White-nose disease, caused by the dermatophyte Pseudogymnoascus destructans, is a devastating pathology that has caused a massive decline in the US bat populations. In Europe, this fungus and the related infection in bats have been recorded in several countries and for many bat species, although no mass mortality has been detected. This study reports for the first time the presence of P. destructans in Italy. The fungus was isolated in the Rio Martino cave, a site located in the Western Alps and included in the Natura 2000 network. Twenty bats, belonging to five different species, were analysed. The fungus was retrieved on eight individuals of Myotis emarginatus. The allied keratolytic species P. pannorum was observed on two other individuals, also belonging to M. emarginatus. Strains were isolated in pure culture and characterized morphologically. Results were validated through molecular analyses. Future work should be dedicated to understand the distribution and the effects of the two Pseudogymnoascus species on Italian bats.