Taxonomists have described chromatic and morphological variation for a number of European bumble bee species, although molecular studies have so far failed to find significant differentiation. We investigated the genetic variation of Bombus pascuorum from 13 locations by sequencing a fragment of the mitochondrial cytochrome b gene. We find considerably more variation than in published COII sequences for B. terrestris. The DNA sequences from these bees suggest that a difference exists between areas north and south of the Alps, and that the bees within these two areas are very similar over large areas. Therefore, mitochondrial cytochrome b sequences may be able to reveal geographic subdivision in B. pascuorum across the Alps.