Perennial plant decline is increasingly threatening the profitability and sustainability of agriculture and forestry worldwide. It results from intricate interactions between microbial communities, the plant host, and abiotic stressors. We investigated the effects of drought and esca disease on mature grapevine phytobiomes. Grapevines display no esca leaf symptoms during droughts, but the impacts of drought and esca expression on fungal communities and wood health in mature plants remain poorly understood. We studied 43 uprooted 30-year-old naturally infected vines in three experimental conditions: well-watered asymptomatic (Control) vines, vines with esca symptoms (Esca), and vines subjected to water deficit (WD) over two consecutive summers. We profiled trunk, cane, stem and petiole Ascomycota communities by DNA metabarcoding with primers specifically designed for grapevine trunk-associated Ascomycota, and quantified wood necrosis. The Ascomycota communities of trunks and younger organs clearly differed, and drought and esca had different impacts on the Ascomycota communities of perennial and young organs. In the trunk, drought significantly decreased fungal diversity in healthy wood and increased the abundance of wood pathogens (e.g. Phaeomoniella chlamydospora, Botryosphaeria dothidea). In young organs, esca expression decreased the species richness and diversity of the Ascomycota community to a greater extent than drought. We also found that the relative proportion of healthy wood was smaller in plants with esca symptoms than in control plants. Thus, drought increased Ascomycota pathogen abundance in the trunk but did not increase wood degradation and esca expression, highlighting the need to investigate the molecular basis of plant-microbiome interactions under multi-stress conditions.