usarium wilt disease is a major biotic factor limiting watermelon production in the derived savanna agroecology of southeast Nigeria. The study hypothesized that variety could interact with the fungicide spraying regime to improve the agronomic performance of watermelon (Citrulus lanatus (Thunb) Matsum and Nakai). The objectives of the study were: to assess the effect of fungicide spraying regimes on disease incidence, severity, and agronomic performance of three watermelon varieties; and to identify and characterize the fungal causal organism of Fusarium wilt of watermelon in the derived savanna agroecology of southeast Nigeria. Three varieties of watermelon (Jubaly, red diamond, and sweet sangaria) and four fungicide spraying regimes (No spray, weekly, bi-weekly, and tri-weekly spray) were evaluated for agronomic performance, and disease incidence and severity in a 3 × 4 factorial in a randomized complete block design replicated thrice. Data were collected on phenological, growth and yield indices, and disease incidence and severity. Analysis of variance was done using Genstat 16th edition and GraphPad Prism 9 was used to construct the graphs. Variety significantly affected agronomic traits as well as disease incidence and severity. The Jubaly variety performed better than other varieties in most of the growth and yield parameters measured despite recording higher disease incidence and severity scores. Weekly application of fungiforce recorded the lowest disease incidence (16.67, 16.67, and 50%) and severity (6.7, 6.7, and 10%) at 9, 10, and 11 weeks after planting, respectively. The interaction effect of the Jubaly variety and weekly spray was consistently higher in vine length, node number, fruit number, and fruit yield. Morphological isolation and identification of the fungal causal organism showed the presence of Fusarium species. Further molecular characterization by DNA extraction from the Fusarium isolate and sequencing using the Sanger sequencing method confirmed the presence of Fusarium equiseti. The basic local alignment search tool result showed 94–100% similarity with F. equiseti strain WZ-98 from the NCBI Gene bank. The organism was identified as F. equiseti based on the Phylogeny.