This research characterized the genetics of resistance of wild barley accession PI 466423 to a widely virulent pathotype of Cochliobolus sativus . Breeding lines were identified that combine the Midwest Six-rowed Durable Resistance Haplotype and resistance to the virulent isolate ND4008. Spot blotch, caused by Cochliobolus sativus, is a historically important foliar disease of barley (Hordeum vulgare L.) in the Upper Midwest region of the USA. However, for the last 50 years this disease has been of little consequence due to the deployment of resistant six-rowed malting cultivars. These durably resistant cultivars carry the Midwest Six-rowed Durable Resistant Haplotype (MSDRH) comprised of three Quantitative Trait Loci (QTL) on chromosomes 1H, 3H and 7H, originally contributed by breeding line NDB112. Recent reports of C. sativus isolates (e.g. ND4008) with virulence on NDB112 indicate that widely grown cultivars of the region are vulnerable to spot blotch epidemics. Wild barley (H. vulgare ssp. spontaneum), the progenitor of cultivated barley, is a rich source of novel alleles, especially for disease resistance. Wild barley accession PI 466423 is highly resistant to C. sativus isolate ND4008. To determine the genetic architecture of resistance to isolate ND4008 in PI 466423, we phenotyped and genotyped an advanced backcross population (N = 244) derived from the wild accession and the recurrent parent 'Rasmusson', a Minnesota cultivar with the MSDRH. Disease phenotyping was done on BC2F4 seedlings in the greenhouse using isolate ND4008. The Rasmusson/PI 466423 population was genotyped with 7842 single nucleotide polymorphic markers. QTL analysis using composite interval mapping revealed four resistance loci on chromosomes 1H, 2H, 4H and 5H explaining 10.3, 7.4, 6.4 and 8.4 % of the variance, respectively. Resistance alleles on chromosomes 1H, 4H and 5H were contributed by PI 466423, whereas the one on chromosome 2H was contributed by Rasmusson. All four resistance QTL are likely coincident with previously identified QTL. Agronomically advanced two- and six-rowed lines combining the MSDRH and resistance alleles to isolate ND4008 have been identified and are being utilized in breeding. These results reaffirm the value of using wild relatives as a source of novel resistance alleles.