In the years 2016 to 2019 the virulence of the wheat leaf rust population in Slovakia was tested on Thatcher near-isogenic lines Lr1, Lr2a, Lr2b, Lr2c, Lr3a, Lr9, Lr10, Lr11, Lr13, Lr15, Lr17a, Lr19, Lr21, Lr23, Lr24, Lr26 and Lr28. A total of 111 rust isolates were analysed. Resistance genes Lr19 and Lr9 were effective to most of the tested isolates. Average virulence frequency to these resistance genes was only 1%. The gene Lr24 conditioned resistance to 92% of the tested rust isolates, gene Lr28 to 86% of the tested rust isolates. Low virulence frequency was also recorded to Lr2a (13%), Lr2b and Lr2c (both 12%). Average virulence to other tested Lr genes exceeded 82%. Thirty-eight winter wheat cultivars registered in the Slovak Republic were tested for the presence of Lr10, Lr19, Lr24, Lr26, Lr28, Lr34 and Lr37 by molecular markers. The gene Lr34 was the most frequent one in the tested cultivars, followed by Lr37 (both genes for adult plant resistance). The most widespread pathotypes in separate years and leaf rust severity in thirty-eight cultivars are also recorded.