We describe a new species of long-eared bat, genus Plecotus, from the island of Sardinia (Italy). The new species is clearly distinguishable from other European Plecotus species by its mitochondrial 16S rRNA gene (4.1–9.6% sequence divergence) as well as by a unique combination of morphological characters such as brownish colour of dorsal pelage, a relatively large thumb and thumb claw, an almost cylindrical form of the penis and the characteristic shape of the baculum. The most important morphological diagnostic character is a relatively long (≥ 18 mm) and wide (≥ 6 mm) tragus. The new species is currently known from three localities on Sardinia. In addition to the new species we discovered a lineage of P. auritus, which is substantially differentiated from continental P. auritus at subspecific level (1.2–2.7% of sequence divergence of the 16S rRNA gene). The existence of these two endemic bat taxa on Sardinia highlights the island's importance in the conservation of the European bat community.