Extracellular vesicles (EV) are membrane particles released by cells into their environment and are considered to be key players in intercellular communication. EV are produced by all domains of life but limited knowledge about EV in plants is available, although their implication in plant defense has been suggested. We have characterized sunflower EV and tested whether they could interact with fungal cells. EV were isolated from extracellular fluids of seedlings and characterized by transmission electron microscopy and proteomic analysis. These nanovesicles appeared to be enriched in cell wall remodeling enzymes and defense proteins. Membrane-labeled EV were prepared and their uptake by the phytopathogenic fungus Sclerotinia sclerotiorum was verified. Functional tests further evaluated the ability of EV to affect fungal growth. Spores treated with plant EV showed growth inhibition, morphological changes, and cell death. Conclusive evidence on the existence of plant EV is presented and we demonstrate their ability to interact with and kill fungal cells. Our results introduce the concept of cell-to-cell communication through EV in plants.