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Heritage trees have important ecological, historical, and landscape values in cities. Rapid urbanization may cause dramatic change of ecosystem functions of cities, thereby inevitably affecting the growth performance of ancient trees. However, few studies have explored their species diversity and spatial differentiation on the medium scale in the scenario of urbanization in China. Here, we took Jiangsu Province in China, with developed economy in recent decades, as a typical case. Based on the provincial forest inventory data, we addressed the abundance, species richness, tree density, and species diversity of ancient trees in 13 cities, and their tree habitat, growth status, and tree age, as well. Then, we compared the spatial differentiation of tree attributes by 13 districts and nine tree habitats. We also applied detrended correspondence analysis (DCA) and redundancy analysis (RDA) to determine the leading factor influencing their distribution pattern. The 7678 heritage trees in Jiangsu belonged to 215 species. More than half of the trees were native with domination by Ginkgo biloba. Villages and farmlands accommodated the most heritage trees while parks and gardens harbored the most species. This indicates that sparsely-populated rural community and scenic areas with open space are conducive to accommodating more urban heritage trees. The tier 3 heritage trees (100–299 years) accounted for about 80% of the total. Overall, most ancient trees in Jiangsu grew well. The species diversity index (H) of 13 cities was between 1.98 and 3.39. The H value among the 13 cities was largely affected by elevation range shift, while the tree density by GDP per capita. DCA showed that the ratio of unique species was >40%, and that dominant species presented little habitat preference. Therefore, species diversity among different cities are affected by climate and topography, as well as human factors. With the accelerating urbanization process, tree habitat, cultural tradition, and urban history should be taken into consideration for management and conservation of heritage trees in the future. [ABSTRACT FROM AUTHOR]