Abstract: Trichoderma species are cosmopolitan soil fungi, with some strains having potential as biocontrol organisms, since they can induce systemic resistance (ISR) in plants against leaf-borne pathogens. The mechanisms of Trichoderma-mediated ISR have mainly been investigated in Arabidopsis, while few studies have attempted to characterize the ISR in the Trichoderma-tomato system. In this study, we tested the ISR-inducing ability of Trichoderma hamatum T382 against the necrotrophic fungal pathogen Botrytis cinerea in tomato. For this purpose, a lab-scale hydroponics set-up was first optimized, which allowed us to obtain a robust ISR phenotype, as indicated by a significant reduction of necrotic lesion diameters on the leaves. In a next step we tested several tomato mutants and transgenic lines to investigate the role of the major plant hormones in the ISR in our tripartite system. We focused on jasmonic acid (JA; mutants spr1 and spr2), salicylic acid (SA; transgenic line nahG) and ethylene (ET; ethylene insensitive mutant Nr). Our results indicate that the ISR triggered by T. hamatum T382 in tomato is JA biosynthesis-dependent, while a role for SA and ET signaling was not found.