Extracellular vesicles regulate yeast growth, biofilm formation, and yeast-to-hypha differentiation inCandida albicans

Abstract

The ability to undergo morphological changes during adaptation to distinct environments is exploited by Candida albicans and has a direct impact on virulence. In this study, we investigated the influence of fungal extracellular vesicles (EVs) during yeast growth, biofilm formation, and morphogenesis in C. albicans. Addition of C. albicans EVs (Ca EVs) to the culture medium positively affected yeast growth. Using crystal violet staining and scanning electron microscopy (SEM), we demonstrated that Ca EVs inhibited biofilm formation by C. albicans in vitro. By time-lapse microscopy and SEM, we showed that Ca EV-treatment stops filamentation promoting pseudohyphae formation with multiple sites for yeast budding. The ability of Ca EVs to regulate dimorphism was further compared to EVs isolated from different C. albicans strains, Saccharomyces cerevisiae, and Histoplasma capsulatum. Ca EVs from distinct strains robustly inhibited yeast-to-hyphae differentiation with morphological changes occurring in less than 4 hours. A minor inhibitory effect was promoted by EVs from S. cerevisiae and H. capsulatum only after 24 hours of incubation. The inhibitory effect of Ca EVs was promoted by a combination of lipid compounds identified by gas chromatography-tandem mass spectrometry analysis as sesquiterpenes, diterpenes, and fatty acids. Remarkably, Ca EVs were also able to reverse filamentation, transforming hyphal growth to yeast forms. Transcriptomic analysis demonstrated that treatment with Ca EVs modified the expression of more than 300 genes. The most effectively upregulated pathways were related to DNA metabolism. The downregulated genes were mostly associated with extracellular and adhesion proteins. Finally, yeast cells treated with Ca EVs for 24 hours lost their agar invasive ability and were avirulent when inoculated in Galleria mellonella larvae. In summary, our results indicate that fungal EVs can profoundly modify C. albicans growth and regulate yeast-to-hypha differentiation inhibiting biofilm formation and virulence.

Fungal extracellular vesicles are involved in intraspecies intracellular communication

“…albicans EVs on invasive hyphae development and virulence, as shown in Galleria mellonella model [31]. Moreover, that study was conducted exploring EVs obtained from yeast cultures, and it is reasonable that the cargo and the message within this EV contribute to cells remain in this form.…”

“…Many QS molecules might also be found within fungal EVs; however, it is also possible to speculate a cross-regulation between small RNA content in EVs and QS phenomena in fungi. A recent study showed the involvement of QS molecules such as farnesol derivatives and medium-chain fatty acids in EVs obtained from yeast culture as potential regulators of cellular proliferation and yeast-to-hyphae development [31]. The EVs from C. albicans reduced the hyphae and biofilm formation, and the impact of C. albicans EVs on invasive hyphae development and virulence, as shown in Galleria mellonella model [31].…”

“…A recent study showed the involvement of QS molecules such as farnesol derivatives and medium-chain fatty acids in EVs obtained from yeast culture as potential regulators of cellular proliferation and yeast-to-hyphae development [31]. The EVs from C. albicans reduced the hyphae and biofilm formation, and the impact of C. albicans EVs on invasive hyphae development and virulence, as shown in Galleria mellonella model [31]. Moreover, that study was conducted exploring EVs obtained from yeast cultures, and it is reasonable that the cargo and the message within this EV contribute to cells remain in this form.…”

“…We demonstrated that the conditions that underwent EVs uptake presented an increase in mitochondrial activity, suggesting a potential function of EVs in heightening cellular metabolism and/or proliferation. EVs were previously shown to enhance cellular proliferation in C. albicans [31] as well as in C. neoformans within macrophages. The high activation of the C. neoformans proliferation rate was named the “division of labor” mechanism [11].…”

“…Another report showed that the supernatant from a highly virulent strain of C. neoformans culture stimulated the pathogenic potential of a less virulent isolate, an effect also attributed to EVs [30]. Recently, a study advocated the role of EVs from C. albicans as messaging compartments involved in growth, morphogenesis, and biofilm production [31]. These studies postulated a role for fungal EVs in intra-species communication; however, at the molecular level, many aspects related to the mechanisms switched on by EVs remain to be investigated.…”