Toxicological Effects Of Nanoplastic And Nano-TiO₂ On Litopenaeus Vannamei Through Dietary Exposure

The development of nanotechnology will inevitably lead to the release of a certain amount of nano materials into the marine ecosystem.Nanoplastics and titanium dioxide nanoparticle(nano-TiO₂)are two kinds of nano materials that are widely used and ubiquitous in the environment.Therefore,the ecotoxicological effect caused by nanoparticle pollution is the current research hotspot in the field of environmental toxicology.It is a kind of nano metal oxide particles with the most extensive research in the field of ecotoxicity.In this study,polystyrene(PS)nanoplastics and nano-TiO₂were used as the research objects,Litopenaeus vannamei was selected as the experimental organism,to evaluate the distribution of nano-PS and nano-TiO₂ in L.vannamei through dietary exposure.In addition,the effects of nano-PS plastics and nano-TiO₂ single exposure on the growth,immunity,oxidative stress,histological structure of intestine and hepatopancreas and intestinal microbiota was also analyzed.The results can provide technical support for the ecological risk assessment of nanoparticles.The main experimental results are as follows:(1)The abundances of nano-PS and nano-TiO₂ in intestine and muscle of L.vannamei showed an increasing trend after exposed to the diets spiked with nano-PS and nano-TiO₂.During the depuration process,the concentrations of nano-PS decreased exponentially.However,there was still a large amount of nano-TiO₂ in the tissues after5 days of depuration.Compared with nano-PS,nano-TiO₂ is irregular in shape and easier to agglomerate,so it is more difficult to be excreted by L.vannamei.(2)The exposure of two kinds of nano materials had a certain inhibitory effect on the growth of L.vannamei.The results showed that the weight gain rate and specific growth rate of L.vannamei decreased after exposure of nano-PS and nano-TiO₂.Compared with the control group,the weight gain rate and specific growth rate of high concentration of nano-PS treatment group decreased significantly.(3)The nano-PS and nano-TiO₂ exposure had caused the oxidative stress and affected the immune system of L.vannamei.The results showed that both nano-PS and nano-TiO₂ caused a significant increase in the apoptosis rate of hemocytes.Besides,nano-PS and nano-TiO₂ led to different degrees of increase in the activities of T-AOC,SOD and GST in hepatopancreas,while the significant decrease of Na⁺/K⁺-ATPase activity was observed in the treatment groups.(4)The hepatopancreas and intestine of L.vannamei showed concentration dependent histological changes.Both nano-PS and nano-TiO₂ caused expansion of the stellate tubules of the hepatopancreas tissue,and partial cavitation appeared and ruptured.Compared with the control group,the intestinal mucosa was slightly loose,and the number of epithelial cells in the mucosa was less.Besides,proliferative cells appeared after nano-PS exposure.(5)Exposure to nano-PS and nano-TiO₂ changed the expression levels of immune-related genes in L.vannamei.After 14 days of exposure,the gene expressions of Beta-glucan binding protein,LPS/?-glucan binding protein,HSP 70,TLR and Toll 1 were significantly up-regulated in the high concentration nano-TiO₂ treatment group.The expression levels of Beta-glucan binding protein,LPS/?-glucan binding protein,HSP90 and TLR genes were significantly increased after exposure to high concentration of nano-PS.After 28 days of exposure,high-concentration nano-TiO₂ exposure resulted in a significant increase in the expression of HSP 90 and Toll 1 and a significant decrease in the expression of LPS/?-glucan binding protein.Nano-PS significant up-regulated the expression of TLR and significant down-regulation the expression of Toll1.(6)After exposure to nano-PS and nano-TiO₂,significant alterations of intestinal microbiota were found.The abundance and diversity of intestinal microbiota were reduced.Moreover,the abundance of Proteobacteria and Actinobacteria were significantly increased,while the abundance of Bacteroidetes were significantly reduced.Compared with the control group,the proportion of Vibrio,Tenacibaculum,Microbacterium,Bradyrhizobium and Demequina in the treatment group increased in varying degrees.The function of the intestinal microbiota of L.vannamei was predicted by Tax4Fun analysis,and the results showed that the exposure of the two nano materials led to the pathways of"metabolism","environmental information processing","cellular processes"and"organismal systems"were significantly different compared with the control group.These indicated that the two types nanoparticles caused the variation of intestinal microbiota and may affect the metabolic function of intestinal microbiota.(7)There were certain differences in the toxic effects of the two types nano materials on L.vannamei.Among them,the main effects of nano-PS exposure on L.vannamei were:induction of proliferative cells in the intestinal tissues,disturbing the homeostasis of the intestinal flora,and reducing the weight gain rate and specific growth rate of L.vannamei.Compared with nano-PS exposure group,nano-TiO₂exposure significantly increased the activity of GST in hepatopancreas of L.vannamei,and significantly inhibited the activity of Na⁺/K⁺-ATPase.Integrated biomarker response(IBR)index value showed that the oxidative stress,energy metabolism and blood cell apoptosis rate of L.vannamei exposed to nano-TiO₂ were higher than those exposed to nano-PS.