硝化抑制剂对稻田土壤N2O排放和硝化、反硝化菌数量的影响

【目的】本研究旨在明确硝化抑制剂对稻田土壤氮素周转的影响,探讨抑制剂提高氮肥利用率及微生物响应机理。【方法】以草甸黑土发育的水稻土为研究对象,进行了两组培养试验 (25℃),培养周期均为150天。共设4个处理:1) 不施肥 (CK);2) 单施尿素 (Urea);3) 尿素 + 双氰胺 (Urea + DCD);4) 尿素 + 3, 4-二甲基吡唑磷酸盐 (Urea + DMPP)。一组试验从培养第1天起,抽取气体样品,用气相色谱法测定N₂O排放量。另一组试验从培养第1天直到结束,取土样测定氨氧化细菌和氨氧化古菌数量,采用荧光定量PCR等技术测定nirK基因和nirS基因拷贝数,用常规方法测定土壤无机氮含量。【结果】施用尿素显著增加了N₂O排放量,其中85%的N₂O排放发生在培养开始后的前两周内。Urea + DMPP处理土壤NH₄+浓度在前23天稳定在较高水平,与Urea处理相比,N₂O减排率为78.3%,Urea + DCD处理为21.6%。Urea + DMPP处理排放系数为0.05%,Urea + DCD为0.18%,Urea + DMPP处理显著低于Urea + DCD处理。施用尿素培养,土壤氨氧化细菌 (AOB) 数量显著增加而氨氧化古菌 (AOA) 数量则显著减少。添加DCD和DMPP能显著抑制AOB的数量,但对AOA没有影响。培养第3、30和90天,Urea + DMPP处理土壤中的AOB数量显著低于Urea + DCD处理的30%、56%和60%。对于反硝化细菌来说,所有处理中的nirK基因拷贝数均显著高于nirS基因拷贝数。添加DMPP在培养第3和30天显著减少了含nirK和nirS基因的反硝化细菌数量,而添加DCD对两类反硝化细菌数量无明显作用。【结论】东北黑土水稻生产中,硝化抑制剂DMPP降低N₂O排放量和排放系数的效果显著好于DCD,因为DMPP在培养后的30天内,可以显著抑制氨氧化细菌繁衍,降低反硝化细菌数量,从而起到减少N₂O排放、提高肥料利用率的作用。     

生物质炭对酸性菜地土壤N2O排放及相关功能基因丰度的影响

【目的】 生物质炭显著影响土壤氧化亚氮 (N₂O) 排放,但关于其相关微生物机理的研究相对匮乏,尤其是生物质炭对酸性菜地土壤N₂O排放的微生物作用机理。本文通过研究氮肥配施生物质炭对酸性菜地土壤N₂O排放以及硝化和反硝化过程相关功能基因丰度的影响,探讨酸性菜地土壤N₂O排放与功能基因丰度的关系,阐释生物质炭对酸性菜地土壤试验N₂O排放的微生物作用机理。 【方法】 在田间一次性施入生物质炭 40 t/hm2,试验连续进行了3年,共9茬蔬菜。设置4个处理:对照 (CK)、氮肥 (N)、生物质炭 (Bc) 和氮肥 + 生物质炭 (N + Bc)。在施用后第三年,采集土壤样品进行室内培养,应用荧光定量PCR技术检测硝化过程氨氧化古菌 (AOA)、氨氧化细菌 (AOB) 功能基因amoA和反硝化过程亚硝酸还原酶基因 (nirK、nirS) 以及N₂O还原酶基因 (nosZ) 等相关功能基因丰度,同时监测土壤pH值、无机氮 (铵态氮、硝态氮) 含量及N₂O排放。 【结果】 与CK相比,生物质炭 (Bc) 处理的土壤有机碳 (SOC) 提高了27.1%,总氮 (TN) 提高了8.2%,amoA-AOB基因丰度显著降低了11.0%,nosZ基因丰度增加了21.2% (P < 0.05),N ₂O排放没有显著变化 (P > 0.05)。与CK相比,施用氮肥 (N) 显著降低土壤pH ( P < 0.05),显著增加土壤无机氮含量、 nirK、nirS和nosZ功能基因丰度以及土壤N₂O累积排放量 (P < 0.05)。与N处理相比,生物质炭与氮肥联合施用 (N + Bc) 处理显著增加 amoA-AOA、amoA-AOB、nirK、nirS和nosZ基因丰度,增幅分别为68.1%、39.3%、21.1%、19.8%、48.4% (P < 0.05),但 ( nirK + nirS)/nosZ的比值降低,同时N₂O累积排放量显著降低33.3% (P < 0.05)。室内培养期间N ₂O排放峰出现在1～5 d,N和N+Bc处理排放速率分别为 N 1.70 × 103和1.76 × 103 ng/(kg·h)。相关分析结果显示,N₂O排放速率与氧化亚氮还原酶的标记基因nosZ基因拷贝数 (P < 0.05)、NH ₄+-N含量 (P < 0.01) 呈显著正相关,与pH呈显著负相关 ( P < 0.01)。 【结论】 在菜地生态系统中氮肥和生物质炭联合施用可以有效缓解菜地土壤酸化,减少菜地土壤N₂O排放,主要归因于反硝化作用nosZ基因丰度增加,(nirK + nirS)/nosZ比值降低。      

