The Responses In Growth,Reproduction,and Physiological Metabolic Characteristics Of Chloris Virgata To Drought Stress And Nitrogen Addition In Songnen Grassland

Although the ecological restoration of grassland in China has achieved significant progress,the degradation,desertification and salinization of Northeast grassland has not been effectively curbed.The task of ecological restoration of grassland remains challenging.Nitrogen and water are the most critical factors limiting and driving plant growth,development,and vegetation dynamic in arid and semi-arid regions,especially for rain-fed annual grasses,water and nitrogen conditions have a greater impact on their birth and growth.Annual grasses widely distributed in various grassland ecosystems and serve as pioneer plants in the vegetation restoration or succession of severely degraded grasslands and bare land.However,current research on the growth,reproduction characteristics,and physiological traits of plants in Songnen grassland in response to drought stress and nitrogen addition mainly focuses on perennial species such as Leymus chinensis,while research on annual pioneer species is seriously lagging behind,with very few studies reported.To address this gap,this study selected Chloris virgata as the research subject and investigated the effects of drought stress（at 30-35%,50-55%,and 75-80%of soil field water holding capacity）,stress periods（early-drought:post-emergence-booting period,late-drought:emergence-seed maturity period,and continued drought during plant growth）and nitrogen addition(0,10 g m^-2)on growth,development and metabolic physiology.The research firstly studied the effects of both nitrogen addition and drought on the growth and reproductive characteristics.Subsequently,in the view of photosynthetic physiology and metabolic physiology（metabolomics）,the research explored the photosynthetic parameters,and the changes in differential metabolites and metabolic pathways in C.virgata.The aim is clarify how the growth,reproduction and physiological metabolism characteristics of C.virgata respond to drought stress and nitrogen addition,so as to provide a reference basis for future ultilization of C.virgata for vegetation restoration and management of‘degradation,desertification and salinization’grassland.The main findings are as follows:1.As the severity of drought stress increases,the plant height,main stem,tiller and branch dry weight,branch root number and root dry weight of C.virgata decreased significantly,resulting in a substantial decrease in both aboveground and belowground biomass.Compensatory growth was observed through a significant increase in the number of tillers and branches after re-wetting for early-drought treatment,while late and full-stage drought had minimal effects.Early-drought significantly reduced the dry weight of the main stem and its branches,but significantly increased the dry weight of tiller branches.The dry weight of tillers during the late stress period was greater than that in the early period,but it was not significant between various stress periods.Because the tiller traits of plant clusters were the main contributors to biomass,there was no significant difference in aboveground biomass among early-drought,late-drought and ambient water condition treatments,while the whole period of drought significantly reduced aboveground biomass.Nitrogen addition significantly increased the number of branches,tillers and their dry weight,as well as root dry weight and branch root number,resulting in a marked increase in aboveground and belowground biomass,and a significant reduction inroot length.Both drought stress and nitrogen addition could inhibit the main stem advantage of C.virgata and increase the number and biomass of non-main stem organs.Drought stress promoted the ability to adapt to severe drought by increasing the proportion of investment in the belowground part and increasing the root-shoot ratio,while nitrogen addition significantly reduced the root-shoot ratio.2.As the severity of drought stress increases,there was a tendency decrease for the number of branching spikes,total spikes,seed number per spike,spike weight and seed weight.However,the change in thousand seed weight showed the opposite trend.Early-drought stress significantly increases the number of spikes after rewetting,while it has less effect on the number of tiller spikes in the clumps.Early-drought also significantly increases the total number of spikes,resulting in significantly higher total spike weight and seed weight than the late-drought treatment.This indicates that drought stress has adverse effects on reproductive traits in the late-period of growth.Nitrogen addition significantly increases the main reproductive traits such as plant cluster,number of tiller spikes,number of branching spikes,total spikes,number of seeds,total spike weight and total seed weight.The total spike weight and total seed weight increase by 181.55%and 186.36%,respectively.The spike and seed biomass are highest under the condition of nitrogen addition without drought stress.Drought stress,early-drought and nitrogen addition significantly increased the allocation ratio of spike biomass and seed biomass,indicating that the trade-off between growth and reproduction has changed.3.As the severity of drought stress increases,the leaf area and biomass,net photosynthetic rate,stomatal conductance,water use efficiency and transpiration rate of the upper three functional