Germplasm Screening For Salt Tolerance And Physiological Responses To Exogenous Spermidine For Improving Salt Tolerance In Zoysiagrass

Ninty-nine accessions which were collected in the east part of China were used as plant materials for screening salt tolerance and analyzing evaluation ratings in Zoysiagrass(Zoysia japonica Steud.).Based on the screening results,2 accessions,including 1 tolerant accession,and 1 suscuptible accession were identified for further physiological research about an improvement effect of salt tolerance by application of exogenous spermidine in zoysiagrass.The primary results and conclusions are summarized as follows:1.Screening of salt tolerance and evaluation rating evaluation.Two NaCl concentration levels(0 mM and 300 mM)were used for salt tolerance screening.Leaf were measured at 28 days after the tested plants were grown under a NaCl salt stress.Leaf length,leaf weight,and leaf chlorophyll content were recorded.The results indicated leaf weight of a plant were significantly reduced under the NaCl stress treatment comparing with control in both tolerant and susceptible accessions.The ratio of Na/K in leaf went up under NaCl salt stress.Some zoysiagrass accessions,including Z009 and Z057 which fresh leaf weight only reduced by 7%comparison with no NaCl treatment,displayed a good salt tolerance.Cluster analysis showed that accessions collected from the same site had a similar salt tolerance.All zoysiagrass accessions can be divided into five groups through clustering analysis.Correlation analysis showed that leaf dry weight,fresh weight,leaf Na+ and K+ contents can be used for evaluating salt tolerance in Zoysiagrass.2.Effects of salt stress on the zoysiagrass physiological characteristics.Two accessions,including 2057,a tolerance accession to salt and Z081,a sensentive to salt,which were identified by accessions screening were cultured under different NaCl salt concentrations(0 mM,150 mM,300 mM and 450 mM).The leaf contents of Na+,K+,proline,malondialdehyde and the enzyme activities of superoxide dismutase,peroxidase and peroxidase,and leaf SPAD value were measured at different time intervals(0 days,7 days,14 days,21 days and 28 days).In both accessions,leaf Na+ content,leaf proline and malondialdehyde contents,and the activities of superoxide dismutase,peroxidase and peroxidase were increased by salt treatment.No big changes in these traits were recorded after 21 days,but the maximum values were recorded at 28 days under 450 mM NaCl treatment.The leaf K+ concentration was decreasing with the increasing of NaCl salt concentrations.Under 150 mM and 300 mM NaCl salt treatments,the leaf SPAD value in both assessions zoysia were higher than the value of contral,but the values were lower than that of control at 450 mM NaCl salt treatment.3.The effects of exogenous spermidine application on the polyamines forms in the polyamine metabolic paths for improving salt tolerance.Two accessions,including Z057,a tolerance accession to salt and Z081,a sensentive to salt,which were identified by accessions screening,were cultured with four treatments(CK,150 mM NaCl,0.15 mM Spd and 150 mM NaCl+0.15 mM Spd).Comparison with the control,leaf fresh weight were reduced by a salt stress,but free(Spd and Spm),soluble(Put,Spd and Spm)and insoluble(Put,Spd and Spm)in both accessions were increased under salt stress.Moreover,the activities of arginine decarboxylase(ADC),ornithine decarboxylase(ODC),S-adenosylmethionine decarboxylase(SAMDC)and diamine oxidase(DAO)were enhanced by a salt stress.Polyamines metabolisam was changed by exogenous Spd application,ADC,SAMDC and DAO activities were increased,but free Put concentration level was reduced under the salt stress treatment in both accessions.In addition,Path Analysis showed that ODC,SAMDC and DAO significantly contributed to PA metabolism,and endogenous Spd levels also contributed to endogenous Spm levels.Free PAs are surposed to be the primary factor which cause the value variation in other PA forms.4.The effect of exogenous spermidine concentration on polyamine metabolism and salt tolerance in zoysiagrass.Two accessions,including Z057,a tolerance accession to salt and Z081,a sensentive to salt,which were identified by accessions screening,were cultured under different Spd concentrations(0 mM,0.15 mM,0.30 mM,0.45 mM,0.6 mM)under salt sress.Polyamine(diamine putrescine(Put),tetraamine spermine(Spm)and Spd),H2O2 and malondialdehyde(MDA)contents,polyamine metabolic(ADC,ODC,SAMDC,PAO and DAO),and antioxidant(superoxide dismutase,catalase,and peroxidase)enzyme activities were measured.The results showed that salt stress induced increases in Spd and Spm contents and ornithine decarboxylase(ODC),S-adenosylmethionine decarboxylase(SAMDC)and diamine oxidase(DAO)activities in both accessions.Exogenous Spd application did not alter polyamine contents via regulation of polyamine-degrading enzymes,and an increase in polyamine biosynthetic enzyme levels was observed.Increasing the concentration of exogenous Spd resulted in a tendency of the Spd and Spin contents and ODC,SAMDC,DAO,and antioxidant enzyme activities to first increase and then decrease,peaking in both accessions with exogenous 0.3 mM Spd.H2O2 and MDA levels significantly decreased in both accessions treated with Spd.The ratio of(Spd+Spm)/Put first increased and then decreased,peak value is 3.1 and 2.24 in Z057 and Z081 with 0.3 mM exogenous Spd.Additionally,in both accessions,positive correlations between polyamine biosynthetic enzymes(ADC,SAMDC),DAO,and antioxidant enzymes(SOD,POD,CAT),but negative correlations with H2O2 and MDA levels.The results indicated that the salt tolerance of zoysiagrass species was significantly different under salt stress.The changes of Na+,K+,proline and MDA content and antioxidant enzyme activity were closely related to salt tolerance of zoysiagrass.Exogenous Spd plays an important role in improving salt tolerance of zoysiagrass.