生物硝化抑制剂对黔西南黄壤硝化作用及N2O排放的影响

为探索生物硝化抑制剂对贵州黔西南地区黄壤硝化作用及氧化亚氮(N2O)排放的影响,通过三周的室内培养试验,研究两种生物硝化抑制剂对羟基苯丙酸甲酯(MHPP)和丁香酸(SA)对黄壤中的无机态氮素含量、氨氧微生物功能基因以及N2O排放量的影响。结果表明,与对照CK相比,MHPP和SA在黄壤上均能明显抑制硝化作用,对土壤硝化速率的抑制率分别为6%~43%和5%~51%。MHPP和SA均抑制了黄壤氨氧化古菌AOA(12%~22%,27%~41%)与氨氧化细菌AOB(6%~19%,26%~46%)amoA基因的丰度。整个培养期内,黄壤的硝态氮含量与AOB的amoA基因丰度显著正相关,而与AOA的amoA基因丰度无显著相关,表明AOB对黄壤硝化作用起了主导作用。在N2O排放方面,MHPP和SA分别显著抑制了黄壤51%和21%的N2O排放积累量,MHPP的减排效果优于SA。MHPP降低了黄壤N2O排放的峰值,而SA主要延缓了黄壤N2O产生高峰的出现。总之,生物硝化抑制剂MHPP和SA在贵州黔西南黄壤上具有氮肥减施增效的潜力,这为今后烤烟新型绿色专用肥的开发提供了理论依据。     

有机和无机肥配比对黄褐土硝化和反硝化微生物丰度及功能的影响

【目的】 土壤硝化与反硝化作用是氮循环的两个关键环节,本文研究不同比例的有机、无机肥配施对硝化和反硝化进程产生的影响,为高效施肥提供理论基础。 【方法】 在安徽农业大学农翠园试验基地的黄褐土上进行了小麦–玉米轮作田间试验。试验以不施氮肥为对照 (CK),在小麦、玉米总施氮量相同的条件下,设置5个处理,分别为单施无机肥 (T1)、无机肥∶有机肥 = 2∶1 (T2)、无机肥∶有机肥 = 1∶1 (T3)、无机肥∶有机肥 = 1∶2 (T4)、单施有机肥 (T5)。在小麦拔节期,取0—20 cm土壤样品,利用荧光定量PCR技术测定反硝化和氨氧化微生物丰度,并结合反硝化能力、N₂O/(N₂O+N₂) 产物比、土壤呼吸、硝化势和氨氧化细菌 (AOB) 与古菌 (AOA) 对硝化势相对贡献率的测定,分析江淮地区长期有机和无机肥配施对黄褐土硝化、反硝化微生物丰度及其功能的影响。 【结果】 单施无机肥或有机肥处理的硝化势均高于不同配比处理。与添加有机肥相比,增施无机肥会显著增加AOA的丰度和硝化贡献率。在反硝化方面,反硝化能力和土壤呼吸随着有机肥投入量的增加而增加,单施有机肥处理显著高于其它处理。nirS和nosZ型反硝化菌丰度随着有机肥的增加而增加,而nirK型反硝化菌丰度呈减少趋势。相关分析表明,反硝化能力与nirS型、nosZ型反硝化菌丰度、有机质和可溶性有机碳含量极显著正相关,与nirK相关性不强。 【结论】 与单施无机肥或有机肥处理相比,有机和无机肥适当配施可降低土壤硝化势,并能调控AOA和AOB在硝化过程中的作用,有效地降低土壤反硝化损失。      

