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Downy mildew (Plasmopara viticola) is the primary disease in viticulture worldwide, and your control requires synthetic fungicides applications to avoid quality and yield loss in the grapevines. However, alternatives to reduce synthetic fungicides are needed to ensure the consumer’s health and the environment. Essential oils (EOs) are amongst the most promising natural plant protection alternatives because of their antifungal properties on several crop diseases. The present study objective was to determine the effect of Eucalyptus staigeriana EO in vitro and in vivo against P. viticola. The EO exhibited the highest activity in vitro, inhibiting 90% of the incidence and severity of disease caused by P. viticola in leaves of grapevines in the greenhouse. In the field (in vivo), treatment with EO could not control the disease; however, treatment with EO in consortium with conventional treatment reduced approximately 50% of the incidence and more than 90% of the severity of downy mildew disease in leaves, decreasing the application of synthetic fungicides by 50%.