Four vahlkampfiid amoebae were isolated from freshwater samples taken on the island of Tenerife. Ribosomal DNA sequence analysis identified two isolates to belong to the genus Naegleria, representing novel species with their closest relatives N. gallica and N. philippinensis respectively. The two other isolates belong to the genus Vahlkampfia and represent also novel species, with as their closest relatives V. avara and V. inornata respectively. The new Vahlkampfia spp. are more divergent from their nearest relatives than the new Naegleria spp. are to their closest relatives. The latter is probably due to the already high number of described Naegleria spp. in contrast to the few known Vahlkampfia spp. There are no morphological features in these new species of either genus that allow their identification. Only molecular typing can discriminate species within the vahlkampfiids.