Project description:Powdery mildew (Erysiphe necator) is a widespread and economically important disease of grapevines. Large quantities of fungicides are used for its control, accelerating the incidence of fungicide-resistance. A shotgun approach was applied to sequence and assemble the E. necator genome of five isolates, and RNA-seq and comparative genomics were used to predict and annotate protein-coding genes. In addition, a collection of 98 E. necator isolates collected from diverse locations was used for SSR profiling and was screened for copy number variation and presence/absence of a single point mutation (Y136F) in the CYP51 gene, a key target for DMI fungicides. Our results show that the E. necator genome is exceptionally large and repetitive and suggests that transposable elements are responsible for genome expansion. Frequent structural variations were found between isolates and included copy number variant (CNV) in CYP51. We show that CYP51 copy number correlates with expression level and that an increase in copy number is detected in isolates collected from fungicide-treated vineyards. This copy number variation was usually detected with the CYP51 mutant allele (Y136F), suggesting that an increase in copy number becomes advantageous only when the allele is mutated. We also show that CYP51 copy number correlates with fungal growth in the presence of DMI fungicide in vitro. These results suggest that copy number variation can be adaptive in the development of resistance to DMI fungicides in E. necator. Detach Carignan leaves were sprayed with a Erysiphe necator C-strain conida suspension, leaves were collected in 4 time points (12 hours, 24 hours, 3 days, and 6 days) post inoculation. For each time point, two leaves from the same seedling were pooled and frozen in liquid nitrogen, and each time point was performed in triplicate.
