The contemporary races of Puccinia striiformis f. sp. tritici (Pst) in Egypt during 2016–2018 were differentiated based on virulence and molecular patterns. Virulence patterns based on the reaction of the 17 World/European differential sets carrying stripe rust resistance genes (Yr genes) resulted in ten races including four new (first recorded in Egypt) and six old (previously recorded in Egypt). The new races were identified as 64E0 (virulence [V] Yr4, Su), 0E16 (V Yr8, 19), 66E0 (V Yr4, 7, 22, 23, Su) and 4E130 (V Yr2, 6, 7, 25, HVII), while the old were 0E0 (avirulence), 2E0 (V Yr7, 22, 23), 2E16 (V Yr7, 8, 19, 22, 23), 4E0 (V Yr2, 6), 6E4 (V Yr2, 6, 7, 22, 23, 25) and 70E4 (V Yr2, 4, 6, 7, 22, 23, 25, Su). Cluster analysis differentiated Pst races based on virulence frequency to Yr genes. Simple sequence repeat (SSR) markers were used to detect the molecular polymorphism of the Pst races. Clustering separated the old and new races into two groups, indicating their common ancestry since the new races were very distinct from the old races. Although clustering based on virulence revealed some evolutionary patterns, where the new races 64E0 and 66E0 may have probably evolved from the old races (2E16, 2E0, 6E4, 70E4) and the new race 4E130 may be evolved from the joint race 4E0. However, clustering based on molecular patterns indicated that the new races appear to be genetically distinct and may represent an exotic introduction rather than a mutation in isolates of the old races. A weak association between virulence and molecular patterns revealed that they are independent of each other. The SSR markers did not correspond to the virulences in the pathogen. Further studies on the potential virulence genes of the detected Pst virulences are needed.