Esca is one of the main grapevine trunk diseases affecting vineyards worldwide. Phaeoacremonium minimum and Phaeomoniella chlamydospora are thought to be two of the main causal agents of this disease. However, the molecular mechanisms underlying plant defense responses in the grapevine trunk against esca-associated pathogens are poorly understood. To provide a first glimpse on the trunk responses to P. minimum and P. chlamydospora, transcriptomic and metabolomic analyses were performed to compare and contrast host responses to these pathogens. Transcriptomic analysis revealed different gene expression reprogramming in the trunk in response to each fungus. Main significant differences were found among genes associated with Secondary Metabolism, Signaling and Hormone Signaling. An untargeted liquid chromatography–high resolution mass spectrometry metabolomic approach performed 3 weeks after inoculation was used and dereplication mainly highlighted flavonoids and stilbenes as plant defense metabolites in the infected trunk. Some metabolites were overproduced with both fungi, but specific responses were also observed. Particularly, a lipophilic flavonoid cluster was emphasized after P. minimum inoculation. The assessment of fungal infection 6 wpi showed a higher number of copies of P. minimum than P. chlamydospora. This dissimilarity in the level of colonization could be linked somehow to the metabolomic responses observed. Our results reveal both different gene expression reprogramming and metabolomic specific signatures depending on the wood pathogen. Altogether, these observations suggest that grapevine trunk can differently perceive and respond to P. minimum and P. chlamydospora.