The newly described diploid species Cirsium greimleri (2n = 34; 2C = 1929.1±60.6 Mbp) belongs to Cirsium sect. Cirsium and is distributed sparsely throughout the Eastern Alps and Dinarides, whereas the closely related tetraploid vicarious species C. waldsteinii (2n = 68; 2C = 3682.3±69.8 Mbp) is endemic to the south-eastern Carpathians. The ploidy, genetic and morphological separation of both taxa is confirmed using flow cytometry, AFLP and morphometric analyses of 169 plants from 27 populations covering representatively distribution ranges of the respective species. The species differ in flower colour, with those of C. greimleri ruby red to brownish-purple and those of C. waldsteinii pinkish-purple to purple. The colour difference remains consistent when both species are cultivated together under the same conditions. Differences between the species in the size of the stomata, achenes, corollas, styles and pappus are statistically significant and congruent with differences in the ploidy level. Because both species are gynodioecious (their populations contain female and hermaphrodite plants), the generative features should be compared carefully with respect to gender (e.g. females with females) because the between-gender differences within the same species could be larger than the between-species differences. The basal and median cauline leaves of C. waldsteinii are narrower and more deeply lobed than those of C. greimleri. A slight difference in the flowering period is detected when both species are cultivated together, with C. waldsteinii flowering two weeks earlier than C. greimleri. Both species share ecological/habitat preferences for subalpine woodland tall-forb vegetation. While C. waldsteinii hybridizes extremely rarely with co-occurring diploid congeners, C. greimleri produce hybrids very often, which increase the risk of its extinction via genetic erosion. K e y w o r d s: