Isolating genetic markers is often costly and time-consuming for non-model fungal species. However, these markers are of primary importance to identify the origin of invasive species and to infer their reproductive mode and dispersal ability. We slightly modified a recent molecular method to quickly isolate and validate single-nucleotide polymorphism (SNP) markers, from a first Erysiphe alphitoides draft genome, one of the main causal agent of oak powdery mildew in Europe. Although the draft assembly was strongly fragmented (555,289 contigs), we successfully isolated 1700 SNPs from 75 single-copy genes conserved in most fungal genomes. Ninety percent of them allowed to clearly distinguish the two main Erysiphe species reported on European oaks: E. alphitoides and E. quercicola. Thirty-six SNPs, located in distinct genes, were then validated using a strategy of MassArray genotyping on 95 E. alphitoides isolates sampled in Europe. This genotyping showed that only monospore isolates had the expected haploid signature, whereas direct genotyping from field leaves showed signature of mixed infection. Considering haploid isolates, these markers led to the first results of population genetic diversity, and suggested that E. quercicola may have a more asexual reproduction than its sister species, E. alphitoides.