Andean streams cover steep altitude gradients and locally leach metal-rich bedrock, creating highly selective habitat conditions. Chironomids are among the few dominant insect taxa present under the harshest conditions in Andean high altitude streams, but it remains unclear whether their dominance is due to the adaptive capacity of a few species (population differentiation) or to a diversity of species with different capacities to cope with environmental extremes (species composition). Therefore, the aim of this study was to assess whether metals and altitude drive the genetic diversity of chironomids in Andean streams. We measured metal concentrations and altitude-related conditions, such as UV-B radiation, water temperature and oxygen, in reference and metal-rich streams located at both 3000 and 4000 m a.s.l. The genetic composition of the chironomid communities from these streams was determined by mitochondrial cytochrome oxidase I (COI) gene sequencing, and a phylogenetic tree was constructed. The concentrations of all metals were higher in the metal-rich streams than in the references streams. The UV-B radiation level at 4000 m was notably higher than at 3000 m, while the water temperature followed the opposite trend. At 3000 m, the reference site was inhabited by six putative species, completely different from the three present at 4000 m. Only one putative species, which did not occur at the reference sites, was present at both the metal-rich sites at 3000 and 4000 m. The differences in putative species composition between 3000 and 4000 m indicated a strong sorting of species according to altitude. However, the unique putative species present at the metal-rich sites at both 3000 and 4000 m indicated that altitude-driven selection was overridden by the extreme selection pressure exerted by metal exposure. We conclude that altitude limits the distribution of chironomid taxa, but metal selection leads to predominance of a unique metal-tolerant taxon.