Aims: To assess the antimicrobial efficacy of ascopyrone P (APP), a secondary metabolite formed by the fungi Anthracobia melaloma, Plicaria anthracina, Plic. leiocarpa and Peziza petersi belonging to the order Pezizales. Methods and Results: In vitro testing using a well diffusion procedure showed that APP at a high concentration (approximately 5%) inhibited the growth of Gram-positive and Gram-negative bacteria. Using an automated microbiology reader, growth curve analysis showed that 2000-4000 mg l(-1) APP caused total or significant bacterial inhibition after incubation for 24 h at 30degreesC. Against certain yeast strains, 1000- 2000 mg l(-1) APP enhanced growth, although at higher concentrations inhibition of some yeasts was observed. Clostridium and fungal strains were not sensitive to 2000 mg l(-1) APP. No significant cidal effect was observed after 2 h against Listeria monocytogenes or Escherichia coli . Results were identical whether the APP samples tested had been produced enzymatically or chemically. Conclusions: At a level of 2000 mg l(-1) , APP demonstrated growth inhibitory activity against a broad range of bacteria, but not yeasts or moulds. Significance and Impact of the Study: A possible application for this novel natural antimicrobial is in food preservation, to control the growth of Gram-negative and Gram-positive bacteria in raw and cooked foods. Effective dosage levels would be 500-4000 mg kg(-1), depending on food type. The efficacy, organoleptic and safety aspects of this compound in food still need to be assessed.