Accumulation And Depuration Of Cadmium And Its Toxic Effect On Litopenaeus Vannamei

Litopenaeus vannamei is the highest production shrimp in China.This species is commonly used as seawater target crustacean to study the physiological responses to environmental contaminants.The study of the ecotoxicological effects of heavy metals on marine organisms can provide biological basis for the risk assessment of metal pollutants and aquatic safety.Cadmium(Cd)is a toxic,carcinogenic,undegradable heavy metal pollutant,which is a great threat to aquatic animals.This study is carried on the acute toxicity experiment for L.vannamei,from the transcriptome and metabolite levels,to explore Cd toxic effects on shrimp.It provides the theoretical support for the safety production of L.vannamei,new ideas and research methods for exploring the mechanism of toxicity of Cd stress on aquatic organisms.The main results are as follows:1.In this study,the 24 h LC₅₀(50%lethal concentration)of L.vannamei(11.00±0.53 cm)was 10.5 mg/L Cd Cl₂·2.5H₂O(Cd²⁺concentration was 5.174±0.019 mg/L).Compared to the control group,ROS production significantly increased at 3-48 h(P<0.01)and peaked at 24 h.The apoptosis rate peaked to 87.99%at 3 h(P<0.01)and significantly increased at 12-48 h(P<0.05).The relative expression levels of Cu-Zn SOD and HSP60 observably increased at 3-48 h and 12-48 h,respectively(P<0.05).GPx expression was significantly upregulated at 12 h and 48 h(P<0.05).The expression profiles of CAT,MT and HSP60 significantly decreased at 3 h(P<0.05),while CAT increased significantly at 12 h(P<0.05),and MT and HSP60 increased at 24-48 h(P<0.05).The expression of HSP70 showed no significant differences at 3-24 h(P>0.05),whereas it increased significantly at 48 h(P<0.05).caspase-3 expression increased significantly at 12-24 h(P<0.05)but was not different at 3 and 48 h(P>0.05).This research showed that Cd induced a large amount of ROS production in hemocytes of L.vannamei and affected the gene expressions of various antioxidant enzymes.With the extension of exposed time,the antioxidant enzyme system could not remove excess ROS,and eventually induced apoptosis.Cu-Zn SOD and HSP60 were sensitive to Cd,while GPx,MT and HSP70 might play a protective role in the later stage of Cd exposure.2.The accumulation of Cd in gills and hepatopancreas increased in the accumulated time,but the muscle showed irregular changes.The highest concentration of Cd was found in hepatopancreas(2418.00±51.07 ng/g),followed by gills(1697.39±17.38 ng/g),and muscle tissue(159.42±1.70 ng/g)after 48 h exposure time.After 48 h of 5.25mg/L Cd Cl₂·2.5H₂O exposure(¹/₂ 24 h LC₅₀,Cd²⁺concentration was 2.858±0.014 mg/L),shrimp was transferred to clean seawater,the depuration of Cd in gills firstly increased in 1 and 5 d,and then decreased in 15 d.Cd concentration in hepatopancreas rapidly decreased to 837-985 ng/g in 1 and 15 d,indicated shrimp?s ability to depurate heavy metals.During the depuration time of 1-15 d,Cd concentration in muscle was significantly lower than that accumulated time 48 h,whereas depurated slowly.Therefore,the accumulation and depuration of Cd in gills,muscle and hepatopancreas were tissue-specific.3.Illumina Hi Seq sequencing platform was used to analyze the changes of hepatopancreas transcription in L.vannamei after Cd exposure,and bioinformatics analysis was used to identify the differentially expressed genes to explore the immune response and detoxification metabolic pathways in response to heavy metal stress.This study has found 2615 differentially expressed genes,including 1461 down-regulated genes and 1154 up-regulated genes.KEGG analysis showed that the differential genes were mainly related to the pathways of immunity,heavy metal detoxification and apoptosis.Cd inhibited the expression of lysozyme and Rab7 gene in order to affect the non-specific immune response of L.vannamei.MT gene may regulate p53 expression.4.The results of metabonomics analysis showed that Cd significantly up-regulated11 compounds,including lysopc 18:0,lysopc 17:0 and isonicotinic acid.30 compounds including D-Glucose 6-Phosphate,Ribulose-5-phosphate,glycine and methylcysteine were significantly down-regulated.Combined analysis of transcriptome and metabolome data showed that Cd significantly affected phosphonate and phosphinate metabolism,arachidonic acid metabolism,glycerolipid metabolism,drug metabolism-other enzymes,cholesterol metabolism and glutathione metabolism.Cd inhibited the process of pentose phosphate pathway,to reduce energy synthesis.Cd might induce oxidative stress and apoptosis by inducing the increase of lysophosphatidylcholine.By inhibiting arachidonic acid metabolism,Cd inhibited the immune and antimicrobial function in shrimp.Cd inhibited glutathione synthesis and detoxification by inhibiting glycine synthesis.