Genetic structure of phytophagous insects has been widely studied, however, relative influence of the effect of geographic isolation, the host plant or both has been subject of considerable debate. Several studies carried out on bark beetles in the genus Dendroctonus evaluated these factors; nonetheless, recent evidence has shown that genetic structuring is a more complex process. Our goal was to examine the effect of geographic isolation on genetic structure of the Douglas-fir beetle Dendroctonus pseudotsugae. We used mtDNA cytochrome oxidase I (COI) sequences and RAPD markers. One hundred-seventy-two individuals were obtained from 17 populations, for which we analyzed 60 haplotypes (among 172 sequences of COI gene, 550 bp long) and 232 RAPD markers (7 primers). Analyses of molecular variance (AMOVA and SAMOVA), F-statistics and linear regressions suggest that the genetic structure of D. pseudotsugae is strongly influenced by geographic distance. We found that D. pseudotsugae has high intra- and inter-population genetic variation compared with several other bark beetles. Genetic differences among populations based on COI and RAPD markers were correlated with geographic distance. The observed genetic differences between northern (Canada-USA) and southern (Mexico) populations on Pseudotsuga menziesii var. glauca confirm that these two sets of populations correspond to previously assigned subspecies.