Potential Secretory Transporters and Biosynthetic Precursors of Biological Nitrification Inhibitor 1,9-Decanediol in Rice as Revealed by Transcriptome and Metabolome Analyses

Biological nitrification inhibitors(BNIs)are released from plant roots and inhibit the nitrification activity of microorganisms in soils,reducing NO_(3)^(‒)leaching and N2O emissions,and increasing nitrogenuse efficiency(NUE).Several recent studies have focused on the identification of new BNIs,yet little is known about the genetic loci that govern their biosynthesis and secretion.We applied a combined transcriptomic and metabolomic analysis to investigate possible biosynthetic pathways and transporters involved in the biosynthesis and release of BNI 1,9-decanediol(1,9-D),which was previously identified in rice root exudates.Our results linked four fatty acids,icosapentaenoic acid,linoleate,norlinolenic acid,and polyhydroxy-α,ω-divarboxylic acid,with 1,9-D biosynthesis and three transporter families,namely the ATP-binding cassette protein family,the multidrug and toxic compound extrusion family,and the major facilitator superfamily,with 1,9-D release from roots into the soil medium.Our finding provided candidates for further work on the genes implicated in the biosynthesis and secretion of 1,9-D and pinpoint genetic loci for crop breeding to improve NUE by enhancing 1,9-D secretion,with the potential to reduce NO_(3)^(‒)leaching and N2O emissions from agricultural soils.

Effect of nitrification inhibitor DMPP on nitrogen leaching, nitrifying organisms, and enzyme activities in a rice-oilseed rape cropping system

DMPP （3,4-dimethylpyrazole phosphate） has been used to reduce nitrogen （N） loss from leaching or denitrification and to improve N supply in agricultural land. However, its impact on soil nitrifying organisms and enzyme activities involved in N cycling is largely unknown. Therefore, an on-farm experiment, for two years, has been conducted, to elucidate the effects of DMPP on mineral N （NH4^＋- N and NO3^--N） leaching, nitrifying organisms, and denitrifying enzymes in a rice-oilseed rape cropping system. Three treatments including urea alone （UA）, urea ＋ 1% DMPP （DP）, and no fertilizer （CK）, have been carded out. The results showed that DP enhanced the mean NH4^＋-N concentrations by 19.1%-24.3%, but reduced the mean NO3^--N concentrations by 44.9%-56.6% in the leachate, under a two-year rice-rape rotation, compared to the UA treatment. The population of ammonia oxidizing bacteria, the activity of nitrate reductase, and nitrite reductase in the DP treatment decreased about 24.5%-30.9%, 14.9%-43.5%, and 14.7%-31.6%, respectively, as compared to the UA treatment. However, nitrite oxidizing bacteria and hydroxylamine reductase remained almost unaffected by DMPP. It is proposed that DMPP has the potential to either reduce NO3^--N leaching by inhibiting ammonia oxidization or N losses from denitrification, which is in favor of the N conversations in the rice-oilseed rape cropping system.

Influences of nitrification inhibitor 3,4-dimethyl pyrazole phosphate on nitrogen and soil salt-ion leaching

An undisturbed heavy clay soil column experiment was conducted to examine the influence of the new nitrification inhibitor, 3,4- dimethylpyrazole phosphate （DMPP）, on nitrogen and soil salt-ion leaching. Regular urea was selected as the nitrogen source in the soil. The results showed that the cumulative leaching losses of soil nitrate-N under the treatment of urea with DMPP were from 57.5% to 63.3% lower than those of the treatment of urea without DMPP. The use of nitrification inhibitors as nitrate leaching retardants may be a proposal in regulations to prevent groundwater contaminant. However, there were no great difference between urea and urea with DMPP treatments on ammonium-N leaching. Moreover, the soil salt-ion leaching losses of Ca^2＋, Mg^2＋, K^＋, and Na^＋ were reduced from 26.6% to 28.8%, 21.3% to 27.8%, 33.3% to 35.5%, and 21.7% to 32.1%, respectively. So, the leaching losses of soil salt-ion were declined for nitrification inhibitor DMPP addition, being beneficial to shallow groundwater protection and growth of crop. These results indicated the possibility of ammonium or ammonium producing compounds using nitrification inhibitor DMPP to control the nitrate and nutrient cation leaching losses, minimizing the risk of nitrate pollution in shallow groundwater.