leaves decreased significantly.However,the change of intercellular CO₂concentration was opposite to these trends.This indicates that the non-stomatal factors limited the photosynthetic performance of the main stem leaves under severe drought stress during the whole period,thereby inhibiting the reproduction of main stem traits.Under nitrogen addition condition,as the severity of drought stress increases,leaf nitrogen content and photosynthetic nitrogen use efficiency increased significantly,Adversely,these parameters decreased significantly without nitrogen addition,and leaf carbon content did not change significantly.Moreover,nitrogen addition had a positive impact on leaf area,biomass,net photosynthetic rate,stomatal conductance,water use efficiency,carbon and nitrogen content,and photosynthetic nitrogen use efficiency of the upper three leaves.Notably,nitrogen addition led to a substantial increase in the net photosynthetic rate and photosynthetic nitrogen use efficiency by 40.13%and 48.88%,respectively.These findings indicate that the effect of nitrogen addition on growth and reproductive was more significant than that of water conditions,but has no significant effect on intercellular CO₂concentration.Among the different leaf positions,the countdown third leaf exhibited the largest leaf area,biomass,and photosynthetic nitrogen use efficiency,while the first leaf had the smallest values,and intermediate in the second leaf,with no significant differences in net photosynthetic rate,intercellular CO₂concentration,and carbon content among the different leaf positions,suggesting that the upper third leaf had a relatively greater impact overall.4.Drought stress significantly increased amino acids,soluble sugars such as glucose,Maltose And Trehalose and organic heterocyclic compounds such as Myristicin,O-norferulic acid ester and Norcamptothecin.Nitrogen addition under drought stress not only resulted in the accumulation of amino acids with high nitrogen contents including Proline,Arginine and Asparagine,but also increased in lignans and nucleosides.However,the organic acids and flavonoids were increased in the treatment without nitrogen addition.These findings suggest that the composition of metabolites accumulated under drought stress is influenced by the presence or absence of additional nitrogen.Furthermore,nitrogen addition led to a substantial accumulation of amino acids,soluble sugars and fatty acids.Amino acids detected included Proline,Proline betaine and Threonine.Soluble sugars included alpha-D-glucose 1-phosphate,arabinose and inositol.Fatty acids such as Dodecanedioic acid,2-Hydroxyadipic acid and Traumatic acid.were also notably accumulated.Moreover,during drought stress with nitrogen addition,there was a significant increase in flavonoids and lignans,whereas nucleosides showed significant accumulation in the absence of drought stress,and glycerophospholipids were depleted.The results suggest that when C.virgata is supplied with nitrogen source,it enhances the synthesis of substances required for growth and development,along with providing energy through the synthesis of amino acids,soluble sugars,fatty acids and nucleotides.Additionally,the nitrogen induces synthesis of antioxidant like flavonoids and lignans,which help mitigate stress-induced damage caused by oxidative stress during drought.5.Drought stress induces changes in cysteine and methionine metabolism,as well as glyoxylic acid and dicarboxylic acid metabolism,indicates that the main pathways responding to drought stress are reactive oxygen scavenging pathways and energy metabolism pathways.Under drought stress,nitrogen addition has significant effects on multiple amino acid metabolism pathways,including phenylalanine metabolism,glycine,serine and threonine metabolism,alanine,aspartic acid and glutamic acid metabolism,as well as pyrimidine metabolism.Nitrogen addition stimulates glycine,serine and threonine metabolism.Moreover,under nitrogen addition,drought also significantly impacts tyrosine metabolism,arginine and proline metabolism,alanine,aspartic acid and glutamic acid metabolism,while nitrogen had a notable effect on pyrimidine metabolism under non-drought conditions.These findings suggest that nitrogen addition can promote growth and reproduction by activating some amino acid metabolism and pyrimidine metabolism.Furthermore,nitrogen addition can promote the synthesis of osmotic regulators by activating additional amino acid metabolism to mitigate the damage caused by drought stress.In summary,optimal water and nitrogen conditions significantly increase leaf traits,photosynthetic performance,nitrogen use efficiency,as well as promote amino acid metabolism and pyrimidine metabolism pathways,leading to a significant enhancement in the growth,reproductive traits,and biomass of C.virgata.The effect of nitrogen is significantly greater than that of water conditions.The damage induced by drought stress was significantly less during the early-drought period compared to the late-drought period.After re-wetting for the early-drought treatment,C.virgata employed a compensatory growth strategy to remedy its growth and reproductive capabilities,by increasing the number of branches.Additionally,nitrogen addition mitigate the damage of drought stress on growth and development by activating more amino acid metabolic pathways,etc.to synthesize osmotic substances and inducing the synthesis of antioxidants such as flavonoids and lignans.