As the third largest freshwater lake in China, Taihu Lake has experienced severe cyanobacterial blooms and associated water quality degradation in recent decades, threatening the human health and sustainable development of cities in the watershed. The Yili River is a main river of Taihu Lake, contributing about 30% of the total nitrogen load entering the lake. Tracing the nitrate sources of Yili River can inform the origin of eutrophication in Taihu Lake and provide hints for effective control measures. This paper explored the nitrate sources and cycling of the Yili River based on dual nitrogen (δ15N) and oxygen (δ18O) isotopic compositions. Water samples collected during both the wet and dry seasons from different parts of the Yili River permitted the analysis of the seasonal and spatial variations of nitrate concentrations and sources. Results indicated that the wet season has higher nitrate concentrations than the dry season despite the stronger dilution effects, suggesting a greater potential of cyanobacterial blooms in summer. The δ15N-NO3− values were in the range of 4.0‰–14.0‰ in the wet season and 4.8‰–16.9‰ in dry, while the equivalent values of δ18O were 0.5‰–17.8‰ and 3.5‰–15.6‰, respectively. The distribution of δ15N-NO3− and δ18O-NO3− indicated that sewage and manure as well as fertilizer and soil organic matter were the major nitrate sources of the Yili River. Atmospheric deposition was an important nitrate source in the upper part of Yili River but less so in the middle and lower reaches due to increasing anthropogenic contamination. Moreover, there was a positive relationship between δ18O-NO3− and δ15N-NO3− in the wet season, indicating a certain extent of denitrification. In contrast, the δ18O-δ15N relationship in the dry season was significantly negative, suggesting that the δ15N and δ18O values were determined by a mixing of different nitrate sources.
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