Study On The Effect Of Calcium On Pinus Massoniana Seedlings

Calcium is one of the essential mid-weight elements for plant growth and development.In the natural condition,calcium deficiency in the soil is a fairly rare occurrence,but it tends to occur in some soil with low salt saturation and severe acid deposition.When the soil is deficient in calcium,the Ca²⁺balance in plants is destroyed and calcium deficiency is likely to occur.However,excessive calcium in the soil not only destroys normal physiological and biochemical processes and nutrient element metabolism in the plant,but also adversely affects plant morphology and internal structural changes,causing toxic effects on plants.Pinus massoniana Lamd.?P.massoniana?is one of the most important industrial timber species in China,with high economic,social and ecological benefits.Long-term planting of P.massoniana will increase the soil and water loss in the forest land,resulting in lack of nutrients in the forest land.Periodic harvesting will remove a large number of nutrient elements,and affect the circulation of calcium in the ecosystem,resulting in net consumption of nutrients and calcium loss in the forest land.As a result,soil fertility declined.meanwhile,acid rain caused leaching of salt ions in P.massoniana forest land,especially the loss of Ca²⁺.In addition,the parent rock is one of the main factors affecting the growth of P.massoniana.The content of CaO of soil is negatively correlated with the high growth of P.massoniana,hence the P.massoniana has a poor growth and development in the soil that carbonate rocks are developed.However,the mechanisms behind these phenomena are still unclear.Calcium is an essential plant nutrient for the growth and development of P.massoniana,but the influencing mechanism of different calcium levels on physiological and biochemical functions,nutrient element metabolism and material composition of P.massoniana is still unknown.In this paper,taking 1-year-old seedlings of P.massoniana as test materials,greenhouse sand culture method was used to study the influence rules of different calcium levels on plant growth,physiological and biochemical characteristics,cell structure,nutrient metabolism,and nutrient metabolize of P.massoniana seedlings,in order to reveal the available calcium concentrations for optimal growth of P.massoniana seedlings,and the influence rules of different Ca²⁺levels on seedlings development.Moreover,this paper also discussed the responsive mechanisms of physiological and biochemical characteristics,nutrient metabolism,anatomical structure and ultrastructure of needles and substance components of P.massoniana seedlings under different Ca²⁺levels.The main results were as follows:1)Under nutrient quartz sand culture experiment in the greenhouse,masson pine seedlings were suitable for growth at a level of 1-3 mmol?L^-1 Ca²⁺,which 2 mmol?L^-1Ca²⁺grew best.In high calcium conditions,seedlings were inhibited more severely than in low calcium environments.The growth index of P.massoniana showed a quadratic function relationship with the calcium supply concentration.The growth indexes of P.massoniana seedlings,including survival rate,root biomass,root tips,main root length,root volume,needle length,needle width,diameter increment and height increment etc,had very closely relationships with their growth and development?p<0.001?.Such relationship could help accurately reflect relationship among source and P.massoniana and Ca²⁺concentrations.2)The decrease in Pn of P.massoniana under low calcium conditions was mainly caused by non-stomatal limitation,but was mainly caused by stomatal limitation under high-Ca conditions.As the dominant factors,the Fv/Fm,chlorophyll?a+b?,chlorophyll a,soluble sugar,Ci,Cond,and chlorophyll a/b of needles of P.massoniana,which were related to the calcium level in the environment,they played a very critical role in physiological and biochemical metabolic processes,osmotic adjusting ability,and light absorption,transformation and utilization of P.massoniana.Under calcium stress conditions,ROS(O₂^?.and H₂O₂)increased,chlorophyll content,Pn,Fv/Fm,Ci,and Cond decreased in needles,which caused damage to the cell membrane and increased MDA content,resulting in inhibition of plant growth.However,seedlings synthesized and accumulated the osmotic adjustment substances such as proline,soluble sugar and protein by increasing the activities of SOD and POD enzymes,so as to relieve the damage caused by calcium stress to P.massoniana seedlings.3)Under low calcium conditions,the absorption of Ca by P.massoniana had a synergistic effect on the absorption of N and P.However,it exhibited an overall inhibitory effect under high-Ca conditions and significantly reduced the absorption of P under excessive calcium conditions.With the increase of calcium levels,the K content in the root of P.massoniana seedlings increased gradually,while the K content in stems,biennials and needles of the year presented the tendency of increasing first and then decreasing.The Ca content in all organs increased,while Mg in roots,stems and current-year needles reduced,showing