Grapevine trunk diseases (GTDs) are a major global threat to viticulture, yet data on their etiology and epidemiology remain scarce in Serbia and Southeast Europe. This study provides the first comprehensive analysis of Botryosphaeriaceae species associated with GTDs in Serbia, combining morphological characterization, temperature-growth assays, multilocus phylogenetic analyses (internal transcribed spacer, beta-tubulin, translation elongation factor-1α) and pathogenicity testing. A total of 46