Fusarium graminearum is an important plant pathogen that causes head blight on wheat and barley.The outbreak of the scab results in directly reduced food production and grain quality, while the infested grains containing mycotoxins pose a serious hazard to food safety as well as human and animals health.However, there is rarely report about the regulatory mechanisms of biological processes, including its development, pathogenicity and secondary metabolism in molecular level.The vacuolar protein sorting protein Vps27, which is a component of ESCRT-0 involved in the multivesicular body (MVB) sorting pathway, is involved in the regulation of diverse cellular processes.In this study, we explored functions of FgVps27 in the F.graminearum using a gene replacement strategy.The deletion mutant exhibited a reduction in growth rate, aerial hyphae formation and hydrophobicity.Deletion of FgVps27 not only caused the lost capacity of conidial production at 28℃, but also unable to generate perithecium in sexual reproduction.Interestingly, while the mutant was cultivated at a relatively lower temperature (24℃), it could produce a few fragmented conidia.Pathogencity assays showed that the FgVps27 deletion mutant was impaired in virulence on flowering wheat head.Mycelia of the FgVps27 mutant had weak cell wall and cytoplasm membrane, and were also hypersensitive to osmotic stress.FgVps27 mutants also showed a significant reduction in red pigment formation in different media.Additionally, FgHse1 and FgVps23, another component of ESCRT, interacting with FgVps27 was identified by yeast two hybrid assays in current study.Taking together, our results indicated that FgVps27 plays a critical role in a variety of cellular and biological processes in the F.graminearum, such as growth, sporulation, pathogen invasion and secondary metabolism.