Preliminary Study On The Toxic Effect Of Microplastics On Eriocheir Sinensis

Plastic contamination is an increasing environmental problem in marine systems,which poses a great threat to the entire ecosystem and has attracted the attention of more and more researchers.In this study,polystyrene microspheres and E.sinensis were selected as the experimental material and organism,respectively,to evaluate the effects of microplastics on the growth,uptake,and accumulation of microplastics along with effects of microplastics on oxidative stress,humoral immune factors and intestinal microflora.The findings provide basic biological data for follow-up studies on the toxic effects of new pollutant microplastics on aquatic organisms.The main experimental results are as follows:1.Accumulation of microplastics in tissues of Eriocheir sinensis and its effect on growth and ATPase activity in gill tissuesIn this study,fluorescent microplastic particles(diameter = 0.5 ?m)with a concentration of 40mg/L and the live and healthy female crabs were used as experimental materials for 7d accumulation experiment.The results demonstrate that fluorescent microplastic particles accumulated in the gill,liver and gut tissues of E.sinensis after seven days of exposure.A 21-day toxicity test suggested that the rate of weight gain,specific growth rate,and hepatosomatic index of E.sinensis decreased with increasing microplastic concentration(0,0.04,0.4,4 and 40mg/L).The ATPase activity in the gill tissue increased after 7 and 14 days microplastic exposure,while at 21 days,the high concentration microplastic exposure group significantly inhibited the ATPase activity in the gill tissue.The above results indicate that microplastics can accumulated in the gill,liver and gut tissues of E.sinensis,and microplastics have a negative impact on the growth index and the activity of ATPase in gill tissues.2.Effects of microplastics on oxidative stress in hepatopancreas of Eriocheir sinensisThe effects of microplastics on the hepatopancreas of Eriocheir sinensis were evaluted by examining the activity of a series of enzymes and their gene expression in hepatopancreas.These include the activity of alanine aminotransferase(GPT)and aspartate aminotransferase(GOT),which reflect hepatopancreatic tissue damage,acetylcholinesterase(ACh E)activity in neurotoxicity,and oxidative stress-related parameters: superoxide dismutase(SOD),catalase(CAT),glutathione peroxidase(GPx),glutathione transferase(GST),glutathione(GSH),malondialdehyde(MDA),and lipid peroxide(LPO).In addition,the relative m RNA expression levels of SOD,CAT,GPx,GST and several important genes in the MAPK pathway closely related to oxidative stress were further examined from a molecular perspective.The results showed that the activities of SOD,GOT,GSH and GPx increased first and then decreased.The activities of ACh E,CAT and GPT were significantly lower than those of the control group.The contents of MDA and LPO increased significantly compared with the control group.Upon exposure to increasing microplastic concentrations,the expressions of the antioxidant SOD,CAT,GPx and glutathione S-transferase genes in the liver decreased after first increasing.Exposure to microplastics increased the expression of p38 gene in the MAPK signaling pathway and significantly decreased the expressions of ERK,AKT,and MEK genes.The results of this study demonstrate that exposure to microplastics can induce oxidative stress in the liver of E.sinensis.3.Effects of microplastic exposure on non-specific immune responses and intestinal microflora of juvenile Eriocheir sinensisThe effects on humoral immune factors were evaluated by analyzing the following immune parameters in the hemolymph and hepatopancreas: ceruloplasmin(CP)content,activity of alkaline phosphatase(AKP),acid phosphatase(ACP),lysozyme(LSZ),and phenoloxidase(PO)as well as expression of six immune-related genes;Hc,ACP,AKP,LSZ,caspase and myeloid differentiation factor 88(My D88).In addition,effects on intestinal microflora were evaluated based on the diversity and composition of the intestinal community.The results showed that under microplastic-induced stress,the activities of most immune-related factors declined after an initial rise.The trends in Hc,ACP,AKP,and LSZ gene expression was consistent with the corresponding changes in enzyme activity.Moreover,hemocyte expression of caspase and My D88 under microplastic-induced stress was higher than that in the Control group,while the expression levels in hepatopancreas were first increased and then decreased.Furthermore,Saccharibacteria,Firmicutes,Actinobacteria,Tenericutes,Bacteroidetes,and Proteobacteria were dominant in the intestines of E.sinensis regardless of the presence or absence of microplastic.However,the relative abundance of Firmicutes and Bacteroidetes decreased following exposure to microplastic at 40mg/L,while that of Fusobacteria and Proteobacteria increased.The above results indicate that microplastics have a significant negative impact on immune-related enzymes and genes in the hepatopancreas and hemolymph of the Eriocheir sinensis.It is speculated that these enzymes and genes may be part of important defense mechanisms in the Eriocheir sinensis to combat inflammatory reaction caused by microplastic.In addition,microplastics have an impact on the diversity and composition of the intestinal flora of Eriocheir sinensis at several levels of phylum,genus and genus.