Urbanization causes the introduction of exotic plant species and landscape interventions that deeply modify native ecosystems. Herein, we analyzed the composition and abundance of urban tree species to investigate similarities between Brazilian cities and regions, discussing the possible existence and scale of biotic homogenization. We performed floristic assessments of 45 Brazilian urban areas in the North (5), Southeast (14), South (12), Northeast (10) and Center-west (5) regions, and analyzed the richness, diversity, occurrence and abundance of urban tree flora and also their similarity. Brazilian cities, generally, presented low diversity with a high dominance of a few native and non-native species. Some cities of the same region were grouped as result of different groups of native and non-native species’ relative abundance. Latitudinal and longitudinal gradients were identified and were associated with differences in temperature, precipitation and altitude. Species had different roles in cities similarity; some native and non-native species contributed to cities differentiation, while other widespread non-native and native species occurring outside their distribution contributed to increased city similarity. So, there is no generalized homogenization process caused by non-native species, and species should be evaluated in relation to their risks and necessity of management, depending on the scale and services needed.