不同形态氮添加对毛竹林土壤N2O排放的影响

  【目的】  氧化亚氮(N₂O)排放是亚热带地区氮损失的主要途径,我们研究了不同形态含氮化合物对土壤N₂O排放的影响。  【方法】  以毛竹(Phyllostachys edulis)林土壤为研究对象进行了室内培养试验。设置土壤中添加KNO₃、NH₄NO₃、NH₄Cl、KCl处理,以去离子水作为对照(CK),在25oC黑暗条件下培养。在培养0.5 h,1、3、5、7、14、28、60天,测定土壤N₂O排放速率,铵态氮(NH₄+-N)、硝态氮(NO₃?-N)、可溶性有机碳(DOC)和水溶性氮(WSN)含量,采用荧光定量PCR技术测定了土壤氨氧化细菌(ammonia-oxidizing bacteria, AOB)、氨氧化古菌(ammonia-oxidizing archaea, AOA)、nirS、nirK、nosZⅠ、nosZⅡ基因丰度。  【结果】  培养第60天,氮添加与KCl添加处理均显著增加了土壤DOC含量,NH₄NO₃、NH₄Cl处理显著增加了WSN含量,但显著降低了土壤pH。氮添加及KCl添加处理均增加了土壤AOA、AOB、nirK基因丰度,降低了nosZⅠ、nosZⅡ基因丰度。氮添加处理N₂O排放速率均在培养第14天达到峰值,且相较于CK处理均增加了N₂O累积排放量,KNO₃、NH₄NO₃、NH₄Cl和KCl处理累积排放量的增幅分别为524.3%、771.1%、652.7%、98.6%。N₂O排放速率与NO₃?、WSN、nirK基因丰度呈显著正相关,而与pH、nosZⅠ、nosZⅡ基因丰度呈显著负相关。  【结论】  铵态氮添加能显著促进毛竹林土壤N₂O的排放,其效果高于硝态氮,NH₄NO₃作为混合氮,外源性NH₄+-N、NO₃?-N同时输入对土壤N₂O排放的促进作用比单独添加NH₄+-N、NO₃?-N更显著,但并未出现叠加效应。     

长期施肥对土nirS型反硝化细菌的影响及其与N2O排放的关系

土壤反硝化作用是土壤N₂O产生的重要过程,亚硝酸盐还原酶(NIR)催化的亚硝态氮(NO^-₂)还原为一氧化氮(NO)是反硝化作用的关键环节,研究长期施肥对反硝化微生物的影响及其与N₂O排放的关系对于全面理解土壤反硝化过程具有重要意义。基于28年的旱作雨养长期施肥试验,通过常规监测、定量PCR和高通量测序等探讨了长期不同施肥(不施肥CK、偏施肥的单施氮肥N和氮钾配施NK、以及氮磷钾平衡施肥NPK)下土N₂O排放和nirS反硝化细菌群落特征及两者之间的关系。结果表明:长期化肥施用(N,NK和NPK)均显著提高了N₂O累积排放量,其中平衡施肥(NPK)最高。长期化肥施用对nirS基因丰度和nirS型反硝化细菌的α-多样性无显著影响,但长期平衡施用化肥提高了uncultured_bacterium_2303和Rhodanobacter_sp._D206a的相对丰度,降低了unclassified_k_norank_d_Bacteria和unclassified_p_Proteobacteria的相对丰度,从而改变了nirS型反硝化细菌的群落结构组成。雨养旱作条件下,土壤有机碳(SOC)、全氮(TN)、有效磷(AP)和pH等土壤性质是土nirS型反硝化细菌群落结构组成变化的主要影响因素。土nirS型反硝化细菌群落结构组成对土壤N₂O排放具有显著影响,而nirS基因丰度和nirS型反硝化细菌多样性并没有显著影响。     

硝化抑制剂对毛竹林土壤N_2O排放和氨氧化微生物的影响

为了探索硝化抑制剂在毛竹生产中的施用技术,通过培养试验研究3,4-二甲基吡唑磷酸盐(DMPP)和双氰胺(DCD)两种硝化抑制剂对毛竹林施用尿素后土壤N2O排放、氮素转化和相关氨氧化细菌(AOB)、氨氧化古菌(AOA)群落结构和丰度的影响。试验设(1)对照(CK)、(2)单施尿素(Urea)、(3)尿素+1%DMPP(DMPP占总N的1%,下同);(4)尿素+1.5%DMPP;(5)尿素+10%DCD;(6)尿素+15%DCD等6个处理,测定N2O的排放动态以及气体排放转折点时的土壤特征指标。结果表明:与单施尿素相比,160 d的时间内两种DMPP用量处理的土壤N2O累积排放减排幅度均为54%,而10%DCD和15%DCD处理的土壤分别减少28%和41%。DMPP和DCD处理50 d和90 d时土壤的NH4+-N含量均显著高于(p0.05)单施尿素处理,而NO3--N含量和表观硝化率则恰好相反,但两种抑制剂间无差异。DMPP处理的AOB群落结构的变化从10 d开始显现,至50 d和90 d时仍保持明显的抑制状态,而DCD处理则至90 d时抑制作用基本消失。单施尿素AOB功能基因(amo A)的丰度均显著高于硝化抑制剂处理(90 d时尿素+10%DCD处理除外);在整个培养期内,尿素和对照土壤的AOA群落结构相似,硝化抑制剂反而增加了AOA功能基因的丰度,表明硝化抑制剂对AOA丰度无明显抑制作用。即两种硝化抑制剂主要通过抑制AOB起作用;调节土壤p H至中性范围,并在1%DMPP施用条件下,硝化抑制剂的抑制效果最显著。     

