Understanding pathogen evolution over time is vital for plant breeding and deployment of host resistance. In the context of a soilborne pathogen, the potential of host-directed evolution of a Verticillium dahliae race 1 isolate and genotypic variation of V. dahliae associated with two major hosts (lettuce and tomato) were determined. In total, 427 isolates were recovered over 6 years from a resistance screening nursery infested with a single V. dahliae race 1 isolate. In a separate study, an additional 206 isolates representing 163 and 43 isolates from commercial lettuce and tomato fields, respectively, were collected. Analyses of isolates recovered from the screening nursery over 6 years revealed no changes in the race and mating type composition but did uncover seven simple sequence repeat (SSR) variant genotypes. No significant genotypic variation in V. dahliae was observed between or within fields of either lettuce or tomato but pathogen populations were significantly differentiated between these two hosts. Replicated virulence assays of variant SSR genotypes on lettuce differential cultivars suggested no significant difference in virulence from the wild-type race 1 isolate introduced into the field. This suggests that deployed race 1 host resistance will be robust against the widespread race 1 populations in lettuce-growing regions at least for 6 years unless novel pathogen genotypes or races are introduced into the system.