Pesticides usually occur as mixtures of multiple chemicals in the natural aquatic ecosystem, so research based on the toxicity data of a single compound on aquatic organisms is not enough to accurately assess the actual toxicity risk of pesticides. There is still a gap in the research on the reproductive toxicity of combined insecticides, herbicides and fungicides on zebrafish (Danio rerio). In this study, zebrafish were used to systematically investigate the separate and combined reproductive toxicity of imidacloprid (IMI), acetochlor (ACT) and tebuconazole (TBZ), which are commonly used in rice fields. Adult zebrafish were exposed to the three pesticides individually and in combination for 28 days, and the number, heartbeat, deformation rate, body length, and swim bladder development of F1 offspring embryos were observed and the reproductive hormones testosterone (T), estradiol (E2), and vitellogenin (VTG) contents and the expressions of nine reproductive genes (ar, esr2a, vtg1, gr, star, fshr, hmgcrb, 3βhsd and vasa) in the testes of the male and the ovaries of the female F0 zebrafish adults were measured to evaluate the individual and combined effects. The results showed that exposure to the mixtures of IMI, ACT and TBZ resulted in a decrease in heartbeat, body length and swim bladder development and an increase in the deformity rate of F1 offspring embryos compared to the individual exposure groups. In the combined exposure group, the content of T decreased significantly and the content of VTG increased significantly in the testes of the males; the content of T significantly increased, while the content of E2 and VTG significantly decreased in the ovaries of the females, indicating that combined exposure showed a more obvious endocrine-disrupting effect compared to the individual exposures. In addition, the expression of nine reproductive genes was significantly altered compared to the individual exposure groups. Therefore, our results indicated that the mixture of IMI, ACT and TBZ caused fewer number of F1 embryos, higher developmental defects of F1, greater disruption in the content of reproductive hormones and the expression of reproductive genes compared to the individual pesticides at the corresponding doses. Therefore, the presence of pesticides in mixtures in the real water environment is likely to increase the toxic reproductive effects on zebrafish and cause more serious impacts on aquatic ecosystems.

Due to its high persistence in soil, oxyfluorfen has negative effects on environmental and human health. To reduce soil contamination and impacts on non-target organisms, biochar is introduced into soils to immobilize and sequestrate oxyfluorfen as a remediation practice. Three types of soils common in China were selected and biochar (rice hull, BCR) was added to investigate the desorption and bioavailability of oxyfluorfen after aging BCR for 0, 1, 3, and 6 months. Four chemical extraction methods were used to predict oxyfluorfen bioavailability. Results indicated that after addition of 0.5–2% unaged BCR, the desorption values of oxyfluorfen increased from 64–119 to 176–920 (μg/g)/(mg/L)ⁿ in the three soils compared with unamended soil. The bioaccumulation factor (BCF) values of oxyfluorfen in earthworms decreased from 0.80–1.7 to 0.10–1.56 after BCR addition. However, the desorption values decreased from 170–868 to 144–701 (μg/g)/(mg/L)ⁿ after aging. The bioavailability of oxyfluorfen in earthworms also increased after the aging treatments, while the BCF was still lower than with unaged BCR. The reduced BCF indicated lower exposure risk of oxyfluorfen to earthworms after amendment with biochar, even after aging 6 months. The bioavailability after extraction by Tenax showed a high linear correlation with uptake in earthworms, even after the 6-month aging treatment (R² > 0.80). Consequently, BCR could be a practical method to remediate contaminated soil and the 6h Tenax method could be a sensitive and feasible tool to assess the bioavailability of oxyfluorfen in soil.