长期施肥对棕壤氨氧化细菌和古菌丰度的影响

【目的】氨氧化是氮转化过程的限速步骤,其由氨氧化微生物所驱动。本研究旨在探明 37 年玉米–大豆轮作施肥条件下影响棕壤氨氧化微生物丰度的主要影响因子及变化规律。【方法】以沈阳农业大学棕壤肥料长期定位试验耕层土壤 (0—20 cm) 为材料,选取其中 9 个施肥处理进行取样分析:不施肥 (CK)、低量氮肥 (N₁)、高量氮肥 (N₂)、氮磷肥 (N₁P)、氮磷钾肥 (N₁PK)、高量有机肥 (M₂)、高量有机肥 + 低量氮肥 (M₂N₁)、高量有机肥 + 氮磷肥 (M₂N₁P)、高量有机肥 + 氮磷钾肥 (M₂N₁PK)。采用实时荧光定量 PCR 技术测定其氨氧化微生物丰度,通过对土壤基本化学性质和氨氧化微生物丰度的冗余分析找出影响氨氧化微生物丰度的主要因素。【结果】施用有机肥处理的土壤 pH、有机质、全氮、碱解氮、速效钾、速效磷、铵态氮、硝态氮含量明显高于不施肥和单施化肥处理。各施肥处理土壤有机质、全氮、碱解氮、速效钾、速效磷的含量总体呈现有机肥处理 > 化肥处理 > CK;与不施肥处理 (CK) 相比,单施化肥处理显著降低了土壤 pH 值,施用有机肥处理显著提高了土壤 pH 值,其中 N₂ 处理的土壤 pH 最低,M₂ 处理的土壤 pH 最高。不同施肥处理氨氧化细菌 (AOB) 的丰度为 0.94 × 106～5.77 × 106 copies/g 干土,氨氧化古菌 (AOA) 的丰度为 3.56 × 106～1.22 × 107 copies/g 干土;施用有机肥处理 AOB 和 AOA 丰度显著高于不施肥和单施化肥处理,其中 M₂ 处理的 AOB 和 AOA 丰度最高,单施氮肥处理的 AOB 和 AOA 丰度最低。冗余分析 (RDA) 表明,影响棕壤 AOB 和 AOA 丰度的主要环境因子有土壤 pH、有机质、全氮、碱解氮、速效磷、速效钾,且与 AOB 和 AOA 丰度呈正相关关系。【结论】长期轮作施肥显著改变了棕壤的化学性质,从而对氨氧化微生物的丰度产生了显著影响。长期施用有机肥显著提高了土壤养分含量及 AOB 和 AOA 的丰度,对维持土壤氨氧化微生物的数量起到十分重要的作用;同时试验结果也为今后通过改变土壤 pH、有机质、全氮、碱解氮、速效磷、速效钾等性质对 AOB 和 AOA 进行调节提供了依据。     

