Cardamine amara, an Eurasian species comprising several subspecies, was examined through morphometric and amplified fragment length polymorphism (AFLP) analyses to re-evaluate subspecific status of tetraploid C. amara subsp. olotensis from Catalonia (NE Spain) and to resolve the taxonomic position of Central Italian populations usually placed within C. amara and/or C. raphanifolia s. l. A presumptive close relative from the Caucasus, diploid C. wiedemanniana, was also analyzed for AFLPs. Morphometric results show Catalonian and Italian populations to be similar to each other, but distinct from other C. amara. With both principal coordinate analysis and the neighbour-joining tree based on AFLP data, three main groups are delimited: (1) tetraploid Catalonian C. amara subsp. olotensis and Italian populations, (2) all other diploid and tetraploid subspecies of C. amara, and (3) diploid C. wiedemanniana. As a result of this study, Italian and Catalonian populations are herein treated as a single species, C. amporitana, distinct from C. amara. Catalonian populations appear genetically depauperate, perhaps having experienced bottlenecks during Pleistocene glaciation, or originating via long-distance dispersal from the Apennine Peninsula.