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Esca disease, which attacks the wood of grapevine, has become increasingly devastating during the past three decades and represents today a major concern in all wine-producing countries. This disease is attributed to a group of systematically diverse fungi that are considered to be latent pathogens, however, this has not been conclusively established. This study presents the first in-depth comparison between the mycota of healthy and diseased plants taken from the same vineyard to determine which fungi become invasive when foliar symptoms of esca appear. An unprecedented high fungal diversity, 158 species, is here reported exclusively from grapevine wood in a single Swiss vineyard plot. An identical mycota inhabits wood of healthy and diseased adult plants and presumed esca pathogens were widespread and occurred in similar frequencies in both plant types. Pioneer esca-associated fungi are …