We used 454-sequencing to identify and characterize 32 microsatellite loci for the Western Hermann's tortoise (Testudo hermanni hermanni) for which primer sequences, characteristics, and multiplexes are provided. These markers have been tested for one continental (Var) and one insular (Corsica) French population as well as for the Eastern Hermann's tortoise (Testudo hermanni boettgeri). For these 3 populations, 24 loci were polymorphic. These microsatellite markers will be particularly useful to assess the genetic diversity and characteristics of the different Hermann's tortoise populations and will have direct applications for its protection, notably with respect to reintroduction and conservation programs.