The development of biological control agents (BCAs) is a promising and environmentally friendly method to control plant pathogens. In a grapevine nursery greenhouse and for the first time on grafted cuttings (scion cv. Cabernet sauvignon), this two-year study demonstrated the significant efficacy of the oomycete Pythium oligandrum as a BCA against two major aggressive fungal pathogens, Neofusicoccum parvum and Phaeomoniella chlamydospora, which are involved in grapevine trunk diseases (GTDs). By considering the reduction in necrosis lengths within the scion stem, treatments with P. oligandrum alone showed the greatest efficacy against the two pathogens (overall average efficacy of 48.3%). This major result was obtained during the two-year bioassay and in cuttings grafted on the two widely used rootstocks, 101-14 and SO4. The biocontrol efficacies of two bacterial strains previously isolated from vineyards, Pantoea agglomerans and Bacillus pumilus, were also assessed, separately or in combination with P. oligandrum. Treatments with each bacterial strain were less effective than treatments with P. oligandrum, and the efficacies were not improved when they were applied in combination with the oomycete.