AimsThe aim of this study was to address the toxicity of recently described polyene macrolide 32, 33-didehydroroflamycoin (DDHR) on a wide range of fungal pathogens and its potential to control plant fungal diseases. Methods and ResultsThe antifungal activity of DDHR invitro was examined against common human and plant pathogenic fungi using a broth microdilution assay and a disk diffusion assay. Minimum inhibitory concentrations ranged from 125 to 35gml(-1). A radial growth inhibition assay showed that DDHR inhibited mycelia growth, inducing mycelial necrosis and affecting sporulation. During the invivo assay on apple fruits administration of DDHR 1h before fungal inoculation inhibited spreading of the infection. Importantly, DDHR exhibited no phytotoxic effects on the model plant, Capsicum annum, verified by the plant growth rate and chlorophyll content. ConclusionsDDHR inhibits growth of various plant pathogens invitro with the strongest activity against Alternaria alternata, Colletotrichum acutatum and Penicillium expansum, and protects apple fruits from decay. Significance and Impact of the StudyThis is the first report of the inhibitory effect of DDHR on important pathogenic fungal isolates. DDHR could be a good scaffold for developing new antifungal agents for fruit and vegetable protection.