| The carbon dioxide generated by the burning of fossil fuels continuously released into the atmosphere.About 1/4～1/3 of the carbon dioxide will be absorbed by the oceans,which led to the decrease of the pH value of surface water,forming the so-called ocean acidification,and causing the serious impact on the Marine system.There are reports that ocean acidification can affect the growth and development,gene expression and metabolic process of marine calcareous organisms.But the researches about the effects of ocean acidification on gene expression and metabolic process of abalone are still few.In addition,many studies have shown that environmental stress will change the biological state of DNA methylation,thus affect the gene expression and adjust the survival and development state of organisms.But the related researches are limited in the marine calcareous organisms.By simulating the future marine environment(control group:400ppm,medium acidification groups:800ppm and high acidification groups:1500ppm),our experiments studied the effects of the short-term and long-term acidification on the growth and survival of Haliotis discus hanai and Halotis diversicolor,and explored the effects on molecular function of abalone from the level of DNA methylation,gene expression and metabolome.The main results were as follows:1,Effects of ocean acidification on the growth and survival of adult Haliotis discus hanai and Halotis diversicolor.The short-term(the time of stress within 2W)moderate acidification could promote the growth of Haliotis discus hanai and Halotis diversicolor,while the short-term high acidification only could promote the growth of Haliotis discus hanai,and inhibited the growth of Halotis diversicolor.The long-term(the time of stress above 4M)moderate and high acidification would inhibit the growth of Haliotis discus hanai and Halotis diversicolor.The short-term acidification would not cause the death of Haliotis discus hanai,but obviously increased the mortality rate of Halotis diversicolor.The long-term high acidification would obviously increase the mortality rate of adult Haliotis discus hanai,while the moderate acidification would not affect the survival of Haliotis discus hanai.But both of the long-term moderate and high acidification could significantly increase the mortality rate of Halotis diversicolor.2,Effects on the larval development when abalone parents cultured in the long-term acidification(10.5 M).In comparison with the larval development of acidified water and normal water,it was found that:(1)In terms of fertilization rate and deformity rate,regardless of whether the parent was treated by acidification in advance,the high acidification could lead to the decrease of fertilization rate and the increase of deformity rate.(2)In the hatching rate,when the parents weren’t treated by the acidification in advance,the acidification would reduce the hatching rate of Haliotis discus hanai;but when one of the parents was treated by the acidification,the hatching rate of acidification group would rise.(3)In the case of the mortality rate and shell length of the veliger,when the male parent was treated with long-term acidification while the female parent was not treated by acidification,the larvae mortality was reduced and shell length was increased.It would benefit the survival and growth of the larvae under the acidification environment.But when the female parent was also treated with long-term acidification,the beneficial effects of the male parent treated by acidification in advance would disappear.Comparing the larval development under conditions that the parents not treated by acidification to ones that the parents treated by acidification in advance,it showed that:(1)In terms of fertilization rate,comparing with the parents not treated by acidification,parents treated by the long-term high acidification could increase the fertility rate,while the parents treated by the long-term moderate acidification would decrease the fertility rate.(2)For the hatching rate,when the female(male)parent had not been subjected to acidification,the male(female)parent treated by long-term acidification in advance would raise the larvae hatching rate in acidified water(compared with male(female)parent had not been subjected to acidification),which was in contrast with the situation in the normal water.(3)For the deformity rate,mortality rate and veliger shell length,when the female parent was not treated by acidification,the male parent treated by acidification in advance would decrease the larvae deformity rate and mortality rate of Haliotis discus hanai,and increase the veliger shell length(compared with male parent had not been treated by acidification),which was in contrast with situation in the normal water.When the female parent is also treated by acidification in advance,the advantages by the male parent treated by acidification on the growth and development will be weakened.In addition,regardless of whether the male parent was treated by acidification in advance,the female parent treated by acidification in advance would increase the deformity rate and mortality rate,and decrease the veliger shell length in the