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SPE–UPLC–MS/MS was used to investigate the occurrence of 18 target antibiotics in the surface water of Poyang Lake over different seasons of 2014–2015. The maximum concentrations of sulfadiazine, oxytetracycline, and doxycycline were 56.2, 48.7, and 39.7 ng/L, respectively. Compared with those in the other lakes or surface waters, the surface water of Poyang Lake contained moderate or below-average levels of antibiotics. The significantly lower concentrations (P < 0.01) of roxithromycin in June 2015 likely resulted from the dilution effect of water flow during the flood season. Antibiotic concentrations were higher in site P3-1 than in other sites (P < 0.01), whereas those in other sites (P1-1, P2-1, P5-1, P6-1, P7-1, P13-1, P16-1, P17-1, P18-1) were not significantly different (P > 0.05). Given that tetracyclines and sulfonamides are common veterinary medicines, the high concentrations of oxytetracycline, doxycycline, and sulfadiazine in site P3-1 might be closely related to agricultural production in the surrounding areas. The risk assessment of the main antibiotic contaminants revealed that the majority of the risk quotients of the target antibiotics were below 0.01, thereby indicating the minimal risk of these antibiotics to organisms at three different trophic levels. Sulfadimidine and sulfadiazine were identified as the main antibiotics that contribute to ecological risk in Poyang Lake, and that the daphnid is the main model organism exposed to these risks. This study provides important data for antibiotic pollution control and environmental protection in the study area and enriches environmental monitoring data on a global scale. [ABSTRACT FROM AUTHOR]