Fusarium wilt of watermelon (Citrullus lanatus), caused by the soilborne fungus Fusarium oxysporum f. sp. niveum, is the most serious disease of watermelon in South Carolina and other southeastern U.S. states. Isolates of F. oxysporum collected from field-grown plants, greenhouse-grown seedlings, and field soil between 1999 and 2018 were inoculated onto three differential watermelon cultivars to identify races. Of 197 isolates obtained from plants, 12% were nonpathogenic, 2% were race 0, 23% were race 1, and 63% were race 2. One collection of isolates from greenhouse seedlings was exclusively race 1 and the other was exclusively race 2. Seventeen of 81 soil isolates were pathogenic: five were race 1 and 12 were race 2. Reactions of C. amarus PI 296341-FR, Carolina Strongback, and SP-6, cultigens with resistance to race 2, did not differ significantly among five highly virulent race 2 isolates and a standard race 2 isolate, indicating a lack of a race 3 phenotype. Forma specialis-specific primers matched phenotypic race identification for 74% of the isolates. Race-specific primers based on a secreted-in-xylem elicitor present in race 0 and 1 isolates matched phenotypic race identification for 66% of the isolates. Because a majority of the F. oxysporum f. sp. niveum isolates from South Carolina were race 2, integrated management practices should be used until commercial cultivars with resistance to race 2 are available.