Forest cover in Europe has substantially increased in recent decades, resulting in extensive secondary forest cover (ca. 10-20 years) that is too young for many specialist forest-dwelling species. In the Mediterranean region, forests have historically been logged with detrimental effects on local biodiversity. Only a few small forest patches remain untouched. Specialist forest-dwelling species are often less studied than other species due to their lower population densities and the inherent difficulties involved in sampling. In fact, some forests species have only recently been discovered or described such as the Myotis alcathoe (Alcathoe bat) and so there is a remarkable dearth of information regarding their natural history, habitat requirements and conservation status. A total of 18 M. alcathoe bats were captured and radio-tracked in a 100-year-old Mediterranean forest which led us to locate and describe 18 different tree roosts. The structural traits of each roost and the surrounding habitat were studied to unravel the factors involved in roost selection. Alcathoe bats were found roosting in a wide variety of sites (holes, cracks and under bark) and tree species. Around 70% of the roosts were found in dead or decrepit trees. However, except for roost height, no special traits were selected. Due to the maturity of some common holly Ilex aquifolium, this typically secondary bush species was often used as a breeding roost. Our results provide new insights into how old and small secondary tree species play an essential role in the conservation of certain forest species. Considering that in 20 years of bat surveys in the region, breeding colonies of this forest species have only been found in this forest characterized by its remarkable maturity (50-100 dead trees/ha), we suggest that current forest management guidelines (recommending densities of 5-10 dead trees/ha), might not be enough to ensure the presence of breeding colonies of these tree-dwelling bats.