Due to the excessive exploitation of traditional energy sources, the attention paid to water energy has increased in recent years. As an important means to effectively utilize water energy, reservoirs play an important role in drinking water, irrigation, flood control, and drought resistance. However, utilizing reservoirs often led to water quality issues resulting from the interaction of nutrients and hydrological conditions, especially due to the special structure of karst areas. Because of the change of hydrological conditions by the effect of dam construction, the dynamic of water quality will be more obvious in karst areas with a fast exchange of water and contaminants between underground and surface streams. In the present study, the change in water quality of a karst reservoir, the Muzhu Reservoir in the Houzhai Catchment, was studied. Long-term monitored datasets (1981-2002) and water quality datasets of more recent years were used to assess the effect on the water quality of reservoir expansion from the underground reservoir to the surface reservoir in a karst area. Long-term series datasets had shown that the hydro-chemistry type had been changed from HCO3-·SO42--Ca2+·Mg2+ type to HCO3--Ca2+ type in the short term after the reservoir's expansion. The chemical components of water originating from a rock background reduced markedly after the reservoir's expansion, whereas the content of the anthropogenic contribution in the water decreased after the expansion, except in April and May. Isotopic characteristics showed that δ15N-NO3- and δ18O-NO3- values were positively correlated before and after the reservoir expansion, but the slope of the linear regression before the expansion was 0.34, while the slope of the linear regression before the expansion was close to 0.7. This indicated that although denitrification and assimilation may occur simultaneously after the reservoir's expansion, the role of denitrification on nitrate removal decreased, which resulted in nitrate accumulation in the karst reservoir. The results highlighted that nitrate accumulation in karst reservoirs should be monitored to decrease nitrate concentration in the future.