脲酶抑制剂与硝化抑制剂对稻田土壤硝化、反硝化功能菌的影响

[目的]在农业生产中,脲酶抑制剂(urease inhibitor,UI)与硝化抑制剂(nitrification inhibitor,NI)常作为氮肥增效剂来提高肥料利用率。本文研究了在我国南方红壤稻田施用脲酶抑制剂与硝化抑制剂后,土壤中氨氧化细菌(ammonia oxidizing bacteria,AOB)、氨氧化古菌(ammonia-oxidizing archaea,AOA)以及反硝化细菌的丰度以及群落结构的变化特征,旨在揭示抑制剂的作用机理及其对土壤环境的影响。[方法]试验在我国南方红壤稻田进行,共设5个处理:1)不施氮肥(CK);2)尿素(U);3)尿素+脲酶抑制剂(U+UI);4)尿素+硝化抑制剂(U+NI);5)尿素+脲酶抑制剂+硝化抑制剂(U+UI+NI),3次重复。脲酶抑制剂与硝化抑制剂分别为NBPT[N-(n-butyl)thiophosphrictriamide,N-丁基硫代磷酰三胺]和DMPP(3,4-dimethylpyrazole phosphate,3,4-二甲基吡唑磷酸盐)。通过荧光定量PCR(Real-time PCR)研究水稻分蘖期与孕穗期抑制剂对三类微生物标记基因拷贝数的影响,并分析土壤铵态氮、硝态氮与三种菌群丰度的相关性;利用变性梯度凝胶电泳(DenaturingGradient Gel Electrophoresis,DGGE)分析抑制剂对土壤AOB、AOA以及反硝化细菌群落结构的影响,并对优势菌群进行系统发育分析。[结果]1)荧光定量PCR结果表明,施用氮肥对两个时期土壤中AOB的amoA基因与反硝化细菌nirK基因的拷贝数均有显著提高,而对AOA的amoA基因始终没有明显影响;AOB与nirK反硝化细菌的丰度与两个时期的铵态氮含量、分蘖期的硝态氮含量呈极显著正相关,与孕穗期的硝态氮含量相关性不显著;DMPP仅在分蘖期显著减少了AOB的amoA基因拷贝数,表明DMPP主要通过限制AOB的生长来抑制稻田土壤硝化过程;NBPT对三类微生物的丰度无明显影响;2)DGGE图谱表明,在分蘖期与孕穗期,施用氮肥均明显增加了图谱中AOB的条带数,而对AOA却没有明显影响;氮肥明显增加了孕穗期反硝化细菌的条带数;与氮肥的影响相比,抑制剂NBPT与DMPP对AOA、AOB以及反硝化菌的群落结构影响甚微;系统发育分析结果表明,与土壤中AOB的优势菌群序列较为接近的有亚硝化单胞菌和亚硝化螺菌。[结论]在南方红壤稻田中,施入氮肥可显著提高AOB与反硝化细菌的丰度,明显影响两种菌群的群落结构,而AOA较为稳定;NBPT对三类微生物的群落结构丰度无明显影响;硝化抑制剂DMPP可抑制AOB的生长但仅表现在分蘖期,这可能是其缓解硝化反应的主要途径;这也说明二者对土壤生态环境均安全可靠。     

水钠锰矿提高红壤性水稻土N2O释放速率和氨氧化细菌amoA基因丰度

  【目的】  土壤中的氧化亚氮 (N₂O) 来源于硝化与反硝化作用，锰可与硝化或反硝化作用产物反应产生N₂O或氮气，已有研究表明土壤中锰含量高会影响硝化作用。因此，本试验以水钠锰矿 (KMnO₂·H₂O) 与土壤硝化作用与反硝化作用的生物化学耦合反应为切入点，研究水钠锰矿的添加对土壤N₂O释放速率及微生物的影响，进一步认识N₂O释放与土壤环境因子的相互关系。  【方法】  以红壤性水稻土为供试土壤，通过微宇宙培养试验，在土壤中添加不同质量百分比的水钠锰矿 (0%、0.1%、0.3%、0.7%、1.5%)，预培养7 天后，加入硫酸铵N 100 mg/kg继续培养14天。在培养第1、3、7、14天，采用气密性注射器抽取10 mL气体样品，气相色谱仪测定N₂O含量；同时取土壤样品，比色法测定铵态氮与硝态氮含量。培养结束时，测定土壤pH，采用实时荧光定量PCR测定土壤16S rDNA与氨氧化细菌 (AOB) amoA基因拷贝数，高通量测序技术分析微生物群落组成及多样性。  【结果】  水钠锰矿提高了土壤N₂O释放速率，增加了土壤N₂O累积释放量，以添加0.1%水钠锰矿的N₂O累积释放量最高，添加1.5%的最低。土壤铵态氮含量随培养时间的延长而迅速降低，硝态氮含量则迅速增加。水钠锰矿显著提高了土壤pH与表观N₂O产量 (N₂O-N/NO₃?-N)，pH随着水钠锰矿添加量的增加整体提高，N₂O-N/NO₃?-N则随着水钠锰矿添加量的增加呈降低趋势。适量水钠锰矿显著增加了土壤细菌16S rDNA与氨氧化细菌 (AOB) amoA基因拷贝数，并显著提高了土壤16S rDNA与AOB amoA基因拷贝数的比值，但随着水钠锰矿添加量的增加，细菌16S rDNA和AOB amoA基因拷贝数的增加量整体降低；放线菌、变形菌与拟杆菌是所有处理中的优势菌门，通过非度量多维尺度分析发现不同处理间的微生物群落结构差异显著，未添加水钠锰矿处理与添加水钠锰矿1.5%处理的微生物群落结构差异最大，其他处理的微生物群落结构介于两者之间。  【结论】  土壤中添加0.1%质量比的水钠锰矿，可以通过增加AOB的数量促进红壤性水稻土N₂O的释放，显著影响微生物物种丰度与群落结构。但水钠锰矿高添加量处理对AOB的刺激作用减弱，因此，应将土壤锰含量作为影响土壤N₂O释放的因素加以考虑。     

