Comparative volatilomics identifies ubiquitous sulfur compounds inhibiting the fungal pathogen Rasamsonia argillacea

Members of the phylum Actinomycetota, particularly Streptomyces species, are prolific producers of bioactive metabolites, and bioprospecting in unique environments may uncover novel species producing previously undescribed antifungal compounds. In previous work, Streptomyces strains isolated from cave moonmilk deposits completely inhibited the growth of Rasamsonia argillacea, an emerging fungal pathogen associated with chronic granulomatous disease (CGD) and cystic fibrosis (CF). Cross-streak and bipartite Petri dish assays revealed that R. argillacea inhibition occurred specifically when bacteria were cultivated on Mueller-Hinton agar (MHA) and was mediated by volatile compounds rather than diffusible metabolites. This antifungal effect was not strain specific, as it was reproduced by phylogenetically diverse bacteria grown on MHA, suggesting the involvement of ubiquitous volatile molecules. Similar inhibitory effects were observed on MHA against other fungi and yeasts relevant to CGD and CF, supporting the hypothesis that these volatiles are broadly toxic rather than species-specific. To identify candidate ubiquitous antifungal VCs, we conducted comparative volatilomics of two phylogenetically distant bacterial strains that consistently inhibited fungal growth. Among the 143 VCs detected, only dimethyl trisulfide (DMTS) and dimethyl disulfide (DMDS) showed consistent presence in both bacterial volatilomes, with significantly increased production under conditions that promoted fungal inhibition. Exposure assays with pure compounds confirmed that both DMDS and DMTS strongly inhibited R. argillacea growth, with DMTS exhibiting greater potency. Our findings position these ubiquitous sulfur compounds as valuable models for exploring novel agents against human pathogens.

Our findings suggest that antifungal activity against human pathogens may arise from common metabolic pathways shared across diverse microbes rather than from unique biosynthetic systems. Because dimethyl disulfide and dimethyl trisulfide can also be generated through microbial and dietary sulfur metabolism, it is plausible that the human microbiome may produce similar volatiles depending on diet composition, particularly following consumption of sulfur-rich foods. This study, therefore, underscores the critical influence of culture conditions on revealing bioactive volatile production and opens intriguing perspectives on the ecological and physiological roles of ubiquitous microbial metabolites in regulating fungal colonization and microbiome-host interactions.