The genus Vitis is highly diverse and includes more than 60 species. In scion breeding, hybrid grapevines are created to combine fruit quality of V. vinifera cultivars with a wide range of adaptive traits from their wild relatives. Among these, resistance to disease is the most representative example. In this work, the goals were to dissect genetic regions that control disease resistance and pest-related traits in V. rupestris B38, V. cinerea B9, V. vinifera 'Chardonnay', Illinois 547-1 and the complex hybrid 'Horizon'. For this, we studied the segregation of powdery mildew (Erysiphe necator) and phomopsis (Diaporthe ampelina) disease intensities; predatory mite (Phytoseiid) abundance; and trichome trait densities, such as domatia, leaf hairs and bristles, in bi-parental F1 families. We used genotyping-by-sequencing (GBS) to rapidly identify single nucleotide polymorphisms across the whole genome. In order to make full use of this technology in heterozygous crosses, we developed novel methods to generate dense genetic maps and validated them by correctly localizing the sex locus. Multiple QTL mapping was used to identify genetic regions controlling resistance traits. For powdery mildew, we identified a moderate effect locus controlling susceptibility on chromosome 9 of the cultivar, 'Chardonnay'. Two qualitative loci, Rda2 and Rda1, were found to be responsible for phomopsis cane and berry rot resistance on chromosome 7 of 'Horizon' and chromosome 15 of V. cinerea B9, respectively. Further, transcriptome analyses in V. cinerea B9 characterized a resistance reprograming and expression QTL mapping (eQTL) narrowed down Rda1 candidate genes to three NBS-LRR genes. Genetic determinism of leaf trichomes was more complex, with several major and minor QTL controlling trait expression on leaf blades and veins. Among these, a QTL located on chromosome 1 of Illinois 547-1 had major effect on all trichome traits and was also correlated with the abundance of predatory mites. While providing knowledge of the mechanisms of powdery mildew resistance, phomopsis cane resistance and abundance of predatory mites, this work also delivered tools for the creation of dense genetic maps in heterozygous crosses and for marker assisted selection in grapevine breeding.