A common vesicle proteome drives fungal biofilm development.

Extracellular vesicles mediate community interactions among cells ranging from unicellular microbes to complex vertebrates. Extracellular vesicles of the fungal pathogen are vital for biofilm communities to produce matrix, which confers environmental protection and modulates community dispersion. Infections are increasingly due to diverse species, such as the emerging pathogen , as well as mixed communities. Here, we define the composition and function of biofilm-associated vesicles among five species across the genus. We find similarities in vesicle size and release over the biofilm lifespan. Whereas overall cargo proteomes differ dramatically among species, a group of 36 common proteins is enriched for orthologs of biofilm mediators. To understand the function of this set of proteins, we asked whether mutants in select components were important for key biofilm processes, including drug tolerance and dispersion. We found that the majority of these cargo components impact one or both biofilm processes across all five species. Exogenous delivery of wild-type vesicle cargo returned mutant phenotypes toward wild type. To assess the impact of vesicle cargo on interspecies interactions, we performed cross-species vesicle addition and observed functional complementation for both biofilm phenotypes. We explored the biologic relevance of this cross-species biofilm interaction in mixed species and mutant studies examining the drug-resistance phenotype. We found a majority of biofilm interactions among species restored the community's wild-type behavior. Our studies indicate that vesicles influence the development of protective monomicrobial and mixed microbial biofilm communities.