Conservation biogeography, which applies principles, theories, and analyses of biodiversity distribution patterns to address conservation challenges, can provide valuable insight and guidance to policy making for protection of biodiversity at multiple scales. The temperate and tropical ecosystems of the Nearctic-Neotropical transition in the small western state of Colima, Mexico, support a mosaic of remarkably diverse fauna and flora and provide a rare opportunity to determine spatial distribution patterns of terrestrial vertebrate species, assess human-induced threats, and identify potential conservation strategies. We analyzed the spatial distribution patterns and correlated them with the current land cover and extent of the protected areas. Despite its limited geographic extension, 29% (866) of all vertebrates, and almost a quarter of both endemic and threatened species in Mexico, live in Colima. Our analysis identified clear high-richness concentration sites (i.e., "hotspots") coincident for all groups and that elevation and both temperate and tropical ecosystems composition exert significant influence on richness patterns. Furthermore, current species´ distribution also showed significant correlation with natural and disturbed landcover. Significant hotspots for all species groups coincided poorly with the limited protected areas in the state (only 3.8%). The current state of natural land cover (less than 16%) in the state, coupled with its remarkable biological importance, highlights the need for further complementary conservation efforts including expansion and creation of new protected areas, significant restoration efforts and other conservation measures to maintain this uniquely biogeographic and biological diverse region of the country.