Geothermal soils in Central Italy are characterized by high temperatures (up to 60°C), low pH values, and sparse vegetation. The biodiversity of entomopathogenic soil fungi and nematodes has never been assessed in these areas. Soil samples (N = 12) separated from each other by at least 100 meters were collected in the areas of Le Biancane Natural Park (Grosseto) and Sasso Pisano (Pisa). The Galleria bait method was used to assess the presence of entomopathogenic fungi and nematodes. A total of nine species of fungi and one nematode were isolated and molecularly identified. All these species came from samples collected in the immediate surroundings of geothermal spring, apart from one Beauveria pseudobassiana collected at about 16 meters from the nearest spring. Moreover, Pristionchus uniformis previously reported only from North America and few European countries is recorded for the first time in Italy.