDSCs of P. brasiliensis-infected mice, resulting in enhanced immunity. This protective effect can be viewed as a potential immunotherapeutic tool for PCM.