Human disturbances have altered global stream ecosystems in multiple ways. Understan-ding spatio-temporal dynamics of aquatic organism composition and their relationships with environmental factors play an important role in revealing changes in the structure and function of river ecosystems. The macroinvertebrates and environmental parameters were sampled in the Yongding River Basin at spring (March), summer (July), and autumn (November) in 2017. The results showed that chironomids were the dominant group in the Yongding River Basin, including Orthocladius, Chironomus, Glyptotendipes, and Polypedilum. Results of the cluster analysis showed significant spatial variations of macroinvertebrate community. Sampling sites were clustered into two groups based on the similarities of macroinvertebrate composition in the Yongding River. Sampling sites belonging to group 1 mainly located in the Sanggan River and Dongyang River, with Orthocladius sp.being the dominant species. Sampling sites belonging to group 2 mainly located in the Yang River, Guishui River and main section of the Yongding River, with Glyptotendipes sp. being the dominant species. The biomass, Shannon index, Margalef index, Pielou evenness index and taxa richness of group 2 were significantly higher than those of group 1. Higher values of biological indices indicated better ecological condition of the stream ecosystem. The density, biomass and diversity indices of summer were significantly lower than those of both spring and autumn. Results of Canonical correspondence analysis (CCA) showed that the spatio-temporal distribution of macroinvertebrate in the Yongding River was mainly influenced by water temperature, ammonium, pH, dissolved oxygen, and total phosphorus. From the perspective of hydrological characteristics, flow velocity and flow capacity were significantly correlated. The significant positive correlations between hydrological parameters of velocity and capacity and biological indices of density and EPT% in group 2 indicated the preference of fast flow to sensitive taxa of EPT (Ephemeroptera, Plecoptera, and Trichoptera). The negative correlations between flow capacity and biodiversity indices implied the adverse effects of high flow on macroinvertebrate biodiversity. Our results confirmed that the composition and biodiversity of macroinvertebrate were affected by both water environment and hydrology, which reflected the multiple human activities in aquatic ecosystems of the Yongding River. It is important to control water contamination and sustain environmental flow at the specific reaches for the restoration of Yongding River.