It reports the preliminary analysis of the mitochondrial DNA of the last nomadic ethnic group of hunter-gatherers from Ecuador: the Waorani. The control region (HVI and HVII) of 111 Waorani individuals from two Amazonian communities was studied. It founds only two different haplotypes, which can be included in two (A and D) of the four major Native Americans mtDNA haplogroups (A–D). The haplogroup A2 was the most prevalent in both communities. The high homogeneity observed in the Waorani population might mirror the geographical and social isolation that have distinguished this population. Further studies will be necessary to deepen in the anthropological meaning of these results.