| Phytoremediation is an efficient and environmentally friendly solution to heavy metal pollution.Woody plants such as willow and poplar grow rapidly,have large biomass and deep roots,and have good potential to remediation environmental pollution.However,there are still few studies have been devoted to the tolerance of woody plants to compound heavy metal pollution and their tolerance mechanism.In view of this,Cd and Zn,which have similar chemical properties,are selected as experimental element(Zn is an essential element for plant growth,while Cd is an unnecessary element for plant growth);In this study,Salix matsudana Koidz and S.fragilis L.are selected as experimental materials to investigate the tolerance and mechanism of the two species to Cd-Zn compound pollution by atomic absorption spectrometry,chemical continuous fractionation extraction,differential centrifugation and enzyme activity analysis and other methods of physiological and biochemical indexes.Meanwhile,the mechanism of plant tolerance to different heavy metals was revealed by comparing the differences in the accumulation rules and existing forms of Cd and Zn in S.fragilis tissues.The main results were summarized as follows:1.Under hydroponic conditions,different concentrations of complex heavy metals(low stress concentration Cd(25μmol/L),high stress concentration Cd(50μmol/L),low stress concentration Zn(50μmol/L),high stress concentration Zn(100μmol/L)were used in pairs,namely CK、Cd25+Zn50、Cd50+Zn50、Cd25+Zn100、Cd50+Zn100)were used to conduct stress experiments on S.matsudana and S.fragilis.The results showed that S.matsudana has stronger tolerance to Cd-Zn combined stress than S.fragilis.Under the same concentration stress,all the growth indexes(fresh weight of shoot,dry weight of shoot,longest stem length,fresh weight of root,dry weight of root)of S.fragilis except for the longest root length showed a more obvious decreasing trend than that of S.matsudana,indicating that the growth of S.fragilis was more seriously inhibited.However,the reduction of root length of S.fragilis under high concentration of complex heavy metal stress was significantly less than that of S.matsudana,suggesting that reducing root length is beneficial to reduce the contact between root and metal,which is a strategy of S.matsudana to cope with heavy metal stress.The results also showed that the tolerance index of S.matsudana to different concentrations of CdZn compound stress was significantly higher than that of S.fragilis.2.The mechanism of strong heavy metal tolerance of S.matsudana was revealed from three aspects: immobilization of heavy metal ions,complexation of cellular ligands,and protection of antioxidant defense system.Compared with S.fragilis,S.matsudana retained more heavy metals in the roots to protect the aboveground organelles from damage.Cd and Zn showed antagonistic effects in the shoots and roots of the two willows.However,the increase of Cd stress concentration significantly increased the content of Zn in chloroplasts of S.matsudana,in order to alleviate the damage of chloroplast caused by Cd stress.Both S.matsudana and S.fragilis fixed a large amount of heavy metals in the cell wall to protect the protoplasts from heavy metal damage,and the cell wall of S.matsudana had a stronger ability to immobilize heavy metals,especially under high concentration of heavy metal stress.There was no significant difference in the compartmentalization ability of soluble fraction to heavy metals between the two species.However,the proportions of heavy metals in mitochondria,plastids and nucleus of S.matsudana were lower than those in S.fragilis.Compared with S.fragilis,more heavy metal ions in S.matsudana cells combined with cell ligands such as pectin and protein to transform into heavy metal forms with low activity and low mobility,especially under high concentration of heavy metal stress.With the increase of Zn and Cd concentrations,the content of malondialdehyde(MDA)in the shoots and roots of both species increased,indicating that the cell membranes of both species were damaged to varying degrees.The superoxide dismutase(SOD)activity of S.matsudana increased with the increase of heavy metal stress concentration,and the SOD activity of S.fragilis increased first and then decreased.Meanwhile,the SOD activity of S.matsudana was significantly higher than that of S.fragilis under high concentration of complex heavy metal stress.The ability of glutathione(GSH)synthesis of S.matsudana was significantly stronger than that of S.fragilis under heavy metal stress.3.Low stress concentration Cd(25μmol/L),high stress concentration Cd(50μmol/L),low stress concentration Zn(50μmol/L)and high stress concentration Zn(100μmol/L)were used to treat S.fragilis separately(CK,Cd25,Cd50,Zn50,Zn100),to study the difference of tolerance mechanism of S.fragilis to Zn and Cd.The results showed that low or high concentration of Cd stress significantly inhibited the growth indexes(shoot fresh weight,shoot dry weight,longest stem length,root fresh weight,root dry weight,longest root length)of S.fragilis.Low concentration Zn stress significantly inhibited the fresh weight of shoot,dry weight of shoot and longest root length of S.fragilis,while high concentration Zn stress significantly inhibited all growth indicators.There are differences in the effects of Cd and Zn on photosynthetic pigments of S.fragilis.Both of Cd and Zn could significantly reduce the content of photosynthetic pigments and the ratio of chlorophyll a to chlorophyll b.However,Zn stress had the most significant effect on the content of chlorophyll a,while Cd stress had the most significant effect on the content of carotenoid,followed by chlorophyll a and chlorophyll b.4.The tolerance mechanism of S.fragilis to Cd and Zn was investigated from the perspectives of the subcellular distribution and chemical forms of Zn and Cd.The results showed that the cell wall and soluble fraction were the main parts of the fixation of Cd,while more Zn entered the cells and was compartmentalized in the soluble fraction.Cd and Zn also showed different accumulation patterns in sensitive organelles,with Cd preferentially accumulating in chloroplasts rather than mitochondria,while Zn was almost evenly distributed in the two organelles.The results also showed that Cd and Zn have different interactions with different cell ligands.The ligands such as pectin and protein play an important role in the process of S.fragilis alleviating Cd stress,while Zn mainly combined with phosphate and oxalate to reduce mobility and activity.5.The differences in tolerance mechanism of S.fragilis to Zn and Cd were investigated from the perspective of response of antioxidant defense system.The results showed that MDA content was significantly higher under high concentration of Cd than that under Zn stress,indicating that lipid peroxidation induced by Cd was stronger than that induced by Zn.The activities of SOD and GSH increased significantly under Zn stress,indicating that these two substances played an important role in relieving the physiological stress induced by Zn.Under low concentration of Cd stress,the activities of SOD and GSH in shoots and roots increased,but with the increase of Cd stress concentration,SOD activity decreased,and root GSH content also slightly decreased,indicating that high concentration of Cd stress weakened the role of S.fragilis free radical clear system.In conclusion,this study found that the tolerance of S.matsudana to Cd-Zn combined stress was significantly stronger than that of S.fragilis.Compared with S.fragilis,S.matsudana can transform heavy metal ions into low active forms by combining with cell ligands such as pectin,and store them in the roots or cell wall or other metabolically inactive parts.At the same time,S.matsudana can enhance its tolerance to Cd-Zn complex stress through its strong active oxygen scavenging function of antioxidant defense system.The growth of S.fragilis was significantly inhibited under the stress of Cd or Zn at certain concentration.The non-essential element Cd was mainly enriched in chloroplasts to reduce the content of carotenoids.The tolerance of S.fragilis to Cd was mainly through binding with pectin or protein and stored in cell wall and soluble fraction;while the essential element Zn mainly reduces the chlorophyll a content,the tolerance of S.fragilis to Zn was mainly through binding with phosphate or oxalate and compartmentalize in soluble components. |
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