Australia is a major contributor to the global production of natural pyrethrins, derived from pyrethrum (Tanacetum cinerariifolium). However, crown and root rot diseases, along with plant stunting and winter losses, pose significant challenges in some cultivation areas. Members of the Fusarium oxysporum species complex (FOSC) have been identified as primary pathogens causing crown and root rot in pyrethrum, although their genetic diversity and pathogenicity remain poorly understood. This study investigated F. oxysporum isolates collected from symptomatic and asymptomatic pyrethrum tissues in the field. Identification was carried out through morphological assessments of fungal cultures and multigene phylogenetic analyses of partial sequences of the cmdA (calmodulin), rpb2 (RNA polymerase II subunit), tef1-α (translation elongation factor 1-alpha) and tub2 (β-tubulin) genes. Results revealed substantial genetic variation among the isolates, and placed these isolates within the context of different published FOSC taxonomies. Glasshouse trials successfully replicated the crown and root rot symptoms observed under field conditions. These findings emphasize the need for effective management of soil-borne pathogens to mitigate yield losses in pyrethrum. Moreover, the results will facilitate the identification of highly aggressive isolates for screening plant resistanceand the development of more resilient pyrethrum varieties.