Opportunistic fungal pathogens, responsible for over 300 million severe cases and 1.5 million deaths annually, pose a serious global health threat, especially in immunocompromised individuals. Among these, Candida albicans is a major cause of both superficial and invasive infections, which can progress to systemic candidiasis. One of the critical factors in C. albicans pathogenicity is the yeast-to-hyphal transition, which enables biofilm formation and promotes tissue invasion through the secretion of candidalysin, a cytolytic peptide toxin encoded by the ECE1 gene. In this study, metabolites produced by Streptomyces ambofaciens CJD34, isolated from soil samples, were screened for antifungal activity. Methoxy-apo-enterobactin (compound 1) was identified as a potential inhibitor of C. albicans virulence. Treatment with compound 1 significantly suppressed ECE1 expression and candidalysin production. In a murine subcutaneous infection model, topical application of compound 1 reduced subcutaneous colonization by C. albicans. Molecular docking analysis suggested that the inhibition of ECE1 expression was not mediated by direct binding to known upstream transcription factors, indicating an indirect mechanism of action. Collectively, these findings highlight compound 1 as a promising antivirulence agent targeting candidalysin-mediated pathogenicity in C. albicans.