Since our goal was to determine characteristic differences between Bromus richardsonii and B. ciliates, a discriminate analysis (DA), principal components analysis (PCA), multidimensional scaling (MDS), bivariate analysis, and an amplified fragment length polymorphisms (AFLP) analysis were undertaken on 93 herbarium specimens and 31 field-collected populations. A cytological survey of B. ciliates, B. richardsonii, and B. mucroglumis confirm previous reports that the first species is diploid (2n = 14) and the latter two are tetraploid (2n = 28). All taxa were correctly classified in the DA and important characters for each of the species were identified. Bromus richardsonii has lemmas with scattered hairs on the lower half between the mid nerve and margins [glabrous in B. ciliatus], anthers (1.2) 1.6-2.7 (3.4) mm long [(0.9) 1-1.4 (1.6) mm long in B. ciliatus], second glumes (7.8) 8.9 - 11.3 (13.2) mm long [(6.2) 7.1-8.5 (9.5) in B. ciliatus); and basal sheaths with dense, short to medium hairs [glabrous or with long hairs in B. ciliatus]. The PCA easily separated B. ciliatus and B. richardsonii into two well-defined groups and MDS mirrored the principal components analysis but displayed more overlap of individuals between the two groups. The AFLP-derived UPGMA dendrogram separated 154 individuals into two distinct clusters, one consisting entirely of B. ciliatus individuals and the other consisting of B. richardsonii individuals with six individuals of B. mucroglumis embedded within. Our study clearly indicates that there are distinctive morphological, cytological, and genetic differences to distinguish B. richardsonii and B. ciliatus as separate species.