Knowledge of the responses of reservoir ecosystems to anthropogenic and climatic impacts on multi-centennial timescales is relatively poor, but is crucial for the management of numerous reservoirs in China. In this study, the history of an ancient reservoir system was reconstructed for the last ∼ 700 yr based on multiple dating methods (210Pb, 137Cs, and 14C) and geochemical data (e.g., the mass accumulation rates, loss-on-ignition values, total organic carbon, total nitrogen, total phosphorus contents, and stable carbon isotope ratios of organic matter) obtained for sediment cores from Gongqing Reservoir in Guizhou Province, southwestern China. Combined with historical literature and regional climatic records, the response of the aquatic ecosystem in Gongqing Reservoir to water level fluctuations, anthropogenic activity, and climatic changes during the last ∼ 700 yr was assessed. The results suggest that dramatic changes in the water level due to dam heightening in 1950 s significantly changed the aquatic ecosystem of Gongqing Reservoir. During periods of low water levels (average water depth = 1–2 m; 1345–1957 CE), aquatic macrophytes contributed more to the sedimentary organic matter, whereas phytoplankton increasingly contributed more during periods of high water levels (∼5 m; 1957–2020 CE). During the late stage (1700–1957 CE) of low water levels, the reservoir ecosystem was significantly affected by human activities. Combined with regional geological and historical records, we consider that increased population and enhanced human activities have markedly affected vegetation conditions in western Guizhou Province since ∼ 1700 CE. After 1957 CE, anthropogenic activities were key in affecting the aquatic ecosystem of Gongqing Reservoir. Especially in the last decade, large housing developments and enhanced agricultural activities in the watershed have led to enhanced catchment erosion and transport of more nutrients into the reservoir, thus promoting the algae productivity. These results provide new insights into how reservoir ecosystems respond to water level fluctuations, anthropogenic activity and climatic changes on multi-centennial timescales.