Mitigating N2O and NO Emissions from Direct-Seeded Rice with Nitrification Inhibitor and Urea Deep Placement

Soil-emitted nitrous oxide(N2O) and nitric oxide(NO) in crop production are harmful nitrogen(N) emissions that may contribute both directly and indirectly to global warming. Application of nitrification inhibitors, such as dicyandiamide(DCD), and urea deep placement(UDP), are considered effective approaches to reduce these emissions. This study investigated the effects of DCD and UDP, compared to urea and potassium nitrate, on emissions, nitrogen use efficiency and grain yields under direct-seeded rice. High-frequency measurements of N2O and NO emissions were conducted using the automated closed chamber method throughout the crop-growing season and during the ratoon crop. Both UDP and DCD were effective in reducing N2O emissions by 95% and 73%, respectively. The highest emission factor(1.53% of applied N) was observed in urea, while the lowest was in UDP(0.08%). Emission peaks were mainly associated with fertilization events and appeared within one to two weeks of fertilization. Those emission peaks contributed to 65%–98% of the total seasonal emissions. Residual effects of fertilizer treatments on the N2O emissions from the ratoon crop were not significant;however, the urea treatment contributed 2%, whereas UDP contributed to 44% of the total annual emissions. On the other hand, cumulative NO emissions were not significant in either the rice or ratoon crops. UDP and DCD increased grain yields by 16%–19% and N recovery efficiency by 30%–40% over urea. The results suggested that the use of DCD and UDP could mitigate N2O emissions and increase grain yields and nitrogen use efficiency under direct-seeded rice condition.

N_2O emissions from a cultivated Andisol after application of nitrogen fertilizers with or without nitrification inhibitor under soil moisture regime

The aim of this work was to examine the emission of N 2O from soils following addition of nitrogen fertilizer with a nitrification inhibitor(+inh) or without the nitrification inhibitor(-inh) at different soil water regime. Higher soil moisture contents increased the total N 2O emissions in all treatments with total emissions being 7 times larger for the CK and >20 times larger for the fertilizer treatments at 85% WFPS(soil water filled pore space) than at 40% WFPS. The rates of N 2O emissions at 40% WFPS under all treatments were small. The maximum emission rate at 55% WFPS without the nitrification inhibitor(-inh) occurred later (day 11) than those of 70% WFPS (-inh) samples (day 8). The inhibition period was 4—22 d for 55% WFPS and 1—15 d for 70% WFPS comparing the rates of N 2O emissions treated (+inh) with (-inh). The maximum emission rates at 85% WFPS were higher than those at the other levels of soil water content for all treatments. The samples(+inh) released less N 2O than (-inh) samples at the early stage. Nevertheless, N 2O emissions from (+inh) samples lasted longer than in the (-inh) treatment. Changes in mineral N at 55%, 70% and 85% WFPS followed the same pattern. NH + 4-N concentrations decreased while NO - 3-N concentrations increased from the beginning of incubation. NH + 4-N concentrations from 40% WFPS treatment declined more slowly than those of the other three levels of soil water content. Nitrification was faster in the (-inh) samples with 100% NH + 4-N nitrified after 22 d(50% WFPS) and 15 d(70% and 85% WFPS). N 2O emissions increased with soil water content. Adding N-fertilizer increased emissions of N 2O. The application of the nitrification inhibitor significantly reduced total N 2O emissions from 30.5%(at 85%WFPS) to 43.6%(at 55% WFPS).

Wastewater Treatment in a Hybrid Biological Reactor (HBR): Nitrification Characteristics

Objective To investigate the nitrifying characteristics of both suspended- and attached- biomass in a hybrid bioreactor. Methods The hybrid biological reactor was developed by introducing porous ceramic particles into the reactor to provide the surface for biomass attachment. Microorganisms immobilized on the ceramics were observed using scanning electron microscopy (SEM). All chemical analyses were performed in accordance with standard methods. Results The suspended- and attached-biomass had approximately the same nitrification activity. The nitrifying kinetic was independent of the initial biomass concentration, and the attached-biomass had a stronger ability to resist the nitrification inhibitor. Conclusion The attached biomass is superior to suspended-biomass for nitrifying wastewater, especially that containing toxic organic compounds. The hybrid biological reactor consisting of suspended- and attached-biomass is advantageous in such cases.

