Time-dependent toxicity and key toxic components of five neonicotinoid mixtures under environmental concentrations in the Yangtze River Basin

Neonicotinoids (NEOs), as widely used insecticides, have raised increasing concerns regarding their ecological impacts. Current knowledge on the mixture toxicity of NEOs, particularly under environmental concentrations, remains insufficient. This study investigated the occurrence of five representative NEOs, including imidacloprid (IMI), thiamethoxam (THM), clothianidin (CLO), acetamiprid (ACE), and thiacloprid (THA), in surface water from the Linjiang (LJ) and Xuebu (XB) River Basins within the Yangtze River Basin. The total concentrations of the five NEOs ranged from 0.16 to 72.31 ng/L, with higher levels detected in the LJ Basin. High-throughput microplate toxicity analysis showed that the single THA and THM exhibited stimulatory effects (−17 % to 47 %) on Vibrio qinghaiensis sp.-Q67 (Q67). The 72 mixtures of the five NEOs exhibited not only inhibitory effects (up to 34 %) on Q67 at 12 h but also previously unobserved stimulatory effects (down to −32 %). These mixtures showed three different time-dependent toxicity characteristics during 0.25–12 h: 23 mixtures consistently exhibited stimulatory effects, 30 mixtures shifted from stimulatory to no effect, and 19 mixtures shifted from no effect to inhibitory effects. CLO, although showing no direct effect as a single compound, was identified as a key driver influencing mixture toxicity, exhibiting interactions with other components. Ecological risk assessment indicated that IMI, THM, CLO, and ACE posed potential ecological risks in the LJ Basin (risk quotients≥0.1). Based on the findings, management of NEOs in the LJ Basin may need to be strengthened, with particular attention to CLO due to its single ecological risk and key driver influencing mixture toxicity. This study provides comprehensively understanding of single and mixture toxicities of NEOs at environmental concentrations. • Five neonicotinoids (NEOs) were detected in the Linjiang and Xuebu River Basins. • 72 NEO mixtures at environmental levels reflected diverse exposure scenarios. • Mixtures showed time-dependent and biphasic (stimulatory-inhibitory) toxicity. • Clothianidin was identified as the key driver influencing the toxicity of NEO mixtures. • Four of the five NEOs posed potential ecological risks in the Linjiang River Basin.