| Pesticides have been widely used in agriculture to control diseases,pests,and weeds,improve crop yields and ensure food security.Herbicide is used to control weeds and small shrubs of a class of pesticides.Pyroxsulam is a typical triazopyrimidine sulfonamide herbicide,which plays an important role in weed control in the wheat field.With the spread of pyroxsulam,it will inevitably enter the aquatic environment.In addition,pyroxsulam degrades slowly and remains for a long time in the water,which is easy to pollute the water environment and harm the safety of aquatic organisms.However,the previous studies on pyroxsulam mainly focused on weed resistance and control effects,and its ecotoxicological research on aquatic organisms was lacking.In this study,pyroxsulam was used as a pollutant,and zebrafish embryos and adults（Danio rerio）at different growth stages were used as test organisms.The acute toxicity of pyroxsulam to zebrafish embryos and adults was studied.According to the residual concentration of pyroxsulam in the environment,the effects of different concentrations of pyroxsulam（0.2,2 and 10μg/L）on oxidative stress,and DNA damage of zebrafish embryos（120 h）and adults（14 and 28 d）were systematically studied.The quantitative real-time polymerase chain reaction（qRT-PCR）was used to analyze the effects of pyroxsulam on apoptosis-related genes in zebrafish embryos and adults.The integrated biomarker response index（IBR）was calculated to evaluate the combined toxicity of pyroxsulam to zebrafish embryos and adults.This study comprehensively evaluated the toxicity of pyroxsulam at different growth stages of zebrafish,providing valuable data for understanding its effects on aquatic ecosystems.The main results of this study are as follows:（1）The 96 h median lethal concentration(LC50)for zebrafish embryos and adults was greater than 100 mg/L,indicating low toxicity.（2）Pyroxsulam reduced the content of reactive oxygen species（ROS）in embryos but increased the content of ROS in adults.Pyroxsulam inhibited the activities of superoxide dismutase（SOD）and catalase（CAT）and induced the activity of glutathione peroxidase（GSH-Px）.When the concentration was greater than 2μg/L,the activities of CAT and GSH-Px in embryos did not change with the increase of the concentration.Pyroxsulam also inhibited SOD activity in adults,but induced CAT activity and inhibited glutathione S-transferase（GST）activity.The higher the concentration is,the more obvious the inhibition effect is.Pyroxsulam increased the content of malondialdehyde（MDA）,a lipid peroxidation product,in embryos and adults,resulting in lipid peroxidation.（3）In both embryos and adults,pyroxsulam could significantly increase the content of8-hydroxydeoxyguanosine（8-OHdG）,which leads to DNA damage in zebrafish,and there was a dose-response relationship,namely,with the increase of pyroxsulam exposure concentration,the more serious the DNA damage to zebrafish.However,DNA damage in adults showed no time-effect relationship.Pyroxsulam has toxic effects on zebrafish in different life stages.（4）After 120 h of exposure,pyroxsulam significantly down-regulated the human tumor suppressor gene（p53）in zebrafish embryos and inhibited cell apoptosis through the mitochondrial pathway;The ratio of Bcl-2 gene（Bcl-2）to Bcl-2-associated X protein（Bax）was also increased,and the medium concentration（2μg/L）was a threshold.The Bcl-2/Bax ratio did not change significantly when the concentration increased.In addition,the relative expression levels of caspase 3,caspase 8,and caspase 9 genes in the cysteinyl aspartate specific proteinase（caspase）family were significantly down-regulated.The down-regulation of caspase 9 had a dose-response relationship.The changes of p53,Bcl-2,Bax,caspase 3,caspase 8,and caspase 9 genes will eventually inhibit cell apoptosis in the embryos.（5）After exposure to pyroxsulam,the expression trend of apoptosis-related genes in adults was similar to that in embryos at the early stage of the experiment（14 d）,and apoptosis was inhibited.At the end of the experiment（28 d）,Bcl-2/Bax and caspase 8 returned to the level of the solvent control group,but p53,caspase 3,and caspase 9 were regulated,and the medium concentration（2μg/L）was a threshold.There was no significant change in the expression level of caspase 9 when the concentration increased,which eventually induced apoptosis of cells in adults.（6）By calculating IBR,apoptosis is more easily affected than oxidative damage in both embryos and adults.The toxicity of pyroxsulam to zebrafish embryos and adults increased with the increase of concentration,showing a dose-response relationship,and the oxidative stress response in zebrafish became more obvious with the increase in concentration.With the increased time,the toxicity of pyroxsulam in adults decreased,especially at low concentration（0.2μg/L）and medium concentration（2μg/L）. |
