Previous morphological, allozyme and chloroplast DNA data have suggested that Pinus densata originated through hybridization between P. tabuliformis and P. yunnanensis. In the present study, sequence and restriction site analyses of maternally inherited mitochondrial nad1 intron were used to detect variation patterns in 19 populations of P. tabuliformis, P. yunnanensis and P. densata. A total of three mitotypes (A, B, C) were detected. All but one of the populations of P. yunnanensis possessed mitotype B while all populations of P. tabuliformis had mitotype A. Pinus densata populations, on the other hand, harboured both mitotypes A and B, which are characteristic of P. tabuliformis and P. yunnanensis, respectively. This result gives strong additional evidence supporting the hybrid origin of this diploid pine. The distribution of mitotypes indicated very different mating compositions and evolutionary history among P. densata populations. It seems that local founder populations and backcrosses may have played important roles in the early establishment of P. densata populations. The uplift of the Tibetan Plateau had a significant impact on the distribution of maternal lineages of P. densata populations.