Processes And Its Thermodynamic Characteristics On Nitrous Oxide Production In Greenhouse Vegetable Soils

Nitrous oxide(N2O)is a long-lived greenhouse gas and also contributes to ozone depletion in the stratosphere,and agricultural soil is a major source of N2O.Based on the flux measurements for decades in situ,N2O emissions have been attributed to N2O emission peaks resulted from natural anthropogenic or perturbations.Greenhouse vegetable production is characterized by high nitrogen(N)application rates,high cropping index and frequent farming practices,which results in more frequent and higher N2O emission peaks.Multiple pathways of N2O production occur in soil,but their significance and dependence on these mitigation strategies is poorly understood.To develop a comprehensive understanding of how different greenhouse vegetables soil types interact with these multiple controls and to quantify their influence on N2O emissions.In the present studies,soil samples were collected from six typical concentrated greenhouse vegetable production region,for laboratory incubations.Therefore,we used the incubation and field experiments simultaneously in the present study,which were divided into the following six sections:(1)Here,the combined approaches of a 15N tracing technique,DNA and mRNA analyses,and modeling were used to investigate the mechanisms underlying rapid nitrate(NO3-)accumulation and identify the relative contributions of autotrophic nitrification,heterotrophic nitrification,codenitrification and denitrification to nitrous oxide(N2O)production in six vegetable soils;(2)We used the inhibition method combined with the 15N-isotopomer analyses of N2O to estimate the role of ammonia oxidizers(AOA and AOB)and reactive nitrification intermediates(NH2OH and NO2-)in soil NO and N2O production and investigate the ?15Nbulk,?18O,and SP values of N2O produced from different biotic and abiotic processes that could possibly occur in soils;(3)Inhibitors of 1-octyne and 2-pheny 1-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide(PTIO)in combination with potassium chlorate were used to examine the importance of AOA and AOB in NO2-production and consumption,to evaluate the soil-specific effects on N2O production under six long-term fertilization greenhouse vegetable soils across China's mainland,and to predict their thermodynamic responses in a temperature gradient of 5-45?using the square root growth(SQRT)and macromolecular rate theory(MMRT)models;(4)The soil potential ammonia oxidation(PAO)and nitrite oxidation(PNO)rates were evaluated over a temperature gradient(5-45?)in six greenhouse vegetable soils from China's mainland and were modeled using both the square root growth(SQRT)and macromolecular rate theory(MMRT)models to quantify their thermodynamic responses.Abundances of ammonia-oxidizing archaea(AOA)and bacteria(AOB),encoded by amoA,and nitrite-oxidizing bacteria(NOB),encoded by nxrA and nxrB were determined using real-time PCR;(5)Combined approaches of the 15N-18O labeling technique and transcriptome analyses Were applied to investigate the pathways of N2O production(nitrifier nitrification,nitrifier denitrification,nitrification-coupled denitrification and heterotrophic denitrification)under 15,25 and 35? in six greenhouse vegetable field soils in China's mainland.The results of our study are summeraized as follow:1.Our results evidenced that the vegetable soils exhibited a leaky N cycle and that the acidic soils had a greater potential capacity to enhance NO3-accumulation,and the N2O production pathways were soil-specific.The soil pH had a positive effect on the NO3-retention capacity(p<0.01)and a negative effect on the net NO3-production rate(p<0.05),resulting in higher NO3-accumulation in acidic soils.Since the N losses via microbe-involved N cycling processes are dominant,thus,the vegetable soils exhibit a leaky N cycle.Meanwhile,autotrophic nitrification accounted for 39-86%of N2O production in alkaline soils,whereas denitrification was responsible for 85%of N2O production in acidic soils.The results of structural equation modeling indicated that autotrophic nitrification-derived N2O production was influenced by soil C/N ratio,the gross NO3-production rate and the ammonia-oxidizing archaea(AOA)amoA/ammonia-oxidizing bacteria(AOB)amoA mRNA ratio.while denitrification-derived N2O production was influenced by pH,the gross NO3-consumption rate and the abundance of nirS mRNA,all of which were influenced directly and indirectly by in situ climate parameters of mean annual precipitation and mean annual temperature.Furthermore,regression analyses revealed that the soil total N content affected heterotrophic nitrification and the pH affected codenitrification-derived N2O production,which contributed 9-76%and 0-7%,respectively,to the total N2O production in vegetable soils.This section revealed the characteristic of N transformation and cleared the N2O production pathways and derived factors through combined approaches of the 15N labeling technique,DNA and mRNA analyses,and modeling.2.Our data presents the suitability of the N2O isotopic analyses in disentangling soil N2O production pathways,and emphasize the coupled biotic-abiotic reactions acted as the crucial role in vegetable soils.The ammonia-oxidizing bacteria(AOB)dominate NO and N2O production in alkaline vegetable soils but that the compounds are primarily produced from the activity of ammonia-oxidizing archaea(AOA)in acidic vegetable soils.NO2-addition in the nonsterile and ?