To understand the relationship between fish community structure and diversity and environmental factors in the middle reaches of the Yangtze River, fish and environmental surveys were conducted in five sections of the middle reaches of the Yangtze River (Yichang, Shishou, Jiayu, Wuhan and Hukou) in June and November 2021. Using these data, fish community structural characteristics, the spatial and temporal distribution pattern and its relationship with environmental factors were analyzed. A total of 16 335 individuals belonging to 76 species, 50 genera, 13 families, and 6 orders were collected, among which, Cyprinidae was the most abundant with 52 species, accounting for 70%. Among the three living habits, the number of resident fish species was the largest (80%). Among the three feeding types, the number of carnivorous fish species was the largest (46%). Among the three spawning types, sticky-egg fish had the largest number of species (50%). According to index of relative importance (IRI), the dominant fish species were small fishes, such as Pelteobagrus vachelli, Xenocypris argentea, Megalobrama terminalis, Coreius heterodon. The average Margalef richness index was 5.11, the average Pielou evenness index was 0.63, the average Shannon-Wiener diversity index was 3.23, and the average Simpson dominance index was 0.26. The abundance/biomass comparison curve showed that the fish communities in Yichang and Hukou were seriously disturbed, and those in Shishou, Jiayu, and Wuhan were relatively stable. The results of Cluster analysis and non-metric multidimensional scale ranking (NMDS) analysis showed that the fish communities in the middle reaches of the Yangtze River could be divided into three groups, and Yichang belonged to one group. Shishou, Jiayu and Wuhan belonged to one group, and Hukou belonged to one group. Redundancy analysis (RDA) was used to analyze the relationship between fish community and environmental factors. It was found that ammonia nitrogen (NH₄⁺-N) and bottom type were the main factors affecting the differences in fish community structure in the middle reaches of the Yangtze River. The results showed that compared with the historical data, the fish community structure in the middle reaches of the Yangtze River has changed significantly, the proportion of economic fish has decreased, and the fish individuals have become smaller and younger. These changes were caused by the joint effect of long-term human activities and natural environmental changes.