a significant antagonistic effect on the absorption and accumulation of Ca in the corresponding organs.The contents of trace elements in the organs of P.massoniana were different at various Ca²⁺levels.The overall performance was that there was relatively more absorption and accumulation of Mn,and less absorption and accumulation of Cu.There was a clear synergy between Fe and Ca under low calcium conditions.However,there was an antagonistic effect between Fe and Ca under high calcium conditions.Ca and Mn have some synergistic effects,while Ca and Zn have some antagonism.Under the conditions of suitable calcium or high calcium,Ca and Cu showed antagonistic effects,while the relationship between Ca and Cu was not significant in low calcium conditions.Metabolism characteristics of N,Ca and Mn of seedlings under different Ca²⁺levels were less affected,but it was influenced largely under different Ca²⁺levels.Meanwhile Zn migration and transportation from the roots was obviously influenced by different Ca²⁺levels.Moreover,metabolism characteristics of P and Fe of P.massoniana under high Ca²⁺levels were distracted,and thus the effect growth impact.The cell wall was the largest calcium reservoir in needles of P.massoniana,whose Ca content increased with the increasing Ca²⁺levels and played a key role in balancing Ca²⁺concentration in cells.Among the intracellular components of needles,calcium was mainly present in the nuclear protein and soluble components,followed by mitochondria and chloroplasts.With the different Ca²⁺levels,Ca contents of cell wall in 2-year-old needles was increased and the proportion of Ca content was decreased when compared with 1-year-old needles.The proportion of non-active calcium in subcellular components of P.massoniana needles was relatively high,including calcium oxalate and calcium silicate etc.The content of calcium oxalate increased significantly with the increase of Ca²⁺?p<0.05?.The formation of calcium oxalate,under high Ca^2+levels,was beneficial to regulating Ca²⁺concentration in cells.Under high calcium conditions,the proportion of calcium in subcellular components of 2-year-old needles decreased,while the proportion of calcium in only some subcellular components decreased in low calcium conditions.In insufficient calcium environments,the proportion of active calcium in needles of P.massoniana was increased,while the proportion in high calcium conditions was decreased.4)The Ca²⁺level was positively correlated with the cuticle thickness of P.massoniana?p<0.05?,and was significantly positively correlated with the thickness of the epidermis,the perimeter of the section and the area of needles?p<0.01?.Under the conditions of calcium stress,the injury caused by calcium stress could be alleviated by increasing the tightness of the arrangement of epidermal cells,the diameter of the resin tract,the thickness of the mesophyll and the perimeter and area of the vascular tissue.Under calcium stress,the ultrastructure of P.massoniana needles was destroyed.The chloroplast lamella structure was blurred or even disintegrated.The mitochondrial deformation or double membrane structure disappeared.The content of starch granules was decreased.The osmiophilic granules were increased.The plasmolysis phenomenon was appeared.The growth of P.massoniana was inhibited to varying degrees.The seedlings could enhance its stress resistance by inducing some cells death.5)Under calcium stress conditions,there were some common typical absorption peaks in the same organs of different treatments of P.massoniana,and some new characteristic peaks or deletions;some of the same absorption peaks in different organs of P.massoniana were displaced.There were also some absorption peaks missing or increasing,and some differences in the change rules of the absorption peaks.According to the peaks of the absorption peaks at the locations of 2924 cm^-1,1626 cm^-1 and 1064 cm^-1 in needles of P.massoniana,the degree of calcium stress in P.massoniana was determined.Specific absorption peaks,including the nearby locations of 1552 cm^-1 in needles and 1159 cm^-1 in the stems in the low Ca²⁺levels and 1036 cm^-1 in needles and 1390 cm^-1 in the stems in the Ca²⁺levels,could be used as an important indicator in the screening and cultivation of different adaptive new germplasms of P.massoniana.The growth of P.massoniana seedlings under 1-3 mmol?L^-1 Ca²⁺,and the seedlings grew best in 2 mmol?L^-1 Ca²⁺.Under calcium stress condition,P.massoniana seedlings could enhance their adaptation by some particular approaches,including increasing the activity of antioxidases,actively accumulating the osmotic substances,decreasing the photosynthesis rate,promoting the partial cellular senescence,adjusting the anatomy characteristics of needles,changing the nutrient metabolic characteristics,readjusting the composition of organic matter,decreasing the growth rate,and so on.A reference method was provided for calcium fertilizer application of forests and seedlings of P.massoniana,and the foundations for the creation of germplasms with high tolerance to different calcium concentrations was emerging.