Lepidium papilliferum of southwest Idaho was previously treated as an infraspecific variety of Lepidium montanum. Chloroplast (cpDNA) sequences, nuclear ribosomal internal transcribed spacer (ITS) sequences, and AFLPs were used to test species delimitations and other possible evolutionarily significant units (ESU) based on genetic differentiation, isolation by distance (IBD), and genetic admixture among 32 L. montanum and 21 L. papilliferum collections from the western US. The L. papilliferum AFLP genotypes formed a monophyletic clade. However, the AFLP genotypes of L. montanum samples from eight western sites were more similar to L. papilliferum, which together comprise a regionally significant West clade showing significant differentiation from eastern L. montanum collections (East clade). Bayesian analysis of AFLP genotypes detected possible admixture between L. papilliferum and related western L. montanum collections. Neither taxa nor regionally significant AFLP clades displayed reciprocally monophyletic cpDNA or ITS sequences, but the AFLP clades showed stronger cpDNA differentiation and unique ITS alleles. The East and West clades fit models of speciation with relatively strong IBD within groups and weak IBD between groups, based on correlations between the average number of AFLP differences and geographic distances among collection sites, but comparisons between taxa did not fit this model. Conversely, relatively strong partial correlations between AFLP and taxonomic differences, controlling for geography, support taxonomic delimitations. Results suggest that L. papilliferum is a distinct subgroup of L. montanum influenced by speciation. However, gene flow or common ancestry between L. papilliferum and western forms of L. montanum provide a basis for other possible ESUs.