Inhibition of nitrification in soil by metal diethyldithiocarbamates

Nitrification acts as a key process in determining fertilizer use efficiency by crops as well as nitrogen losses from soils. Metal dithiocarbamates in addition to their pesticidal properties can also inhibit biological oxidation of ammonium(nitrification) in soil. Metal [M=V(Ⅲ), Cr(Ⅲ), Mn(Ⅱ), Fe(Ⅲ), Ni(Ⅱ), Cu(Ⅱ), Zn(Ⅱ) and Co(Ⅱ)] diethyldithiocarbamates (DEDTC) were synthesized by the reaction of sodium diethyldithiocarbamate with metal chloride in dichloromethane/water mixture. These metal diethyldithiocarbamates were screened for their ability to inhibit nitrification at different concentrations(10 μg/g soil, 50 μg/g soil and 100 μg/g soil). With increasing concentration of the complex, capacity to retard nitrification increased but the extent of increase varied for different metals. At 100 μg/g soil, different complexes showed nitrification inhibition from 22 36% to 46 45%. Among the diethyldithiocarbamates tested, Zn(DEDTC) 2 proved to be the most effective nitrification inhibitor at 100 μg/g soil. Manganese, iron and chromium diethyldithiocarbamates also proved to be effective nitrification inhibitors than the others at 100 μg/g soil. The order of percent nitrification inhibition in soil by metal diethyldithiocarbamates was: Zn(Ⅱ) > Mn(Ⅱ) > Fe(Ⅲ) > Cr(Ⅲ) >V(Ⅲ) > Co(Ⅱ) > Ni(Ⅱ) > Cu(Ⅱ).

Reduction of N2O emissions by DMPP depends on the interactions of nitrogen sources(digestate vs. urea) with soil properties

The inhibition of nitrification by mixing nitrification inhibitors(NI)with fertilizers is emerging as an effective method to reduce fertilizer-induced nitrous oxide(N_(2)O)emissions.The additive 3,4-dimethylpyrazole phosphate(DMPP)apparently inhibits ammonia oxidizing bacteria(AOB)more than ammonia oxidizing archaea(AOA),which dominate the nitrification in alkaline and acid soil,respectively.However,the efficacy of DMPP in terms of nitrogen sources interacting with soil properties remains unclear.We therefore conducted a microcosm experiment using three typical Chinese agricultural soils with contrasting pH values(fluvo-aquic soil,black soil and red soil),which were fertilized with either digestate or urea in conjunction with a range of DMPP concentrations.In the alkaline fluvo-aquic soil,fertilization with either urea or digestate induced a peak in N_(2)O emission(60μg N kg^(-1)d^(-1))coinciding with the rapid nitrification within 3 d following fertilization.DMPP almost eliminated this peak in N_(2)O emission,reducing it by nearly 90%,despite the fact that the nitrification rate was only reduced by 50%.In the acid black soil,only the digestate induced an N_(2)O emission that increased gradually,reaching its maximum(20μg N kg^(-1)d^(-1))after 5–7 d.The nitrification rate and N_(2)O emission were both marginally reduced by DMPP in the black soil,and the N_(2)O yield(N_(2)O-N per NO2–+NO3–-N produced)was exceptionally high at 3.5%,suggesting that the digestate induced heterotrophic denitrification.In the acid red soil,the N_(2)O emission spiked in the digestate and urea treatments at 50 and 10μg N kg^(-1)d^(-1),respectively,and DMPP reduced the rates substantially by nearly 70%.Compared with 0.5%DMPP,the higher concentrations of DMPP(1.0 to 1.5%)did not exert a significantly(P<0.05)better inhibition effect on the N_(2)O emissions in these soils(either with digestate or urea).This study highlights the importance of matching the nitrogen sources,soil properties and NIs to achieve a high efficiency of N_(2)O emission reduction.

Recent Advances on the Technologies to Increase Fertilizer Use Efficiency

To increase fertilizer use efficiency （FUE） and to minimize its negative impact on environment have been the focal points in the world for a long time. It is very important to increase FUE in China for its relatively low FUE and serious losses of nutrients. Recent advances of the technologies to increase FUE are reviewed in this article. These include site-specific and real-time nitrogen management, non-destructive quick test of the nitrogen status of plants, new types of slow release and controlled release fertilizers, site-specific nutrient management, and use of urease inhibitor and nitrification inhibitor to decrease nitrogen losses. Future outlook in technologies related to FUE improvement is also discussed.

