Fusarium oxysporum, a cross-kingdom pathogen, causes notorious diseases across diverse hosts, including plants, animals, and humans. Within the F. oxysporum species complex (FOSC), formae speciales were defined based on their host specificity, such as F. oxysporum f. sp. vasinfectum (Fov) causing Fusarium wilt in cotton. However, the genetic diversity and evolution of isolates within a formae speciales remain elusive. We sequenced and assembled high-quality genomes of two classical Fov races and two recently evolved isolates in a naturally infected field in Texas, USA. Comparative genomic analysis revealed that Fov isolates share 11 core chromosomes with other FOSC isolates that infect plants and humans, along with a set of distinctive accessory chromosomes. Notably, while core chromosomes exhibited high conservation, accessory chromosomes displayed significant divergence within different Fov races, comparable to the divergence observed among different formae speciales. Phylogenetic analysis supported the notion of independent evolution among various Fov races within FOSC. Extensive microsynteny was identified among accessory chromosomes within the same or different FOSC isolates, suggesting the existence of a shared ancestral accessory chromosome in FOSC. Furthermore, genome sequencing and comparative analysis of additional 27 Texas Fov field isolates with varying virulence unveiled rapid structure and sequence changes in accessory chromosomes within a Fov race, and the association of pathogenicity with copy numbers of putative effector genes in accessory chromosomes. Collectively, our findings indicate that isolates within the same formae speciales of F. oxysporum can evolve independently, and the accessory chromosomes in FOSC may share a common ancestor.