Toxic Effects Of Single And Combined Exposure To Microplastics And Atrazine On Earthworm

With the widespread application of mulch film mulching cultivation technology,the use of mulch film and the cultivation area of mulch film in my country are increasing year by year.However,since the raw materials for the production of mulch film are mostly chemically stable polyethylene or polyvinyl chloride,which are difficult to degrade in the environment,and the current lack of mulch film recycling technology,the problem of mulch film residue has become increasingly prominent.The residual mulch film in the soil is gradually broken and decomposed to form microplastics under the combined effects of farming,ultraviolet radiation,weathering,and microbial degradation.As a new type of pollutant,microplastics themselves have been found to have potential adverse ecological effects,and their compound pollution with coexisting pollutants has also attracted widespread attention from scholars at home and abroad.Atrazine is a triazine herbicide,which is widely used in agricultural production.However,due to its long residual period and years of excessive use,atrazine has been widely detected in soil,groundwater and surface water.Studies have shown that atrazine can be bioaccumulated and produce toxic effects on non-target organisms after entering the environment.Further studies have shown that atrazine can affect the reproductive system,endocrine system,central nervous system and immune system.With the widespread use of mulch film and atrazine,the pollution of mulch film-derived microplastics and atrazine in farmland soil has become more and more serious.However,there are relatively few studies on the combined pollution of mulch filmderived microplastics and atrazine.In this study,unused polyethylene mulch film and farmland residual polyethylene mulch film were used as raw materials to prepare unaged polyethylene mulch film-derived microplastics(PE MPs)and field-aged polyethylene mulch film-derived microplastics(PEaged MPs),atrazine was used as a typical organic pollutant in the soil,and Eisenia fetida was used as the test organism to systematically study the oxidative stress,DNA damage and physiological related genes expression in earthworm after single and combined exposure(28 d)to above-mentioned mulch film-derived microplastics(PE MPs and PE-aged MPs,550～1000μm)and different concentrations of atrazine(0.02 mg/kg and 2.0 mg/kg).The integrated biomarker response index(IBR)was used to evaluate the toxicity and differences after single and combined exposure to mulch film-derived microplastics and atrazine on earthworm from the above three aspects.Our study can provide data support for the ecological risk assessment of the combined pollution of microplastics and coexisting pollutants.The main results of this study are as follows:(1)Single and combined exposure to mulch film-derived microplastics(PE MPs and PEaged MPs)and atrazine(0.02 mg/kg and 2.0 mg/kg)can lead to the contents of reactive oxygen species(ROS)significantly increase,lead to the activities of superoxide dismutase(SOD),catalase(CAT)and glutathione S-transferase(GST)significantly reduce in earthworm,and lead to a significant increase in the content of 8-hydroxydeoxyguanosine(8-OHd G),a product of DNA oxidative damage in earthworm,which in turn causes oxidative damage to earthworm;cause DNA strand breaks in earthworm coelomocyte,that is,olive tail moment(OTM)values significantly increase,which in turn causes DNA damage to earthworm coelomocyte;lead to significant changes in the relative expression levels of calreticulin(CRT),annetocin(ANN),translationally controlled tumor protein(TCTP)and heat shock protein 70(Hsp70)genes in earthworm.(2)Comparing the single exposure to two kinds of mulch film-derived microplastics,it can be found that the IBR values(oxidative stress),IBR values(physiologically related gene expression)and OTM values in the PE-aged MPs exposure group were higher than those in the PE MPs exposure group,indicating that exposure to PE-aged MPs induced higher toxicity on earthworm compared with PE MPs.Comparing the single exposure to different concentrations of atrazine,it can be found that the IBR values(oxidative stress),IBR values(physiologically related gene expression)and OTM values in the 2.0 mg/kg atrazine exposure group were higher than those in the 0.02 mg/kg atrazine exposure group,indicating that the toxic effect of atrazine on earthworm showed an obvious dose-effect relationship,that is,as the concentration of atrazine increased,the toxic effect of atrazine on earthworm was also enhanced.(3)The IBR values(oxidative stress)and OTM values in most of the combined exposure groups of mulch film-derived microplastics and atrazine,especially in the combined exposure groups of mulch film-derived microplastics and high concentration of atrazine(2.0 mg/kg)were higher than those in the corresponding single exposure group,indicating that the combined exposure to mulch film-derived microplastics and atrazine can enhance its oxidative stress and DNA damage on earthworm;in contrast,the IBR values(physiologically related gene expression)in the combined exposure groups of mulch film-derived microplastics and atrazine did not show a uniform trend compared with those in the single exposure group;in addition,the IBR values(oxidative stress)and OTM values in the combined exposure group of PE-aged MPs and 2.0 mg/kg atrazine were higher than those in the combined exposure group of PE MPs and 2.0 mg/kg atrazine,indicating that the combined exposure to PE-aged MPs and 2.0 mg/kg atrazine had a stronger toxic effect on earthworm than the combined exposure to PE MPs and2.0 mg/kg atrazine.