Study On The Physiological Response Mechanism Of Pocillopora Damicornis And Symbiodiniaceae To Seawater Acidification And Cu²⁺ Exposure

Scleractinian corals are the framework organisms in the coral reef ecosystem,and the stable symbiosis relationship with Symbiodiniaceae is the survival characteristic of scleractinian corals.However,the survival and reproduction of scleractinian corals are severely threatened by environmental changes,including the acidification of seawater caused by massive discharge of carbon dioxide(CO₂)and excessive Cu²⁺concentration formed by sewage discharge.Previous studies have shown that both seawater acidification and Cu²⁺pollution can affect the symbiotic relationship between corals and Symbiodiniaceae,but only limited studies have focused on the effects of seawater acidification and Cu²⁺combined exposure on the symbiotic physiology of scleractinian corals.In this study,Pocillopora damicornis and Cladocopium goreaui were used as research objects,and their physiological activities were tested after seawater acidification and Cu²⁺exposure.The relevant results will help deepen the understanding of the symbiosis physiology of stony corals.At the same time,it can provide basic data for environmental risk assessment of coral reef ecosystems.The research includes the following two parts:(1)P.damicornis was exposured to acidified seawater and Cu²⁺,and then the density and chlorophyll a+c₂ content of Symbiodiniaceae,as well as key enzyme activities,cellular energy distribution and macrotranscriptome changes of corals and Symbiodiniaceae were detected.Acidification alone did not produce visible coral bleaching,while Cu²⁺exposure alone and compound exposure both resulted in obvious coral bleaching,and the compound exposure group was more serious.Consistent with the bleaching phenomenon,acidification alone induced the density of Symbiodiniaceae.Both Cu²⁺exposure and compound exposure significantly reduced the density of Symbiodiniaceae,and the compound exposure group was even lower.At the same time,acidification alone inhibited the chlorophyll content of Symbiodiniaceae and promoted its glutamine synthetase activity without significantly disturbing the cellular energy distribution of corals and Symbiodiniaceae;Cu²⁺exposure alone reduced symbiont chlorophyll content,and inhibited coral NADPH-glutamate dehydrogenase activity,and significantly increased and decreased the cellular energy distribution of coral and Symbiodiniaceae respectively;Compound exposure reduced symbiont glutamine synthase,glutamate synthetase and coral NADPH-glutamate dehydrogenase activity,and inhibited the cellular energy distribution of Symbiodiniaceae.The analysis of the macrotranscriptome revealed that acidification alone upregulated the CO₂ transport of coral and downregulated the biological processes of glucose metabolism,amino acid and its derivatives metabolism;Cu²⁺exposure alone upregulated oxidative stress,intracellular transport and carbohydrate biosynthesis,but downregulated DNA replication;Compound exposure can upregulate coral cilia movement,catabolism,intracellular transport,carbohydrate synthesis and other biological processes,and downregulate transcripts involved in DNA replication and cell cycle processes.The above results collectively indicate that seawater acidification may relieve the nitrogen limitation of Symbiodiniaceae and induce the proliferation of Symbiodiniaceae by promoting the CO₂ transport capacity of corals.Short-term acidification exposure has little effect on the growth and development of coral;Cu²⁺exposure alone,and compound exposure can inhibit the energy distribution of Symbiodiniaceae and the ammonia assimilation ability of corals,resulting in a significant decrease in the density of Symbiodiniaceae,and ultimately disrupt the DNA replication and growth and other life processes of coral.Compound exposure can also significantly inhibit the ammonia assimilation ability of Symbiodiniaceae,and downregulate the coral cell cycle,which in turn has a more serious impact on the growth of corals and the proliferation of Symbiodiniaceae.Moremore,P.damicornis may increase their heterotrophic capacity by enhancing ciliary movement,and upregulate intracellular transport,catabolism and carbohydrate biosynthesis to cope with the negative effects of compound exposure.(2)The cultured C.goreaui was exposured to acidified seawater and Cu²⁺for 7days,and then the growth,photosynthesis,organic carbon and nitrogen content,carbon and nitrogen stable isotope content,and metabolome changes were measured.Acidification treatment significantly increased the cell density at the end of the exposure,singular copper treatment and compound treatment significantly depressed the growth of C.goreaui.Furthermore,acidification treatment promoted the maximum photosynthetic efficiency and nutrient accumulation of algae cells,while reducing chlorophyll content;Singular copper treatment inhibited the maximum photosynthetic efficiency and C/N ratio of C.goreaui;and copper/acidification dual treatment caused significant increases in the maximum photosynthetic efficiency,chlorophyll content,and nutrient accumulation.Further analysis revealed that the metabolites in the citrate cycle,glyoxylate and dicarboxylate metabolism,ABC transporter,and nitrogen metabolism pathway increased in the acidification treatment;However,copper treatment-induced metabolites were limited and not enriched in any metabolic pathway;but the acidification/copper dual treatment elicited differential metabolites enriched metabolic pathways such as pentose phosphate pathway,citrate cycle,and biosynthesis of unsaturated fatty acids.In the comparison between dual exposure or and singular acidification treatments,aromatic amino acid in amino acid/secondary metabolite biosynthesis were downregulated.These results suggest that ocean acidification alone can increase the Fv/Fm and nutrient assimilation of C.goreaui,and at the same time increase the citric acid cycle and biomolecule synthesis pathways to promote the growth of algae cells;Cu²⁺exposure alone can inhibite the growth of algae cells by reducing the Fv/Fm and C/N ratio of C.goreaui;Compound exposure can induce the pentose phosphate pathway of C.goreaui.C.goreaui can increase energy through this process to resist the negative effects of Cu²⁺,and downregulation of aromatic amino acid biosynthesis may inhibit the growth of C.goreaui.Taken together,this study shows that both seawater acidification and Cu²⁺exposure could disturb the growth of Symbiodiniaceae,thereby destroying the symbiotic homeostasis between P.damicornis and Symbiodiniaceae;After seawater acidification and Cu²⁺compound exposure,the nutrient absorption and assimilation of Symbiodiniaceae cultured in vitro was sufficient,while the nutrient absorption and energy distribution decreased of Symbiodiniaceae in P.damicornis,indicating that the compound exposure may promote the transfer of nutrients of Symbiodiniaceae to P.damicornis to maintain energy distribution in coral cells;Most of the effects of seawater acidification and Cu²⁺exposure on P.damicornis and Symbiodiniaceae were antagonistic,suggesting that in future ocean acidification scenarios,the effects of metal toxicity on corals may not be as previously predicted based on the low pH metal speciation model serious.