Answers to naysayers regarding microbial extracellular vesicles

Abstract

It is now over 30 years since the discovery of extracellular vesicles (EVs) in Gram-negative bacteria. However, for cell-walled microbes such as fungi, mycobacteria and Gram-positive bacteria it was thought that EV release would be impossible, since such structures were not believed to cross the thick cell wall. This notion was disproven 10 years ago with the discovery of EVs in fungi, mycobacteria, and gram-positive bacteria. Today, EVs have been described in practically every species tested, ranging from Fungi through Bacteria and Archaea, suggesting that EVs are a feature of every living cell. However, there continues to be skepticism in some quarters regarding EV release and their biological significance. In this review, we list doubts that have been verbalized to us and provide answers to counter them. In our opinion, there is no doubt as to existence and physiological function of EVs and we take this opportunity to highlight the most pressing topics in our understanding of the biological processes underlying these structures.

Keywords: EVs; OMV; exosomes; extracellular vesicles; nanovesicles; outer membrane vesicles.

Figure 1.. EVs from Gram + bacteria and fungi.

(A) Negative Staining electron micrograph of L. monocytogenes EVs after Optiprep gradient separation (as prepared in [30]). (B) Transmission electron micrograph of EVs from C. neoformans (as prepared in [58]). (C) Scanning electron micrograph of EVs from C. neoformans (as prepared in [58]). Scale bars 100 nm. Images courtesy of Carolina Coelho and Raghav Vij (A), and Julie M. Wolf (B and C).

Figure 2.. EVs serve as delivery devices in intraspecies and interspecies interactions.

(A) C. albicans deleted for ESCRT subunit HSE1 is defective in biofilm and extracellular matrix. (B) Biofilm and extracellular matrix formation were rescued when wild-type EVs were. Added to HSE1−/− strains. Scale bars 11 μm. (C) EVs from the plant Arabidopsis near the pathogen B. cinerea infection sites. Scale bars, 1 μm. (D) Isolated Tetraspanin8 –GFP-labeled Arabidopsis EVs were taken up by B. cinerea within 2 h of co-incubation. Scale bars 10 μm. (E) L. monocytogenes release EVs when infecting a mammalian host. Bacterial phospholipids were previously loaded with fluorescent dye Bodipy-C12 fatty acid which is incorporated and released in EVs. Images courtesy of Zarnowski et al. [11] (A and B), Cai et al. [47] (C and D), and Coelho et al. [30] (E).