Technical Abstract: The appearance and anticipated spread of race TTKS (syn. Ug99) of Puccinia graminis f. sp. tritici have renewed interest in breeding for durable resistance to stem rust of wheat. In an attempt to determine the current status of stem rust resistance, 67 South African (SA) bread wheat cultivars and lines were tested with USA and East African races of P. graminis f. sp. tritici. Entries were also screened with DNA markers associated with Sr24 (Sr24#50) and Sr31 (iag95). Sr2 DNA marker (stm559n) data were compared with seedling chlorosis scores to validate the use of this marker in SA genotypes. Most cultivars interacted differentially with the races tested. DNA marker analysis confirmed the presence of Sr31 in seven entries. According to phenotype, Sr24 was postulated in several entries, but confirmed in less when combined with the marker data. Stm559n for Sr2 reliably amplified the correct alleles in most local and control lines. However, in several instances Sr2-associated alleles were amplified in presumably non-Sr2 carrying cultivars. This study emphasized that diversification of resistance sources, and more directed breeding for stem rust resistance, are needed as few SA wheat entries appear to have a broad-based resistance to stem rust.