Synergistic Enhancement of Acetamiprid Toxicity by Herbicide and Fungicide: Triple Mixture Outperforms Binary Combination

Pesticides are considered one of the main drivers of insect decline, and complex pesticide mixtures are frequently detected in our landscapes. However, the risk posed by mixtures of active substances is not systematically assessed. In this study using acute contact dose-response testing, we show that binary and triple combinations of relevant surrogates for major pesticide classes can lead to synergistic toxic effects on the non-target insect Oncopeltus fasciatus . First, we tested the individual active substances acetamiprid (insecticide), fluopyram (fungicide) and terbuthylazine (herbicide), detecting acute toxic effects only for acetamiprid. However, the combination of acetamiprid and terbuthylazine significantly increased toxicity compared to acetamiprid alone and the combination of acetamiprid and fluopyram with the effective dose 50 being 2.7 and 2.3 times lower respectively. Strikingly, a triple mixture of all active substances exceeded the toxicity of any other treatment. While the exact mechanisms remain unclear, we speculate that increased physiological stress contributes to the observed synergistic effects, as terbuthylazine and fluopyram have been shown to interfere, though to varying extents, with cellular energy metabolism, oxidative balance, detoxification and neurotransmission in several non-target organisms. These results are concerning and highlight the ecotoxicological uncertainty posed by traces of complex pesticide mixtures present in the air, soil, water and plants across landscapes. We therefore propose a systematic assessment of realistic exposure scenarios reflecting both agricultural practices and observed environmental mixtures.