Late Pleistocene equus hydruntinus mitochondrial DNA from Iberian Peninsula: phylogenetic relationships

Abstract

The European ass (Equus hydruntinus) appeared in the fossil record around 350.000 years ago, and got extinct during the Holocene in Eurasia. Cranial and limb bone measurements have closely related Equus hydruntinus with the extant hemiones. This evidence is also supported by partial mtDNA sequences. Although Equus hydruntinus has been described in Iberian sites, most descriptions are mainly based on size proportions and dental morphology. Here we present for the first time a mtDNA HVR-I analysis of a tooth from a Late Pleistocene Iberian equid, tentatively attributed to Equus hydruntinus. An equid third lower right molar (AT-GE-189) was recovered from Galería de las Estatuas site, Cueva Mayor karst system (sierra de Atapuerca, Burgos, Northern Spain). This fossil was found in the Level 3, Sector 1 and it was directly radiocarbon dated. DNA extraction was performed in the ancient DNA laboratory at Centro Mixto UCM-ISCIII (Madrid, Spain) using silica spin columns. Four primer pairs were used to amplify a mtDNA HVR-I fragment. In order to study the evolutionary relationships of this tooth with other equids, a sample of extant and extinct Equus sp. sequences from caballine and non caballine lineages were collected, with special attention to the subgenus Hemionus. The Galería de las Estatuas sample radiocarbon dating revealed an age of 44.000 ± 1900 years BP. The 4 overlapping primer pairs yielded positive results, obtaining partial mitochondrial DNA sequence (np.15444 – 15946, according to the new Equus caballus Reference Sequence, JN398377). The sample AT-GE-189 displayed the 28 bp deletion previously described in other mtDNA HVR-I Equus hydruntinus sequences as well as from Equus kiang, Equus hemionus onager and Equus hemionus kulan. Moreover, the phylogenetic analyses showed that our Galería de las Estatuas sample clusters with the Equus hydruntinus sample from Scladina (Belgium), dated in around 40.000 years BP. We report a mtDNA sequence from an Iberian Equus hydruntinus and, due to the relationships with the other Equus hydruntinus sequences, we suggest the existence of an Equus hydrutinus group during the Late Pleistocene in Western Europe.