Abstract: As ecological indicator species, aquatic vascular plants and macrobenthos continuously reflect the ecological environment of rivers. To investigate the diversity, distribution characteristics, and their responses to environmental and socio-economic factors in the mainstream of the Yellow River Source Area, field surveys were conducted across eight counties in the source region. Structural characteristics indices (density, biomass, and species number) and diversity indices (Shannon diversity, Pielou's evenness, and Margalef's richness) of aquatic vascular plants and macrobenthos were calculated and analyzed. Redundancy analysis (RDA) was used to investigate the comprehensive effects of environmental and socio-economic factors on the diversity of aquatic vascular plants and macrobenthos. The results showed that a total of 6 aquatic vascular plants and 27 macrobenthos were collected in the mainstream of the Yellow River Source Area. The overall diversity of aquatic vascular plants was not high, and their distribution exhibited spatial heterogeneity. Among the counties, Maduo County recorded the highest values across all the indices for aquatic vascular plants, with all 6 species of aquatic vascular plants found there achieving a density of (248.00±35.55)ind./m² and a biomass of (22.92±4.27)g/m². Except for Maduo County and Gande County, there was no growth of aquatic vascular plants in other counties in the mainstream. The diversity of macrobenthos was abundant and widely distributed, among which Chironomidae had the richest species (6 species), while Chlaenius sp. was the most widely distributed, with records in seven other counties except Xinghai County. The density of macrobenthos in Dari County (206.67±85.61)ind./m² and the total number of species (12 species) were the highest, while the biomass of macrobenthos in Maduo County (7.04±3.31)g/m² was the highest. The RDA analysis results showed that the diversity of aquatic vascular plants and macrobenthos was influenced by multiple environmental factors, such as precipitation (explaining 31.81% of variance) and altitude (16.7%). It was significantly negatively correlated with socio-economic factors, such as population density (11.24%) and year-end livestock inventory (11.18%). To promote the protection and restoration of water ecology in the Yellow River Source Area, this study provided suggestions and countermeasures from both short-term operational measures and long-term mechanisms.