| In China,to develop bioenergy,there is a conflict between planting energy plants and food crops.The energy plants are used to cultivate in the soil contaminated with heavy metals,which can not only alleviate the above issue,but also have the effect of remediation the polluted soil.Italian ryegrass(Lolium multiflorum L),an annual crop,can be rotated with rice.This crop has the characteristics of fast growth,wide distribution and high biomass.In recent years,it has been reported that Italian ryegrass can be used to convert bio-ethanol.Thus,this crop is valuable as an energy plant to be developed.In this study,different cadmium(Cd)-tolerant groups were screened during seed germination by using Italian ryegrass cultivars,Cd tolerant and Cd sensitive cultivars were further identified according to the pot experiment.The difference in Cd tolerance and accumulation were tested in soil and Hydroponics conditions by using Cd tolerant and Cd sensitive cultivars.In this study,the physiological responses of the two cultivars were compared under Cd stress with hydroponics conditions.On this basis,in order to promote the rapid growth of Italian ryegrass under Cd stress,the mechanisms of alleviation Cd toxicity in Italian ryegrass were conducted with supplement of exogenous glutathione(GSH).The main results of the study are as follows.2.The gradient Cd treatment(0-1000 μmol L-1)was conducted to compare the the absolute value and relative value of multiple parameters in seed germination.The subordinate function and standard deviation coefficient weight were used to evaluate the Cd tolerance among 21 cultivars treated with excess Cd.The results showed that significant difference in Cd tolerance among 21 Italian ryegrass cultivars were observed under 0-1000μmol L-1 Cd stress(P<0.01).The tolerance to Cd among 21 cultivars can be classified into four different groups:Cd-tolerant,moderate Cd-tolerant,low Cd-tolerant and Cd-sensitive.As a result,Idyll,Jaianto and Hitachihikari were classified Cd tolerance cultivars,while Nioudachi,Wasehope,Doraian,Harukaze and Tachiwase were regarded as Cd sensitive cultivars.When the seeds treated with Cd,the root growth,seed germination potential and seed vigor of the cultivars were inhibited with different degrees,which the effects of weight on cadmium tolerance is higher than other parameters.2.Pot soil experiment was used to further analyzed the characteristics of Cd indexes,Cd uptake and transport in 10 cultivars with different Cd-tolerant at 40 mg kg-1 Cd treatment.The results showed that the highest Cd index was Idyll among different cultivars,and the lowest one was the Harukaze.Significant difference in Cd uptake was observed among different cultivars.According to Person’s correlation analysis(P<0.05),significant positive correlation between Cd tolerance index and Cd accumulation was observed,while a significant negative correlation between translocation factors and Cd concentrations in roots was found,respectively.After identification,Idyll was a cultivar with Cd tolerance and high accumulation,while Harukaze was another one with low accumulation and Cd sensitive.When the Cd concentrations in pot soil were simulated with 0,20,40,and 80 mg kg-1,the differences in Cd tolerance and accumulation between Idyll and Harukaze were cultivated for 90d,the results showed that the total amount of Cd concentration and Cd accumulation in cultivar Idyll was higher than that of Harukaze.After planting these two cultivars,Cd concentration in soil decreased compared with preplanting,while no significant level was exhibited between cultivars(P<0.05).However,the concentration of Cd in the soil showed an decline after planting,and the decrease of Idyll was lower than that of Harukaze,indicating that Idyll can be planted in Cd contaminated soil as the first cultivar,this cultivar also can be used to remediation contaminated soil.3.Two cultivars Idyll(relative Cd-tolerant)and Harukaze(relative Cd-sensitive)were used as materials in hydroponic experiment,the seedlings were grown for 12 days at Cd concentration of 0-100 μmol L-1 to compare the physiological responses between two cultivars.The results showed that Cd treatments significantly affects the concentration and accumulation of Cd in roots and shoots,significant difference in the above indexes were found between two cultivars.Cd concentration in root and shoot of Idyll was lower than that of the corresponding of Harukaze under the same Cd treatment.The translocation factors of Cd in Harukaze showed an increase with Cd supply in the solution.Excess Cd inhibited the growth and photosynthetic pigment content in two cultivars seedlings,and these effects was more pronounced in Harukaze than that of Idyll.When Cd concentration in solution was not less than 25 μmol L-1,the level of membrane lipid peroxidation in roots and leaves of Idyll was lower than those of Harukaze,and the trend of non-protein sulfhydryl content was reversed.Thus,Idyll has a stronger tolerance to Cd than Harukaze.The uptake and translocation of Cd and essential elements(Zn,Fe,Mn,Mg)were different,which depend on the cultivar,Cd concentration and plant tissues.The differences in Cd tolerance and Cd uptake of two cultivars were related to Cd transport efficiency,non-protein sulfhydryl content in tissues and the level of membrane lipid peroxidation.4.Seedlings of Idyll and Harukaze grown in the condition of hydroponics with exogenous application of 200 μmol L-1 GSH(glutathione)under the stress of 25 μmol L-1 and 50 μmol L-1 Cd.In this experiment,the difference