Decreased nitrous oxide emissions associated with functional microbial genes under bio-organic fertilizer application in vegetable fields

Bio-organic fertilizers enriched with plant growth-promoting microbes(PGPMs)have been widely used in crop fields to promote plant growth and maintain soil microbiome functions.However,their potential effects on N₂O emissions are of increasing concern.In this study,an in situ measurement experiment was conducted to investigate the effect of organic fertilizer containing Trichoderma guizhouense(a plant growth-promoting fungus)on soil N₂O emissions from a greenhouse vegetable field.The following four treatments were used:no fertilizer(control),chemical fertilizer(NPK),organic fertilizer derived from cattle manure(O),and organic fertilizer containing T.guizhouense(O+T,referring to bio-organic fertilizer).The abundances of soil N cycling-related functional genes(amoA)from ammonium-oxidizing bacteria(AOB)and archaea(AOA),as well as nirS,nirK,and nosZ,were simultaneously determined using quantitative PCR(qPCR).Compared to the NPK plot,seasonal total N₂O emissions decreased by 11.7%and 18.7%in the O and O+T plots,respectively,which was attributed to lower NH₄⁺-N content and AOB amoA abundance in the O and O+T plots.The nosZ abundance was significantly greater in the O+T plot,whilst the AOB amoA abundance was significantly lower in the O+T plot than in the O plot.Relative to the organic fertilizer,bio-organic fertilizer application tended to decrease N₂O emissions by 7.9%and enhanced vegetable yield,resulting in a significant decrease in yield-scaled N₂O emissions.Overall,the results of this study suggested that,compared to organic and chemical fertilizers,bio-organic fertilizers containing PGPMs could benefit crop yield and mitigate N₂O emissions in vegetable fields.     

生物质炭对土壤N2O消耗的影响及其微生物影响机理

生物质炭在温室气体减排方面具有很大的发展前景,它不仅能实现固碳,对于在大气中停留时间长且增温潜势大的N₂O也能发挥积极作用。本研究采用室内厌氧培养试验,按照生物质炭与土壤质量比（0、1%和5%）加入一定量生物质炭,土壤重量含水率控制在20%。利用Robotized Incubation平台实时检测N₂O和N₂浓度变化,通过测定土壤中反硝化功能基因丰度（nirK、nirS、nosZ）分析生物质炭对N₂O消耗的影响及其微生物方面的影响机理。结果表明：经过20 h厌氧培养后,0生物质炭处理的反硝化功能基因丰度（基因拷贝数·g^-1）分别为6.80×10⁷（nirK）、5.59×10⁸（nirS）和1.22×10⁸（nosZ）。与0生物质炭处理相比,1%生物质炭处理的nirS基因丰度由最初的2.65×10⁸基因拷贝数·g^-1升至7.43×10⁸基因拷贝数·g^-1,nosZ基因丰度则提高了一个数量级,由4.82×10⁷基因拷贝数·g^-1升至1.50×10⁸基因拷贝数·g^-1,然而nirK基因丰度并无明显变化;5%生物质炭处理的反硝化功能基因丰度并未发生显著变化。试验结束时,添加生物质炭处理的N₂/（N₂O+N₂）比值也明显高于0生物质炭处理。相关性分析结果表明,nirS基因丰度和nosZ基因丰度均与N₂O浓度在0.01水平上显著相关。试验末期nirS基因丰度和nosZ基因丰度均随着N₂O浓度的降低而升高。因此在本试验中,添加1%生物质炭可显著提高nirS和nosZ基因型反硝化细菌的丰度,增大N₂/（N₂O+N₂）比值,促进N₂O彻底还原成N₂。生物质炭对于N₂O主要影响机理是增大了可以还原氧化亚氮的细菌活性,促进完全反硝化。     

Simulated animal trampling of a free‐draining stony soil stimulated denitrifier growth and increased nitrous oxide emissions

