Abstract: In order to understand the species composition and diversity characteristics of the major fish communities in typical riffle-pool reaches of the fluctuating backwater area of the upper Yangtze River, species diversity analyses of fishes in the Guangyangba and Dajiantan reaches were conducted using environmental DNA (eDNA) technology. The results showed that a total of 77 species of fishes of 11 orders, 18 families, 54 genera were monitored in the two river sections, dominated by fishes of the family Cyprinidae, including 14 species endemic to the Yangtze River, 8 invasive species, and 12 species seldom reported in other surveys. Taken together, Cyprinus carpio, Ctenopharyngodon idella, and Hemiculter bleekeri were the main dominant species in this survey. The α-diversity analyses indicated higher species diversity in the tributaries of both tow reaches, and a relatively even distribution of species in the Guangyangba reach compared to the Dajiantan reach. Finally, the results of the classification of fish ecological group showed that the fish in the study reaches were mainly polyphagy, middle-lower, sedentary, adhesive egg-producing, living with cluster and preference for slow-flowing. The results of this study demonstrate the effectiveness of eDNA technology in revealing information on fish species diversity and ecological groups, and provide targeted advice for the protection of fish diversity in the Yangtze River to guide the design of river habitat restoration projects and ensure that the restoration measures can meet the habitat needs of different fish species.