A mutant of Cochliobolus heterostrophus lacking the outer layer of extracellular matrix around its germ tubes and hyphae was obtained by mutagenizing protoplasts. The mutant not only lacks the outer matrix, but also produces much smaller lesions on corn leaves than nonmutant strains; the area of mutant lesions averages 0.6 mm2 compared with 5.8 mm2 for nonmutant lesions. Genetic analysis demonstrated that the failure to produce the outer matrix cosegregates with the reduced lesion size, indicating that the two traits are controlled by the same locus, designated Ecm1 (Extracellular Matrix Deficient). The mutant retains normal growth on media and normal abilities to germinate, form appressoria, and penetrate corn leaves. This indicates that the outer matrix is not necessary for infection prior to entrance of the fungus into the leaf. It also indicates that the pathogenicity defect in this mutant is manifested after penetration. To facilitate future tests of whether the pathogenicity defect is caused by the lack of the outer matrix, Ecm1 was mapped. Seven markers linked to Ecm1 were found by analysis of amplified fragment length polymorphisms. Ecm1 maps to chromosome 4; the closest markers to Ecm1 are 5 cM distant, which is estimated to represent about 115 kb.