Effects Of Liming On Nitrogen Cycling And The Underlying Mechanisms In A Double Rice-cropping System

Rice(Oryza sativa L.)is a staple food for approximately half of the world's population.China is the world's largest rice producer,accounting for 21.8%of global rice production.The double rice-cropping system accounts for nearly 39.0%of the total rice-growing area in China,and thus is important for maintaining rice-growing area,increasing rice production,and ensuring food security.However,excess use of nitrogen(N)fertilizer and inherent low soil p H lead to soil acidification in a large area of paddies in subtropical China.Soil acidification reduces soil microbial abundance and diversity,inhibits root uptake for mineral nutrient,thereby affecting N fertilizer use efficiency and N-cycling in paddy fields.We conducted a four-year(2015-2018)consecutive field experiments(Applying lime only once before the transplanting of early rice in 2015).Results showed that liming significantly increased grain yield and N uptake.However,the effects of liming on N use efficiency,N-cycling processes and the underlying mechanisms remain unclear.Moreover,neither grain yield nor N uptake were affected by liming,and soil p H was back to initial levels in 2018.Therefore,we continued the field experiment by re-applying lime following the harvest of late rice in 2018.A ¹⁵N-labled micro-plot experiment was conducted to clarify the effects of liming on the fates of fertilizer-and straw-N.NH₃ volatilization,N₂O emission,and N leaching were determined to examine the effects of liming on reactive N losses.Meanwhile,a pot experiment was conducted to determine the direct effects of Ca²⁺addition on rice yield and N uptake in order to separate the alkaline effect of lime.Finally,physiological and ecological factors related to N-cycling,and soil microbiological properties were examined to clarify the underlying mechanisms of liming effects on N-cycling in the double rice-cropping system.Main results are as follows:1.Liming increased grain yield and N uptake by 8.7%and 9.6%in early rice,and by9.3%and 12.3%in late rice of the second liming cycle in 2019,respectively.The increase in rice yield was mainly attributed to higher panicle numbers.There was no significant interaction between liming and straw incorporation on grain yield and N uptake.Neither grain yield nor N uptake were affected by applying Ca Cl₂in the pot experiment.Therefore,the increase in grain yield and N uptake with lime application was mainly due to its alkaline effect,rather than the effect of Ca²⁺addition.2.¹⁵N isotope tracing indicated that liming increased rice N uptake from straw(+27.5%and+36.2%in early-and late rice,respectively),while reducing N uptake from urea(-7.8%and-3.1%in early-and late rice,respectively).Liming reduced urea-N recovery rate and retention rate,and consequently increased urea-N loss rate.Liming increased straw-N recovery rate and loss rate,but reduced straw-N retention rate.3.The major pathways of reactive N losses in the paddies were ammonia(NH₃)volatilization and N leaching,while the N₂O emissions were quite low.Liming significantly interacted with straw incorporation to improve NH₃ volatilization in early rice.Liming increased NH₃ volatilization more strongly in plots with straw incorporation by 2.2times compared to plots without straw incorporation by 62.0%in early rice.On average,liming enhanced NH₃ volatilization by 17.4%in late rice,whereas no interactive effect was found on NH₃ volatilization between liming and straw incorporation in late rice.NH₄⁺-N concentration in leachates was not significantly affected by liming in the paddies.Liming increased NO₃^--N concentration in leachates at mid-tillering and panicle initiation stages in early rice as well as at seedling stage in late rice.In addition,total organic nitrogen(TON)concentration in leachates was increased by liming at the seedling stage in early rice,but no significant effect was found in late rice.4.Liming significantly increased leaf SPAD values,except at the mid-tillering stage.Liming interacted with straw incorporation to improve root total absorption area and active absorption area at the heading stage in the rice paddy.With straw incorporation,liming increased root total absorption area and active absorption area by 36.8%and 35.0%for early rice,and by 25.4%and 32.8%for late rice,respectively,with no effect in plots without straw incorporation.In addition,root oxidation activity was increased by liming at the heading stage in late rice.Liming interacted with straw incorporation to enhance root volume at the heading stage for early rice.Liming increased root volume in plots with straw incorporation by 18.3%,but no effect was found in plots without straw incorporation.Root length,root surface area,and root tip numbers were increased by liming at the heading stage for early rice.Liming significantly enhanced root length and root tip numbers for late rice.5.Overall,liming did not affect straw decomposition rate at the early period of late-and early-rice straw incorporation,which,however,enhanced straw decomposition at the late period.Liming significantly increased the decomposition rate at 125th,182th,and203th days following the retention of late-rice straw,and at 30th and 48th days following the retention of early-rice straw.Similarly,liming increased N release in rice straw.Liming increased floodwater p H,total organic carbon(TOC)concentrations and N concentrations in floodwater(e.g.NH₄⁺-N,NO₃^--N,and TON).Overall,liming enhanced soil NH₄⁺-N and NO₃^--N concentrations in the rice paddy.6.Liming significantly increased soil AOB-amo A gene copy numbers,while no effect was found on soil AOA-amo A gene copies in the paddy field.Liming significantly interacted with straw incorporation to reduce soil nir S gene copies at the mid-tillering stage in the early rice season.Liming reduced soil nir S gene copies more strongly in plots without straw(-32.4%)compared to plots with straw incorporation(-27.4%).By contrast,liming increased soil nir S gene copies at the heading stage in the early rice season,without significant effect in the late rice season.Unexpectedly,liming did not affect the nar G,nir K and nos Z gene copies.Liming enhanced nrf A gene copies except at the mid-tillering stage in the early rice season.Liming increased nifH gene copies in the late rice season,but no effect was found in the early rice season.In addition,hzs B gene was not found in the present study.7.Liming increased Alpha diversity indexes of AOA-amo A and AOB-amo A genes.Liming increased the OTU richness of AOA-amo A and AOB-amo A genes in the early rice season,while no effect was found in the late rice season.Liming enhanced Alpha diversity indexes of nir S gene,but no effect was found on Alpha diversity indexes of nar G,nir K and nos Z genes in the present study.Liming increased the OTU richness of nir S gene in both early-and late rice seasons.Liming enhanced the Alpha diversity index and OTU richness of nrf A gene.Liming increased the Alpha diversity index of nifH gene.The OTU richness of nifH gene was enhanced by liming in the early rice season,but no effect was found in the late rice season.Both Redundancy analysis and Spearman correlation analysis revealed that soil p H has a major influence on nir S,nrf A,and nifH community structure on the OTU level.Taken together,we conclude that liming increases grain yield in the double rice cropping system,which is due to the increase in N uptake that enhances panicle numbers.The increase in N uptake with lime application is mainly attributed to its alkaline effect,rather than the Ca²⁺application.Liming promotes straw decomposition,and enhances root oxidation activity and absorption area,which contributes to rice N uptake from straw.In contrast,liming reduces N uptake from urea mainly due to the enhance in NH₃volatilization and N leaching in the rice paddy.On the one hand,liming increases the abundance and diversity of soil AOB-amo A and nir S genes,which may stimulate N losses through nitrification and denitrification.On the other hand,liming enhances the abundance and diversity of soil nrf A and nifH genes,which may benefit N retention and promote biological N fixation.The long-term effect of liming on soil N pool and the relative contribution of different functional microorganisms on N-cycling processes need further investigation in the future.