in Cd uptake,Cd subcelxular distribution and chemical forms in plant tissues were analyzed,respectively.The results showed that Cd concentration in plant roots were significantly increased by application of GSH,and Cd concentration in root of Harukaze showed higher than that of IdyⅡ.Cd is mainly concentrated the cell wall of subcellular components.Meanwhile,GSH treatment caused the decrease in Cd concentration of shoot,most of Cd was accumulated in soluble fraction by application of GSH.The proportion of cell wall in subcellular fraction was significantly increased with supplement of exogenous GSH.The increase of proportion in Idyll was higher than that of Harukaze.Compared with Cd stress alone,the concentration of 80%ethanol-abstracted Cd in Harukaze root decreased significantly when the seedlings were applied exogenous GSH.At the same time,the Cd concentrations of 2%HAC extraction or 0.6 mol L=1 HCl extraction of the two cultivars were significantly increased,and the significant difference in the Cd form which extracted by 2%HAC or 0.6 M HCl were found between two cultivars.The proportion of the above two Cd forms in total Cd forms increased significantly when the seedling were applied exogenous GSH,and the increment of Harukaze was higher that that of Idyll.The concentration of Cd form in the aerial part of plant was reduced or significantly reduced by application of GSH,while the proportion of Cd extracted by 80%ethanol was still about 58%-76%.Based on this,we concluded that the exogenous addition of GSH significantly increased the ability to fix Cd in root cell wall under Cd stress,and promote the soluble high-toxic Cd form transfer to insoluble and low-toxic form in the root,thus reducing the translocation of Cd from root to shoot.5.Two cultivars Idyll and Harukaze were used in hydroponic experiment,seedlings were treated with 50 μmol L-1 Cd alone or together with 200 μmol L-1 exogenous GSH to investigate the responses of sulfhydryl compounds and Cd uptake.The results showed that the concentrations of total sulfhydryl compounds in root and shoot of Idyll were significantly higher than that of the corresponding Harukaze cultivar under Cd stress.Seedlings were treated for 6 days and 10 days by application of 200 μmol L-1 GSH into solution with 50 μmol L-1 Cd,Cd concentration and GSH contents in root of Idyll and Harukaze were significantly increased,and significant difference was observed between two cultivars.Additionally,GSH treatments resulted in the significant decreases in the synthesis of thiols compound such as PC3 and PC4,and the reductions of Harukaze were higher than the corresponding of cultivar Idyll.The concentration of GSH(PC1)and total thiols were increased significantly in leaves of two cultivars by application of GSH.Meanwhile,Cd concentration in shoot of two cultivars were reduced when GSH were applied.The pretreatment of 150 μmol L-1 BSO greatly inhibited the synthesis sulfhydryl compounds of two cultivars,while the translocation of Cd was significantly increased in Harukaze.The synthesis of large molecule sulfhydryl compounds(PC3 and PC4)were reduced by application GSH,thus,the complexation with Cd2+ were reduced,which should be one of the factors that increase Cd uptake in plants root.6.The response of antioxidant system in Idyll and Harukaze were studied under hydroponic conditions when the seedlings treated with Cd and with supplement of 200μmol L-1 GSH,respectively.The results showed that the accumulation of O2·-and H2O2 in plant roots and leaves was significantly reduced under Cd stress by application of GSH.Meanwhile,the activity of SOD and POD showed variation when the seedlings treated by Cd with supplement of GSH,these effects depended on the treatment time and plant tissues.The seedlings of cultivar Idyll treated for 3 and 6 days,application of GSH significantly increased the activity of CAT in plant roots and leaves.Compared with Cd stress alone,the activity of MDHAR in Idyll roots was significantly improved when the seedlings was applied by exogenous GSH.Similar results were also observed in Harukaze roots and leaves.The seedlings treated with Cd at medium and late stage(6 d and 10 d),the activity of APX in two cultivars roots and leaves were significantly increased by application GSH。Additionally,compared with Cd stress alone,a significant increase of the GR activity was observed in root and shoot of Harukaze by application of GSH.In this way,the circulation and regeneration of ASA and GSH was accelerated in Harukaze.The ratio of ASA/DHA and GSH/GSSG in roots and leaves were greatly increased by application of GSH compared with Cd stress alone,thus,the redox status was maintained in the plant.7.Idyll was used to be as material in this experiment under hydroponics.The root morphological and ultrastructure of chloroplast were studied when the seedlings treated with 50 μmol L-1 Cd stress and 200 μmol L-1 GSH,respectively.The results showed that water uptake was promoted when the exogenous GSH was applied into seedlings treated with Cd.AS a result,the root lengthy root area and root volume was greatly increased and then formed the developed root system.Thus,it is beneficial to increase the Cd uptake in plant root.Additionally,compared with Cd stress alone,Fe uptake in plant leaves was signiflcantly increased by application of GSH,the stable structure of chloroplast is maintained.It can contribute to enhance the photosynthesis of Italian ryegrass leaves.Thus,the shoot biomass were promoted. |
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