Chronic Zn²⁺ And Triclosan Exposure Induces Growth And Developmental Toxicity In Bufo.gargarizans Tadpoles

With the standard of living and industrial and agricultural developments,a large number of living and working production waste emissions into the water environments.Different kinds of chemical pollutants in water environments caused scale ecological deterioration,and now have become the focus of attention in the world.As an broad-spectrum antimicrobial agent,triclosan is commonly used worldwide in a range of personal care and sanitizing products,as well as household items,such as soap,detergents,deodorant,toothpaste,and kitchen utensils.,The majority of consumer products containing TCS are eventually rinsed down the drain and discharged with wastewater effluent.Consequetly,the presence of TCS has been detected frequently in all types of aquatic environments,including lakes,rivers,coastal,and estuarine waters.In addition,with the development of industral,a mass of zinc was produced by human activities,such as zinc ore mining,smelting and processing,machinery manufacturing,galvanizing,organic synthesis and other industrial,which caused the increase of the zinc concentration in the aquatic environments.Different kinds of chemical pollutants in water environments will threat the survive of non-target organisms and ecosystem structure,A number of studies have reported the adverse effects of TCS on aquatic organisms,such as fish,mussel,and amphibian.In amphibian species,studies have shown that TCS at sublethal concentrations could reduce survival rate and body weight of tadpoles.Also,TCS was shown to affect the thyroid hormone?TH?signaling pathway and disrupt developmental processes of tadpoles.In addition,excessive Zn²⁺ in aquatic environments can cause toxic effects on organisms.For example,excessive Zn²⁺ caused adverse effects on respiration rate and hatching rate in fish.Unfortunately,to date,little information is available regarding the adverse effects of TCS on amphibian embryos,size and bone development of metamorphic tadpoles,and chronic effects of Zn²⁺ exposure on the growth and development in Bufo gargarizans tadpoles.In this paper,we studied the toxic effects of chronic TCS exposure on B.gargarizans embryos and larvae.Also,the effects of sublethal Zn²⁺ exposure on larval growth and development in B.gargarizans were studied.The results of this study can provide valuable information about the toxicity and safe use of TCS or Zn²⁺ that have the potential to enter aquatic systems.The main results are summarized as follows:1.To evaluate potential effects of TCS exposure on embryonic development of Bufo gargarizans,standard Gosner stage 3 B.gargarizans embryos were exposed to different concentrations of TCS(0?10?30?60?150 ?g·L^-1)during embryogenesis.Survival,total length,body weight,developmental stage,duration of different embryo stages,and malformation were measured.The relative mRNA expression and spatial expression patterns of type II and III deiodinase?D2 and D3?in B.gargarizans embryos were also examined by semi-quantitative RT–PCR and situ hybridization,respectively.Our results showed that inhibitory effects on embryo developmental stage,total length and body weight were found at 30-150?g·L^-1TCS.Moreover,the duration of embryonic development was increased at gastrula,neural,circulation and operculum development stage in TCS-treated groups.In addition,TCS exposure induced morphological malformations in B.gargarizans embryos,which are characterized by hyperplasia,abdominal edema,and axial flexures.Furthermore,semi-quantitative RT–PCR results showed that the expression of D2 in embryos was down-regulated at 60 and 150?g·L^-1.However,the spatial expression patterns of D2 and D3 in embryos was not affected by TCS.2.To examine the effects of chronic exposure to TCS on growth and development,and metamorphosis in Bufo gargarizans.B.gargarizans tadpoles were exposed to waterborne TCS at different concentrations ranging from 10-150 ?g L^-1 from Gosner stage 26 to completion of metamorphosis?G46?.On day 40 and 55 of treatment,total length,body length,body weight,and developmental stage of tadpoles were determined.G46 froglets were measured for body length,body weight,and hind limb length.The skeleton development of G46 froglets was also analyzed by the double-staining method.On day 40,significant increase in total length,body weight,and developmental stage relative to the controls were observed at 150 ?g·L^-1 TCS.On day 55,increased body length was only observed at 150 ?g·L^-1 TCS.In addition,froglet size at completion of metamorphosis was slightly smaller at 60 and 150?g·L^-1 TCS relative to the controls.Moreover,significant reduction in tadpole at G42 deposition of calcium were observed at 60 and 150 ?g·L^-1TCS.3.To evaluate the toxic effects of zinc ion(Zn²⁺)on the larvae of B.gargarizans,tadpoles were exposed to zinc(0,10,50,100 and 500 ?g·L^-1 Zn²⁺)from the beginning of larval period?G26?through metamorphic climax?G42?.Firstly,total length,body length,body weight,and developmental stage of tadpoles were determined on day 15 and 30 after Zn²⁺ exposure.Secondly,metamorphosis and body size?total length,body length,body weight,and hind limb length?at metamorphic climax?G42?were examined.Finally,skeletal systems of tadpoles at G42 were investigated by using double staining methodology.Our results showed that Zn²⁺ at 500 ?g·L^-1 could inhibit tadpole growth and development on day 30 after Zn²⁺ exposure.In addition,there was a significant decline in percentage of tadpoles reaching metamorphosis climax,especially for larvae exposed to 500 ?g·L^-1 Zn²⁺.Moreover,significant reduction in tadpole size?total length,body length,and hind limb length?at G42 and deposition of calcium were observed at 500 ?g·L^-1 Zn²⁺.