Technical Abstract: Gibberella zeae, a self-fertile, haploid filamentous ascomycete, causes serious epidemics of wheat (Triticum aestivum) head blight worldwide and contaminates grain with the mycotoxin deoxynivalenol. Anecdotal evidence dating back to the late nineteenth century indicates that G. zeae ascospores (sexual spores) are a more important inoculum source than are macroconidia (asexual spores), although the fungus can produce both during wheat head blight epidemics. To test this hypothesis the entire mating type (MAT1) locus was deleted from a self-fertile (MAT1-1;MAT1-2), virulent, deoxynivalenol-producing wild-type strain of G. zeae. The resulting mat-deleted (mat-1;mat1-2) strains were unable to produce mature perithecia with ascospores or to mate with the fertile strain from which they were derived. Mat-deleted strains retained the ability to produce macroconidia that were able to cause blight as assessed by direct injection into wheat heads in greenhouse and field tests. In two field tests the relative ability of mat-deleted strains to cause head blight was assessed by placing infested maize stalk pieces on the ground in irrigated experimental plots of wheat. In the first field test, a mat-deleted strain caused less head blight than the wild-type strain from which it was derived, but in the second field test, head blight levels were too low for results to be conclusive, likely due to unusually high temperatures. Availability of mat-null strains provides a means to determine the importance of ascospores in the biology of G. zeae and perhaps to identify novel approaches to control wheat head blight.