The species richness of free-living vertebrates was analysed using mapping of occurrence within individual grid squares (12 x 11.1 km) over the territory of the Czech Republic. The data on species distribution were derived from recent distributional atlases published in the last 15 years, and the records originated mostly in the last 20 years. Altogether, 384 species of cyclostomes, bony fishes, amphibians, reptiles, birds and mammals were included in this study and their presence or absence was recorded in 678 grid squares. The species numbers ascertained in the 523 grid squares situated completely within the Czech Republic varied from 92 to 259 species, with a median of 182 species. The first two principal components explained 44.9 % of the total variance and separated two main habitat gradients based on values of different environmental, topographic, and demographic variables in particular squares. The PC1 represents a gradient from urban habitats at lower altitudes to more homogenous habitats with dominant coniferous forests and meadows situated at higher altitudes. The importance of natural habitats (represented by broad-leaved and mixed forests, as well as by protected areas) and landscape heterogeneity increases along the PC2. Generalized Linear Modelling for each group of vertebrates was fitted using the number of species of individual vertebrate groups as a response variable and the first two principal components as explanatory variables. The species richness of all vertebrate groups except for reptiles is highly dependent on the PC1. The number of fish, amphibian, and bird species in squares decreases with increasing value of the PC1, i.e. it is higher in urban areas at lower altitudes. By contrast, the number of mammal species is higher in uninhabited areas at higher altitudes. The gradient represented by the PC2 is highly significant for species richness of reptiles and mammals, and the number of species of both groups increases with increasing importance of natural habitats.