Enhanced ability of freshwater bacteria to secrete extracellular vesicles upon interaction with virus

The ability of freshwater bacteria to secrete extracellular vesicles (EVs) upon interaction with viruses remains to be established. Here, we investigated for the first time if freshwater virus-infected bacteria release EVs in both natural ecosystems and virus-like particles (VLPs)-enriched cultures. We performed a systematic study using transmission electron microscopy to visualize viruses and EVs at high resolution and single-cell imaging analyses to quantitate nascent EVs at the surface of gram-negative bacteria. First, by analysing freshwater samples from a tropical ecosystem (Negro River/Amazon Basin/Brazil), we captured bacteriophages-infected bacteria releasing EVs from their outer membrane. Next, VLPs isolated from these samples and inoculated in bacterial cultures not only impacted bacteria growth and viability but also led them to a significant release of EVs (~300% increase in numbers/cell section) compared to controls. The numbers of both budding and free EVs and EVs per linear micrometre of cell envelope were significantly higher in infected bacteria. Our findings identify a yet-not recognized capability of freshwater bacteria in generating EVs (overvesiculation) in response to viral infection. Since viruses are abundant members of aquatic ecosystems and bacteria are natural hosts for them, such interaction is an interesting event for microbial communities to be explored in freshwater ecosystems.