Historically, wilt disease caused by Verticillium dahliae has not presented a problem in California spinach production because the crop is harvested well before the symptoms develop after the stem elongation (bolting) stage. However, infested spinach seeds introduce or increase inoculum in the soil for rotational crops such as lettuce. This investigation was designed to identify verticillium wilt-resistant accessions in the U.S. Department of Agriculture (USDA) spinach germplasm collection against races 1 and 2 of V. dahliae, and to examine seed transmission of the pathogen in different spinach genotypes. In a seed health assay of 392 accessions, 21(5.4%) were positive for V. dahliae, and 153 (39%) were positive for Verticillium isaacii. A total of 268 accessions plus nine commercial cultivars were then screened against one race 1 and two race 2 isolates from spinach in replicated greenhouse experiments. Disease incidence, severity, and seed transmission through plating on NP-10 medium and real-time quantitative polymerase chain reaction (qPCR) were assessed. There was wide variation among accessions in their response to V. dahliae with disease incidence ranging from 0% to 100%. The two race 2 isolates differed in their virulence against spinach genotypes. Resistant accessions were identified against both races 1 and 2. Recovery of V. dahliae from seeds plated on NP-10 medium and qPCR results were highly correlated (P = 0.00014). Some accessions identified as resistant based on disease incidence showed little seed transmission of the pathogen. Even though lower wilt incidence and severity generally corresponded with lower seed transmission rates, there were exceptions (r = 0.52). Variation among plants within accessions was also observed. Nevertheless, the sources of resistance identified in this study are useful for spinach cultivar improvement.