-sterilized samples stimulates the production of N2O from NH2OH,and the decomposition of NO2-primarily produces NO,while NH2OH play a more crucial role in biotic or abiotic N2O production in vegetable soils.In addition,most of the NH2OH-and NO2--dependent biotic or abiotic sources produce N2O with 27.4-36.5‰for SP values,which is in the same range as that from the AOA or AOB(25.1-34.2‰).Correlation analyses revealed that the variability of NO2-+NH2OH-induced N2O production can be explained by the soil organic carbon and iron contents.These results demonstrated that the coupled biotic-abiotic reactions acted as the crucial role in N2O production during nitrification,and can be proved by the SP values.3.The relative contributions of AOA and AOB to potential ammonia(NH3)oxidation(PAO)and nitrite(NO2-)consumption(PNC)were determined by thermodynamic parameters.The ammonia oxidizers-driven PAO and PNC exhibited a strong response to temperature with maximum rates attained at 35? or 40? by AOA compared to 30? or 35 ? by AOB.Concomitantly,compared with ammonium amendment,NO2-addition stimulated N2O production by approximately 2-4-fold for AOA and 2-9-fold for AOB at 20-40? in four of six soils.AOA exhibited a broader range of temperatures(-33.2-56.8? vs-16.1-52.3?)and lower relative temperature sensitivity(Q10)than AOB(p<0.05).The heat capacity(?Cp(?))of AOA and AOB in PAO and PNC increased in relation to the optimum temperature(Topt)and exhibited a greater negative value by AOB than AOA,indicating that the niche differentiation between AOA and AOB in PAO and PNC.Structural equation modeling revealed that potential AOA-driven N2O production was influenced by ?CP(?)and Q10 and C/N,while AOB-driven N2O production by ?Cp(?)and Q10 and soil nitrate(NO3-)content,all of which is influenced directly and indirectly by in situ climate parameters.This section mainly revealed that the different driving mechanisms of ammonia oxidation and nitrite consumption can be explained by the difference thermodynamic parameters between AO A and AOB.4.Elevated temperatures(>30?)resulted in the decoupling of ammonia oxidation(PAO)and nitrite oxidation(PNO),leading to nitrite accumulation,which stimulates N2O emissions.Both nitrite accumulation and consumption increased significantly with a temperature increase from 5 to 30 or 35 ?.The key characteristics(minimum temperature-Tmin,optimum temperature-Topt,maximum absolute temperature sensitivity-Tm_sens)for PAO and PNO were significantly different,and both models identified Topt for PAO(34.03 ?)that were significantly greater than those for PNO(25.96?).PAO was positively correlated with AOB-amoA at 20-30? and AOA-amoA at 30-35?,while PNO was positively correlated with Nitrospira-like nxrB at 5-30?,demonstrating that the temperature increase leads to greater differences of the specific growth rates among AOA,AOB and NOB.NO2-and N2O were positively correlated with the(AOA+AOB amoA)to(nxrA+nxrB)ratio,and the concentration of N2O was positively correlated with nitrite accumulation.These results highlight that the mechanisms of NO2-accumulation in "section 1" through the thermodynamic parameters between ammonia oxidation and nitrite oxidation,and microbial functional gene abundances among AO A,AOB and NOB.In addition,this section also explains the previous temperature response of AOA and AOB activity at the molecular level.5.Our findings discerned specific pathways and underlying microorganisms involved in GVF soils and highlighted that the N2O production pathways are affected by the interactions of temperature and soil properties.The results show that heterotrophic denitrification was an important source of N2O in acidic soils,whereas nitrifier-induced pathways(i.e.,nitrifier nitrification,nitrifier denitrification,nitrification-coupled denitrification)were the dominant source of N2O in alkaline soils.The contribution of nitrification-coupled denitrification increased with temperature at 15-35?,whereas nitrifier denitrification increased with temperature at 15-25?.Furthermore,nitrifier denitrification-derived N2O production were negatively correlated to AOA:AOB amoA at 15-25?.Structural equation modeling revealed that the temperature sensitivity(Q10)of N2O emissions was indirectly influenced by in situ climate-induced ammonium sorption capacities through mediation of functional gene transcript abundances,further leading to changes in the Q10 of different N2O production pathways.This section identified the mechanisms of temperature-controlled N2O production pathways through combined approaches of the 15N-18O labeling technique and transcriptome analyses.The results of this study revealed that the main reason for NO3-accumulation is that the N cycle in greenhouse vegetable soil is leaky,and the soil acidification has further increased NO3-accumulation and heterotrophic nitrification,heterotrophic denitrification and AOA derived N2O production.In addition,the coupled biotic-abiotic N2O production is an important mechanism of N2O production during nitrification.Ammonia-and nitrite-oxidation become uncoupled with temperature elevated at 30?,leading to NO2-accumulation and nitrifier denitrification derived N2O production.Furthermore,our results showed that AOA were more active at higher temperature,whereas AOB preferred lower temperature in nitrification.