In the context of urbanization, the degradation of river ecosystems has become increasingly prominent. Consequently, effectively assessment of river habitat conditions is an important foundation for restoration and protection of river ecosystem health. Shenzhen, as a window or opening city of China’s economic development, has extremely limited research conducted related to river habitat conditions. Therefore, in order to accurately elucidate the differences and main influencing factors of river habitats in basins with different degrees of urbanization in Shenzhen, a habitat assessment study was conducted on streams and rivers in two representative basins in Shenzhen. Based on characteristics of river habitats in Shenzhen, quantitative investigation and assessment of habitat conditions were conducted in 13 sampling sites in the highly urbanized Shenzhen River Basin and 12 sampling sites in the less urbanized Pingshan River Basin, in both wet (August) and dry (November) seasons in 2019. We adopted a comprehensive evaluation method to select 10 key habitat indicators from three categories: river bed, river channel, and riparian zone, to construct the habitat assessment index system and assessment method for Shenzhen rivers. The results showed that the overall condition of river habitat in the Shenzhen River Basin was poor, and the habitat condition of river sections with "good", "fair" and "poor" were 7.7%, 38.5%, and 53.8%, respectively. The overall condition of river habitat in the Pingshan River Basin was good, and the habitat condition of river sections with "excellent", "good", "fair" and "poor" were 8.3%, 41.7%, 41.7%, and 8.3%, respectively. The analysis of variance (ANOVA) results showed that there was no significant difference in river habitat conditions between the two seasons, which indicated that the overall river habitat conditions did not change too much in the short time period. There were significant differences in river habitat conditions between the two basins, and the overall river habitat condition of the less urbanized Pingshan River Basin was significantly better than that of the highly urbanized Shenzhen River Basin. Principal components analysis (PCA) results of river habitat assessment indices showed that human activity intensity, riparian stability, river channel modifications, substrate composition, riparian land use and vegetation diversity were the main factors driving river habitat conditions in Shenzhen. Based on the results of the river habitat assessment in Shenzhen, targeted restoration and management recommendations were proposed from three aspects: riverbed, river channel, and riparian zone, which have important implications for river habitat restoration and ecological conservation in Shenzhen.