Answers to naysayers regarding microbial extracellular vesicles.

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.