Leaf rust caused by Puccinia triticina Erikss. (Pt) is the most widely distributed and important wheat disease worldwide. The objective of the present study was to determine the frequency of Iranian Pt races, their virulence to key resistance genes and map quantitative trait loci (QTL) for resistance to different Pt races from 185 globally diverse wheat genotypes using a genome-wide association study (GWAS) approach. The virulence pattern of the 33 Pt isolates from various wheat-growing areas of Iran on 55 wheat differentials showed that the FKTPS and FKTTS were relatively frequent pathotypes among the 18 identified races. The weighted average frequency of virulence on the resistance genes Lrb, Lr3bg, Lr14b, Lr16, Lr24, Lr3ka, Lr11 and Lr20 were high (> 90%). However, low virulence on the resistant genes Lr2a, Lr9, Lr19, Lr25, Lr28 and Lr29 indicates that these genes are still effective against the pathogen population in Iran at present. GWAS on a panel of 185 wheat genotypes against 10 Pt races resulted into 62 significant marker-trait associations (MTAs) belonged to 34 quantitative trait loci (QTL) across 16 chromosomes. Among them, 10 QTLs on chromosomes 1A, 1B, 3B, 3D, 4A, 6D, 7A and 7D were identified as potential novel QTLs, of which four QTLs (QLr.iau-3B-2, QLr.iau-7A-2, QLr.iau-7A-3 and QLr.iau-7D-2) are more interesting, as they are associated with resistance to two or more Pt races. The known and novel QTLs associated with different Pt races found here, can be used in future wheat breeding programs to recombine different loci for durable resistance against leaf rust races.