MCC/eisosomes are membrane microdomains that have been proposed to participate in the plasma membrane function in particular by regulating the homeostasis of lipids, promoting the recruitment of specific proteins and acting as provider of membrane reservoirs. Results: Here we showed that several potential MCC/eisosomal protein encoding genes in the necrotrophic fungus A. brassicicola were overexpressed when germinated spores were exposed to antimicrobial defence compounds, osmotic and hydric stresses, which are major constraints encountered by the fungus during the plant colonization process. Mutants deficient for key MCC/eisosome components did not exhibit any enhanced susceptibility to phytoalexins and to applied stress conditions compared to the reference strain, except for a slight hypersensitivity of the Delta Delta abpil1a-abpil1b strain to 2 M sorbitol. Depending on the considered mutants, we showed that the leaf and silique colonization processes were impaired by comparison to the wild-type, and assumed that these defects in aggressiveness were probably caused by a reduced appressorium formation rate. Conclusions: This is the first study on the role of MCC/eisosomes in the pathogenic process of a plant pathogenic fungus. A link between these membrane domains and the fungus ability to form functional penetration structures was shown, providing new potential directions for plant disease control strategies.