Water erosion is a major threat to biodiversity, according to the European Commission’s Soil Thematic Strategy, as it negatively affects soil structure, soil fertility and water availability for plants. The island of Crete (Southern Greece) has been characterized as a biodiversity hotspot including several Natura 2000 (N2K)-protected areas. The aim of this study was to model the soil loss rate in Crete regarding species richness, habitat types and their conservation status, as well as the MAES (Mapping and Assessment of Ecosystem and their Services) ecosystem types. To this end, the RUSLE soil erosion prediction model was implemented, using freely available geospatial data and cloud-computing processes. The estimated average soil loss in the study area was 6.15 t ha−1 y−1, while there was no significant difference between the terrestrial N2K (6.06 t ha−1 y−1) and non-N2K (6.19 t ha−1 y−1) areas. Notably, the natural habitats of principal importance for the conservation of biodiversity (referred to as “priority” areas), according to Annex I to Directive 92/43/EEC, are threatened by soil erosion with an estimated mean annual soil loss equal to 8.58 t ha−1 y−1. It is also notable that grasslands, heathland and shrubs and sparsely vegetated areas experienced the highest erosion rates among the identified MAES ecosystem types. The results showed that soil erosion is a serious threat to biodiversity in N2K-protected areas. Therefore, there is a need for systematic spatiotemporal monitoring and the implementation of erosion mitigation measures.