Gray leaf spot (GLS) is a serious fungal disease caused by Magnaporthe grisea, recently reported on the important turfgrass and forage species, perennial ryegrass (Lolium perenne L.). This fungus also causes rice blast, which is usually controlled by host resistance, but durability of resistance is a problem. Little GLS resistance has been reported in perennial ryegrass. However, greenhouse inoculations in our lab suggest partial resistance is present, and QTL for GLS resistance have been detected on four linkage groups in an Italian x perennial ryegrass mapping population. Of these, two major QTL, one on linkage group (LG) 3 and the other on LG 6 were noted, with approximately 35% and 10% of the phenotypic variance explained, respectively. To confirm that QTL detected in that population are still detectable in the next generation, a resistant segregant from this population has been crossed with a susceptible perennial clone from a different cultivar background, to form a new mapping population segregating for GLS resistance. QTL analysis has been performed in the new population, using two different ryegrass field isolates and RAPD and RFLP marker based linkage maps for each parent, using many of the same markers as in the original population map. Results indicate the QTL on LG 3 is still significant in the new population, with LOD and percent of phenotypic variance explained ranging from 2.0 to 3.5 and 5% to 10% respectively. Also two QTL were detected in the susceptible parent, with similar LOD and phenotypic variance explained. Although the LG 6 QTL was not detected, the major QTL on LG 3 appears to be confirmed. These results will greatly assist in understanding the genetic architecture of GLS resistance in ryegrass, which will facilitate its use in perennial ryegrass breeding programs.