Fungal extracellular vesicles mediate conserved cross-species communication and immunomodulation

Extracellular vesicles (EVs) play crucial roles in fungal communication and host immune modulation, representing potential therapeutic targets for fungal infections. This study investigated the role of fungal EVs in both intra- and interspecies communication, focusing on their effects on virulence and immune responses. Co-incubation experiments were performed using EVs derived from <i>Candida albicans</i> and <i>Candidozyma auris</i> to assess interactions with <i>C. albicans</i> planktonic cells and biofilms, as well as <i>Cryptococcus neoformans</i> and <i>Cryptococcus gattii</i> EVs interacting with <i>C. neoformans</i> cultures. EVs were observed associating with recipient cell surfaces, suggesting subsequent internalization. Functional assays revealed that EV exposure led to increased expression of <i>CAP59, LAC1, URE1,</i> and <i>ERG11</i> genes, correlating with reduced antifungal susceptibility in both planktonic and biofilm forms. Additionally, EVs facilitated cross-species communication, enhancing biofilm adhesion and dispersion, which emphasizes their role in phenotypic modulation. Macrophages stimulated with fungal EVs exhibited receptor-specific gene expression changes (dependent on the EVs' origin, including variation among species of the same genus), along with a pro-inflammatory phenotype marked by increased <i>iNOS</i> expression, enhanced TBK1/STAT1 production, and elevated levels of IL-1β, IL-6, and IL-8. Collectively, these findings underscore a critical role for fungal EVs in interspecies communication, biofilm regulation, and immune modulation, offering valuable insights into fungal pathogenicity mechanisms.IMPORTANCECurrently, no vaccines exist to prevent fungal infections, underscoring the need for new therapies. As fungal diseases increase globally, understanding fungal biology is essential to identifying treatment targets. Fungi use extracellular vesicles (EVs) to communicate and evade immune responses. EVs mediate cell-cell communication, transporting proteins, polysaccharides, lipids, and nucleic acids, serving as "messages" exchanged within a fungal network. Understanding how these vesicles facilitate communication not only within a single species but also across different fungal species can shed light on their contribution to infection persistence and cross-species adaptability. Moreover, EVs may have a broader role in inter-kingdom signaling, influencing how fungi interact with host immune cells. The impact of fungal EVs on human innate immune responses remains a largely underexplored area, with significant gaps in our knowledge. This study aims to examine how fungal EVs affect immune responses and whether their signaling varies across species, potentially revealing new therapeutic targets.