N₂O Emission Characteristics And Relative Functional Genes Under Flooding-drying Conversion

Nitrous oxide?N₂O?is one of the major greenhouse gases.In recent years,the rapid increase of N₂O concentration contributes to global warming and ozone?O₃?depletion worse.Winter-flooded paddy is a typical rice-based cropping system.Conversion of winter-flooded paddy to rice-wheat rotation has been widely adopted in terms of improving the efficiency of land use and icreasing farmers'income.Management practices including drainage,tillage and fertilization that followed the conversion of winter-flooded paddy to rice-wheat rotation affected microbial activities and soil physicohemical properties,thereby influence N₂O emission.Previous studies have focused on N₂O characteristics after conversion from winter-flooded paddy to rice-wheat rotation However,little is known about the interation between N₂O emission,abundance of nitrify-denitrify functional genes in situ soil sample and soil environmental factors.To better understand N₂O emission processes and mechanisms after the conversion of winter-flooded paddy to rice-wheat rotation,our study was conducted at the Yanting Agro-Ecological Station of Purple Soil,a member station of the Chinese Ecosystem Research Network?CERN?,Chinese Academy of Sciences,in the central Sichuan Province,southwestern China.Three winter-flooded paddy fields historically cultivated in this region were arranged in a split-plot design.Half of each field was converted to rice-wheat rotation and the other half remained as a winter-flooded paddy.Each plot was further divided into conventional N fertilizer application and unfertilized treatments.Each treament has three replicates.During the experiment period,we measured the N₂O fluxes,sampled soil in situ and extracted soil total DNA,quantified the abundance of nitrification and denitrification functional genes,identify and quantify the key soil and environmental factors that control N₂O fluxes after the conversion.And explore the response of the abundances,diversities,and communities of nitrite reductase to the conversion of winter-flooded paddy to rice-wheat rotation.Based on the results in the field experiment,in term of further understanding N₂O emission processes and its controlling factors,we set up the incubation experiment,input different C/N ratios straws into the soil,incubated under flooding-drying and non-flooding condition.During the incubation,we monitored N₂O concentration in the vial,measured physicochemical properties of soil solution,also,we extracted soil DNA and did Q-PCR,trying to figure out key processes such as nitrification and denitrification affecting N₂O production and how microbial functional genes regulate N₂O production.The main results are as follows:1.Conversion of winter-flooded paddy to rice-wheat rotation triggered soil N₂O emissions which increased by 6.6-and 4.4-fold in the first and second years,respectively.High N₂O emission for winter-flooded paddy occurred during the rice season,contributing to 68.1%-75.7%of the total emission.The high emissions in the initial stage of the wheat season represented 45%-82%of the annual cumulative N₂O emissions in N-fertilized and unfertilized treatments.N₂O emission pulses are not only primarily affected by N fertilization but are also influenced by soil moisture and organic matter mineralization.2.N₂O was positively correlated with WFPS.High N₂O fluxes occurred in the first 20days of the wheat season when soil WFPS was in the range of 68%-72%.The contribution rate of WFPS was 22%-43%for both fertilized and unfertilized treatments of the rice-wheat rotation.3.The contribution rate of DOC reached up to 26%-36%.High N₂O fluxes occurred when the ratio of DOC to inorganic N ranged from 0.47 to 1.42.Additionally,N₂O peaked as the DOC/?NH₄⁺+NO₃^-?ratio approached 1.4.nirK gene abundance play a dominant role in winter-flooded paddy,while nirS gene abundance play a dominant role in rice-wheat rotation.Conversion of winter-flooded paddy to rice-wheat rotation inceased the richness and diversity of nirK gene and the richness of nirS gene,but decreased nirS gene diversity.Rice-wheat rotation changed the composition of nirK and nirS gene micro-organisms in the levels of phylum,class,order,family and genus.Compared with winter-flooded paddy,the composition of nirK from rice-wheat rotaion increased euryarchaeota,planctomycetes and firmicutes.The composition of nirS is lack of bacteroidetes and chloroflexi.DOC was the main factor affecting the composition of nirK.WFPS was the main factor affecting the composition of nirS.5.Wheat straw addition enhanced N₂O emission,with greater emissions from the lower C/N ratio amendments.In flooded treatments,the average N₂O fluxes for the low C/N ratio and high C/N ratio treatments were 3 and 2 times higher than for control.In non-flooded treatments,the average N₂O fluxes for the low C/N ratio and high C/N ratio treatments were 2 and 1 times higher than for control.Additionally,N₂O emission from straw addition treatments was exacerbated when soil was under flooding-drying condition.The average N₂O fluxes in flooded treatments were 2 times higher than in non-flooded treatments.6.Compared with the control,straw addition significantly increased 16S rRNA,denitrifier?nirK,nirS and nosZ?gene abundances but did not change nitrifier?AOA and AOB?gene abundances in two soils.However,straw addition increased the AOA/AOB ratio in the UK brown earth soil.The ratio of AOA/AOB for flooded treatments were significantly higher than non-flooded treatments.Flooding altered the ratio of nitrify archaea and bacteria gene.7.In the Chinese paddy soil,68.8%of the N₂O flux variance was explained by denitrifier processes,relative abundance of nirK and nirS,and by abiotic factors?WFPS,soil solution NO₃^-and DOC concentrations?.In the UK brown earth,68.5%of N₂O flux variation was explained by nitrifier and denitrifier processes,relative AOA and nirS and by abiotic factors?WFPS,soil solution DOC,NH₄⁺and NO₃^-concentrations?.The results demonstrated that denitrification process is the predominant pathway of N₂O production during the drying phase.8.N₂O flux was significantly positive linear relationship to CO₂ flux in flooding-drying and non-flooding treatments with different C/N ratios straw addition?R² value varied from 0.23 to 0.31,p<0.01?.For winter-flooded treatment with fertilization,N₂O flux was significantly positive linear relationship to CO₂flux?R² value varied from 0.15 to 0.18,p<0.01?,while,for rice-wheat rotation,N₂O flux was negative linear relationship to CO₂ flux?R² varied from 0.19 to 0.34,p<0.01?.In summary,conversion of winter-flooded paddy to rice-wheat rotation increased N₂O emission.High N₂O emissions occurred in the initial stage of wheat season.Conversion of winter-flooded paddy to rice-wheat rotation inceased the richness and diversity of nirK gene and the richness of nirS gene,but decreased nirS gene diversity.Soil moisture and available carbon were the key factors controlling N₂O emission and nitrify-denitrify functional genes.These findings suggest that N₂O emission during the initial stage immediately after the conversion should be considered when evaluating the environmental consequeces of land-use conversion.These findings also suggest that the conversion of winter-flooded paddy to rice-wheat rotation increased N₂O emissions that could be mitigated by controlling the soil moisture and ratio of available soil carbon to nitrogen.