Winter forage grazing systems in New Zealand cause compaction of soil by grazing animals, especially when the soil is wet. However, there is little information on the effects of animal trampling on denitrifiers in soil, despite their importance for N₂O production. Here, we report a field study of the abundance of the denitrifying genes nirS, nirK, and nosZ and N₂O emissions following the application of dairy cow urine in a free‐draining stony soil. Importantly, we found that simulated animal trampling altered some of the denitrifying microbial communities, thus leading to increased N₂O emissions. Over the 111 day measurement period, the abundance of nitrite (NO₂^?)‐reducing nirS gene copy numbers increased significantly by 87% in the trampled soil with urine (P < 0.01) and increased by 40% in the trampled soil without urine (P < 0.05), but the nirS gene abundance did not change significantly in the nontrampled soil. The abundance of NO₂^? reducing nirK gene copy numbers was not affected by trampling, but increased significantly following urine application. The abundance of N₂O‐reducing nosZ clade I and nosZ clade II gene copy numbers increased significantly in the trampled soil, but did not change significantly in the nontrampled soil. N₂O emissions from the trampled soil were about twice that from the nontrampled soil without urine (1.20 and 0.62 kg N₂O‐N per ha, respectively) and about eight times greater (6.24 kg N₂O‐N per ha) than from nontrampled soil (0.80 kg N₂O‐N per ha) when urine was applied. These results strongly suggest that animal trampling during winter forage grazing can have a major impact on denitrifying communities in soil, which in turn stimulate greater denitrification with increased N₂O emissions.     

Linkage of N2O emissions to the abundance of soil ammonia oxidizers and denitrifiers in purple soil under long-term fertilization

Abstract

Microbial nitrification and denitrification are responsible for the majority of soil nitrous (N₂O) emissions. In this study, N₂O emissions were measured and the abundance of ammonium oxidizers and denitrifiers were quantified in purple soil in a long-term fertilization experiment to explore their relationships. The average N₂O fluxes and abundance of the amoAgene in ammonia-oxidizing bacteria during the observed dry season were highest when treated with mixed nitrogen, phosphorus and potassium fertilizer (NPK) and a single N treatment (N) using NH₄HCO₃as the sole N source; lower values were obtained using organic manure with pig slurry and added NPK at a ratio of 40%:60% (OMNPK),organic manure with pig slurry (OM) and returning crop straw residue plus synthetic NH₄HCO₃fertilizer at a ratio of 15%:85% (SRNPK). The lowest N₂O fluxes were observed in the treatment that used crop straw residue(SR) and in the control with no fertilizer (CK). Soil NH₄⁺provides the substrate for nitrification generating N₂O as a byproduct. The N₂O flux was significantly correlated with the abundance of the amoA gene in ammonia-oxidizing bacteria (r = 0.984, p < 0.001), which was the main driver of nitrification. During the wet season, soil nitrate (NO₃^?) and soil organic matter (SOC) were found positively correlated with N₂O emissions (r = 0.774, p = 0.041 and r = 0.827, p = 0.015, respectively). The nirS gene showed a similar trend with N₂O fluxes. These results show the relationship between the abundance of soil microbes and N₂O emissions and suggest that N₂O emissions during the dry season were due to nitrification, whereas in wet season, denitrification might dominate N₂O emission.     

Tillage system affects fertilizer-induced nitrous oxide emissions

Since the development of effective N₂O mitigation options is a key challenge for future agricultural practice, we studied the interactive effect of tillage systems on fertilizer-derived N₂O emissions and the abundance of microbial communities involved in N₂O production and reduction. Soil samples from 0–10 cm and 10–20 cm depth of reduced tillage and ploughed plots were incubated with dairy slurry (SL) and manure compost (MC) in comparison with calcium ammonium nitrate (CAN) and an unfertilized control (ZERO) for 42 days. N₂O and CO₂ fluxes, ammonium, nitrate, dissolved organic C, and functional gene abundances (16S rRNA gene, nirK, nirS, nosZ, bacterial and archaeal amoA) were regularly monitored. Averaged across all soil samples, N₂O emissions decreased in the order CAN and SL (CAN?=?748.8?±?206.3, SL?=?489.4?±?107.2 μg kg^?1) followed by MC (284.2?±?67.3 μg kg^?1) and ZERO (29.1?±?5.9 μg kg^?1). Highest cumulative N₂O emissions were found in 10–20 cm of the reduced tilled soil in CAN and SL. N₂O fluxes were assigned to ammonium as source in CAN and SL and correlated positively to bacterial amoA abundances. Additionally, nosZ abundances correlated negatively to N₂O fluxes in the organic fertilizer treatments. Soils showed a gradient in soil organic C, 16S rRNA, nirK, and nosZ with greater amounts in the 0–10 than 10–20 cm layer. Abundances of bacterial and archaeal amoA were higher in reduced tilled soil compared to ploughed soils. The study highlights that tillage system induced biophysicochemical stratification impacts net N₂O emissions within the soil profile according to N and C species added during fertilization.     