normal water and acidified water(compared with female parent had not been treated by acidification).3,Effects of marine acidification on DNA methylation of Haliotis discus hannai’s and Haliotis diversicolor’s mantle.The total DNA methylation rate of Haliotis discus hannai’s and Haliotis diversicolor’s mantle was 22.76%～32.46%and 50.32%～57,72%,respectively.The external semi-methylation rates were in the range of 3,40%～7.96%and 12.37%～21.06%,and the internal full-methylation rates were within the range of 18.40%～24.52%and 34.22%～41.83%.The total methylation rate,external semi-methylation rate and internal full-methylation rate of the mantle of Haliotis diversicolor were higher than that of Haliotis discus hannai.But both of them showed that the internal full-methylation rate(CG type)was higher than the external semi-methylation rate(CHG type).For Haliotis discus hannai,there was no significant difference in the total methylation rate between the acidification group and the control group at 24 h,1 W and 1 Y,but all showed that the acidification group was lower than the control group.After stressed for 3M,the total DNA methylation rate in high acidification group was significantly lower than that in control group and moderate acidification group,while there was no significant difference between moderate acidification group and control group.And this significant difference was caused by the significant reduction of internal total methylation rate.However,for Haliotis diversicolor,there was no significant difference in the total methylation rate between the acidification group and the control group at all measured stress points.It showed that the total methylation rate in the acidification group was higher than that in the control group.4,Effects of marine acidification on expression of shell formation related genes in Haliotis discus hannai.In this study,the expression of 11 shell formation related genes was analyzed.Among them,HasSom,TYR and PRX genes could promote the dye of the cuticle,and the expression of HasSom gene in the short-term acidification was significantly increased,while the expression of TYR and PRX genes in the long-term acidification had a trend of increasing,but not reaching a significant level.EPP gene has the function of assisting HasSom gene to participate in the dye of the cuticle.It showed the compensation effect to the HasSom gene when the acidification was less than 1Y,but the compensation effect would disappear when the stress reached 1Y.CaM gene was significantly decreased in short-term acidification,showing the compensation effect on calcification process,while the expression of CaM gene was increased in long-term stress,which could promote the repair of cuticle,but the promoting effect would disappear when the stress reached 1Y.The expression of CA gene showed a trend of decreasing in high acidification stress,which was helpful to alleviate the decrease of pH in the abalone body and beneficial to calcification.NACR gene has the function of shell repair,and acidification has the tendency to enhance its expression.It could play a more rapid repairing effect under moderate acidification conditions.CHI and Pif177 genes had the effect of alleviating acidification on the damage of nacre layer.CHI gene only played a mitigating effect in the moderate acidification,and Pif177 gene played a strong remission in short-term acidification and decreased with prolonged stress time.PLUC and PLUS genes had the effect of promoting the precipitation of aragonite crystal,which could repair the nacre layer.PLUC gene played a strong repair effect in the long-term acidification.In the moderate acidification PLUS gene could play a repair effect in the short time,while in the high acidification,it needed a long-term stress to play a repair role.5,Metabolomics analysis for effects of long-term ocean acidification on Haliotis discus hannai.The OPLS-DA and cluster analysis showed that the control group and the high acidification group were significantly differentiated.The 628 metabolites were detected by GC-MS and 549 metabolites were retained by data filtering.Finally,10 significantly different metabolites were screened,and the expression was up-regulated in high acidification group.The content of fructose-2,threonic acid and 4-hydroxyphenylacetic acid was more than 20,000 times higher than that of the control group.Metabolic pathway analysis revealed that biotin,6-glucose phosphate-1,a-aminoadipic acid and threonic acid were associated with energy metabolism,and fructose-2 was associated with protein and nucleic acid synthesis.Indole-3-acetic acid and 4-hydroxyphenylacetic acid were related to the synthesis of cuticle pigment,indicating that acidification could affect the energy metabolism,protein and nucleic acid synthesis and coloring process of horny stratification of Haliotis discus hannai.The above results showed that biotin,6-glucose phosphate-1,a-aminoadipic acid,fructose-2,threonic acid,indole-3-acetic acid and 4-hydroxyphenylacetic acid had the potential ability of being the biomarker of detecting the effects of acidification on Haliotis discus hannai. |
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