Evaluation of a crop rotation with biological inhibition potential to avoid N_(2)O emissions in comparison with synthetic nitrification inhibition

Agriculture has increased the release of reactive nitrogen to the environment due to crops’low nitrogen-use efficiency(NUE)after the application of nitrogen-fertilisers.Practices like the use of stabilized-fertilisers with nitrification inhibitors such as DMPP(3,4-dimethylpyrazole phosphate)have been adopted to reduce nitrogen losses.Otherwise,cover crops can be used in crop-rotation-strategies to reduce soil nitrogen pollution and benefit the following culture.Sorghum(Sorghum bicolor)could be a good candidate as it is drought tolerant and its culture can reduce nitrogen losses derived from nitrification because it exudates biological nitrification inhibitors(BNIs).This work aimed to evaluate the effect of fallow-wheat and sorghum cover crop-wheat rotations on N_(2)O emissions and the grain yield of winter wheat crop.In addition,the suitability of DMPP addition was also analyzed.The use of sorghum as a cover crop might not be a suitable option to mitigate nitrogen losses in the subsequent crop.Although sorghum–wheat rotation was able to reduce 22%the abundance of amoA,it presented an increment of 77%in cumulative N_(2)O emissions compared to fallow–wheat rotation,which was probably related to a greater abundance of heterotrophic-denitrification genes.On the other hand,the application of DMPP avoided the growth of ammonia-oxidizing bacteria and maintained the N_(2)O emissions at the levels of unfertilized-soils in both rotations.As a conclusion,the use of DMPP would be recommendable regardless of the rotation since it maintains NH_(4)^(+)in the soil for longer and mitigates the impact of the crop residues on nitrogen soil dynamics.

DMPP mitigates N_(2)O and NO productions by inhibiting ammonia-oxidizing bacteria in an intensified vegetable field under different temperature and moisture regimes

Vegetable soils with high nitrogen input are major sources of nitrous oxide(N_(2)O)and nitric oxide(NO),and incorporation of the nitrification inhibitor 3,4-dimethylpyrazole phosphate(DMPP)into soils has been documented to effectively reduce emissions.However,the efficiency of DMPP in terms of soil N_(2)O and NO mitigations varies greatly depending on soil temperature and moisture levels.Thus,further evaluations of DMPP efficiency in diverse environments are required to encourage widespread application.A laboratory incubation study(28 d)was established to investigate the interactive effects of DMPP,temperature(15,25,and 35?C),and soil moisture(55% and 80% of water-holding capacity(WHC))on net nitrification rate,N_(2)O and NO productions,and gene abundances of nitrifiers and denitrifiers in an intensive vegetable soil.Results showed that incubating soil with 1%DMPP led to partial inhibition of the net nitrification rate and N_(2)O and NO productions,and the reduction percentage of N_(2)O production was higher than that of NO production(69.3%vs.38.2%)regardless of temperature and soil moisture conditions.The increased temperatures promoted the net nitrification rate but decreased soil N_(2)O and NO productions.Soil moisture influenced NO production more than N_(2)O production,decreasing with the increased moisture level(80%).The inhibitory effect of DMPP on cumulative N_(2)O and NO productions decreased with increased temperatures at 55%WHC.Conversely,the inhibitory effect of DMPP on cumulative N_(2)O production increased with increased temperatures at 80%WHC.Based on the correlation analyses and automatic linear modeling,the mitigation of both N_(2)O and NO productions from the soil induced by DMPP was attributed to the decreases in ammonia-oxidizing bacteria(AOB)amoA gene abundance and NO_(2)^(-)-N concentration.Overall,our study indicated that DMPP reduced both N_(2)O and NO productions by regulating the associated AOB amoA gene abundance and NO_(2)^(-)-N concentration.These findings improve our insights regarding the implications of DMPP for N_(2)O and NO mitigations in vegetable soils under various climate scenarios.

Autotrophic and Heterotrophic Nitri?cation in a Highly Acidic Subtropical Pine Forest Soil

