Aerobic denitrification is a process reducing the nitrate into gaseous nitrogen forms in the presence of oxygen gas, which makes the nitrification and denitrification performed simultaneously. However, little was known on the diversity of the culturable aerobic denitrifying bacteria in the surface water system. In this study, 116 strains of aerobic denitrifying bacteria were isolated from the sediment, water and biofilm samples in Liangshui River of Beijing. These bacteria were classified into 14 genera based on the 16 S rDNA, such as Pseudomonas, Rheinheimera, and Gemmobacter. The Pseudomonas sp., represented by the Pseudomonas stutzeri, Pseudomonas mendocina and Pseudomonas putida, composed the major culturable aerobic denitrifiers of the river, followed by Ochrobactrum sp. and Rheinheimera sp. The PCA plot showed the unclassified Pseudomonas sp. and Rheinheimera pacifica preferred to inhabit in biofilm phase while one unclassified Ochrobactrum sp. and Pseudomonas resinovorans had higher abundance in the sediment. In the overlying water, the Pseudomonas stutzeri and Ochrobactrum rhizosphaerae were found to have higher abundance, indicating these aerobic denitrifiers had different habitat-preferable characteristics among the 3 phases of river system. The findings may help select the niche to isolate the aerobic denitrifiers and facilitate the bioaugmentation-based purification of the nitrate polluted surface water.