Butterfly species can be sensitive to ecosystem disturbance and, therefore, suitable to be used as indicators of habitat quality. We determined species richness and diversity of butterfly species along five tourist trails in the northeast region of Portugal. These trails were in different landscape structures, varying from urban areas to areas extensively managed for agriculture (i.e., vineyards, meadows) to natural areas (i.e., grasslands, rivers, forests). A total of 522 butterflies representing 45 species belonging to 34 genera and 5 families of Lepidoptera were recorded. Of the taxonomic families represented in the survey, the Nymphalidae were most numerous (362 specimens, 22 species) followed by Pieridae (86 specimens, 11 species) and Lycaenidae (58 specimens, 8 species). Four species have a conservation status, an indicator of the risk of extinction they face at present or in the near future [Euphydryas aurinia (Rottemburg, 1775), Phengaris alcon (Denis & Schiffermüller, 1775), Hipparchia semele (L., 1758) and Melanargia lachesis (Hübner, 1790)], and these represent 6.9% of the total species identified. Among the five trails, diversity parameters varied with high values of species richness and diversity, low dominance of species, and moderate evenness of distribution. Additionally, butterfly species comparison among the trails revealed that Alvão and Vale do Corgo trails have most of the species in common, especially from Pieridae and Nymphalidae, while the Marão trail has more species associated exclusively to this trail. These results were also supported by hierarchical clustering performed with an average linkage aggregation method using Jaccard distance and by comparison between proportions of butterflies among trails within each family.