The occurrence of nitri?cation in some acidic forest soils is still a subject of debate.Identi?cation of main nitri?cation pathways in acidic forest soils is still largely unknown.Acidic yellow soil(Oxisol) samples were selected to test whether nitri?cation can occur or not in acidic subtropical pine forest ecosystems.Relative contributions of autotrophs and heterotrophs to nitri?cation were studied by adding selective nitri?cation inhibitor nitrapyrin.Soil NH^+_4-N concentrations decreased,but NO^-_3-N concentrations increased signi?cantly for the no-nitrapyrin control during the ?rst week of incubation,indicating that nitri?cation did occur in the acidic subtropical soil.The calculated net nitri?cation rate was 0.49 mg N kg^(-1)d^(-1)for the no-nitrapyrin control during the ?rst week of incubation.Nitrapyrin amendment resulted in a signi?cant reduction of NO^-_3-N concentration.Autotrophic nitri?cation rate averaged0.28 mg N kg^(-1)d^(-1)and the heterotrophic nitri?cation rate was 0.21 mg N kg^(-1)d^(-1)in the ?rst week.Ammonia-oxidizing bacteria(AOB) abundance increased slightly during incubation,but nitrapyrin amendment signi?cantly decreased AOB amo A gene copy numbers by about 80%.However,the ammonia-oxidizing archaea(AOA) abundance showed signi?cant increases only in the last 2weeks of incubation and it was also decreased by nitrapyrin amendment.Our results indicated that nitri?cation did occur in the present acidic subtropical pine forest soil,and autotrophic nitri?cation was the main nitri?cation pathway.Both AOA and AOB were the active biotic agents responsible for autotrophic nitri?cation in the acidic subtropical pine forest soil.

Yield-scaled nitrous oxide emissions from nitrogen-fertilized croplands in China: A meta-analysis of contrasting mitigation scenarios

Nitrogen(N) losses in cropland resulting from the application of synthetic fertilizers decrease crop productivity and exacerbate environmental pollution.Mitigation measures, such as reduction in N fertilizer application rates, can have unintentional adverse effects on crop yield. We conducted a meta-analysis of soil N_(2)O emissions from agricultural fields across China under contrasting mitigation scenarios as a novel approach to identify the most effective strategy for the mitigation of emissions of N_(2)O derived from N fertilizer use in China. Current standard agricultural practice was used as a baseline scenario(BS), and 12 potential mitigation scenarios(S1–S12) were derived from the available literature and comprised single and combinations of management scenarios that accounted for crop yield. Mitigation scenarios S6(nitrification inhibitor 3,4-dimethylpyrazole phosphate) and S11(20% reduction in N application rate plus nitrification inhibitor dicyandiamide) in maize, rice, and wheat crops led to an average 56.0% reduction in N_(2)O emissions at the national level, whereas scenario S4(nitrification inhibitor dicyandiamide) led to yield optimization, with a 14.0% increase for maize and 8.0% increase for rice as compared to the BS. Implementation of these most effective mitigation scenarios(S4, S6, and S11) might help China, as a signatory to the 2015 United Nations Framework Convention on Climate Change(Paris Agreement), to achieve a 30% reduction in N_(2)O emissions by 2030.

Nitrapyrin effectiveness in reducing nitrous oxide emissions decreases at low doses of urea in an Andosol

In the tropics,frequent nitrogen(N)fertilization of grazing areas can potentially increase nitrous oxide(N_(2)O)emissions.The application of nitrification inhibitors has been reported as an effective management practice for potentially reducing N loss from the soil-plant system and improving N use efficiency(NUE).The aim of this study was to determine the effect of the co-application of nitrapyrin(a nitrification inhibitor,NI)and urea in a tropical Andosol on the behavior of N and the emissions of N_(2)O from autotrophic and heterotrophic nitrification.A greenhouse experiment was performed using a soil(pH 5.9,organic matter content 78 g kg^(-1),and N 5.6 g kg^(-1))sown with Cynodon nlemfuensis at 60%water-filled pore space to quantify total N_(2)O emissions,N_(2)O derived from fertilizer,soil ammonium(NH_(4)^(+))and nitrate(NO_(3)^(-)),and NUE.The study included treatments that received deionized water only(control,NI).No significant differences were observed in soil NH_(4)^(+)content between the UR and UR+NI treatments,probably because of soil mineralization and NO_(3)^(-)produced by heterotrophic nitrification,which is not effectively inhibited by nitrapyrin.After 56 d,N_(2)O emissions in UR(0.51±0.12 mg N_(2)O-N concluded that the soil organic N mineralization and heterotrophic nitrification are the main processes of NH_(4)^(+)and NO_(3)^(-)production.Additionally,it was found that N_(2)O emissions were partially a consequence of the direct oxidation of the soil's organic N via heterotrophic nitrification coupled to denitrification.Finally,the results suggest that nitrapyrin would likely exert significant mitigation on N_(2)O emissions only if a substantial N surplus exists in soils with high organic matter content.