Abundance of transcripts of functional gene reflects the inverse relationship between CH<Subscript>4</Subscript> and N<Subscript>2</Subscript>O emissions during mid-season drainage in acidic paddy soil

Agricultural management significantly affects methane (CH₄) and nitrous oxide (N₂O) emissions from paddy fields. However, little is known about the underlying microbiological mechanism. Field experiment was conducted to investigate the effect of the water regime and straw incorporation on CH₄ and N₂O emissions and soil properties. Quantitative PCR was applied to measure the abundance of soil methanogens, methane-oxidising bacteria, nitrifiers, and denitrifiers according to DNA and mRNA expression levels of microbial genes, including mcrA, pmoA, amoA, and nirK/nirS/nosZ. Field trials showed that the CH₄ and N₂O flux rates were negatively correlated with each other, and N₂O emissions were far lower than CH₄ emissions. Drainage and straw incorporation affected functional gene abundance through altered soil environment. The present (DNA-level) gene abundances of amoA, nosZ, and mcrA were higher with straw incorporation than those without straw incorporation, and they were positively correlated with high concentrations of soil exchangeable NH₄⁺ and dissolved organic carbon. The active (mRNA-level) gene abundance of mcrA was lower in the drainage treatment than in continuous flooding, which was negatively correlated with soil redox potential (Eh). The CH₄ flux rate was significantly and positively correlated with active mcrA abundance but negatively correlated with Eh. The N₂O flux rate was significantly and positively correlated with present and active nirS abundance and positively correlated with soil Eh. Thus, we demonstrated that active gene abundance, such as of mcrA for CH₄ and nirS for N₂O, reflects the contradictory relationship between CH₄ and N₂O emissions regulated by soil Eh in acidic paddy soils.     

氮肥水平对稻田细菌群落及N2O排放的影响

作为土壤氮素转化的驱动者,微生物群落结构关系着稻田氮素利用及温室气体N_2O排放等问题。本研究分别基于高通量测序和荧光定量PCR技术,分析了不同氮肥水平[CK(不施氮)、N(施N 180 kg·hm-2)、2/3N(施N 120 kg·hm-2)、1/3N(施N 60 kg·hm-2)]下稻田细菌群落及硝化反硝化关键微生物功能基因丰度的变化。结果显示:氮肥水平提高增加了稻田细菌物种丰富度Chao1指数和群落多样性Shannon指数,改变了细菌群落组成,其中与硝化作用相关的硝化螺菌门Nitrospirae和嗜酸的醋杆菌门Acidobacteria的相对丰度随氮肥水平提高而增加,但甲烷氧化菌Methylosinus的相对丰度随氮肥水平提高而降低。氮肥水平对稻田硝化作用关键微生物氨氧化细菌amo A基因丰度的影响较大,0~5 cm和10~20 cm深度土层中的amo A基因丰度均随氮肥用量增加而提高;反硝化作用关键微生物功能基因nir S、qno B和nos Z的丰度在不施肥处理(CK)中显著低于施肥处理(1/3N、2/3N和N)(P0.05),但1/3N、2/3N和N处理的稻田nir S基因丰度没有明显差异;0~5 cm土层中qno B和nos Z基因丰度存在随氮肥水平提高而增加的趋势,10~20 cm土层中nos Z基因丰度在2/3N和N处理下显著高于1/3N处理(P0.05)。N处理的稻田N_2O排放通量显著高于2/3N及1/3N处理(P0.05),后者又显著高于CK处理(P0.05)。相关分析结果表明稻田N_2O排放通量与0~5 cm土层中硝化螺菌门Nitrospirae相对丰度及10~20 cm土层中amo A基因丰度存在显著相关性(P0.05,n=10)。综上所述,氮肥水平提高增加了稻田细菌群落多样性,促进了稻田N_2O排放,且本研究稻田中硝化作用微生物群落及丰度变化与稻田N_2O排放的关系更为密切。     

The effects of silicon fertilizer on denitrification potential and associated genes abundance in paddy soil

This study evaluated the effect of silicate fertilizer on denitrification and associated gene abundance in a paddy soil. A consecutive trial from 2013 to 2015 was conducted including the following treatments: control (CK), mineral fertilizer (NPK), NPK plus sodium metasilicate (NPK + MSF), and NPK plus slag-based silicate fertilizer (NPK + SSF). Real-time quantitative PCR (qPCR) was used to analyze the abundances of nirS, nirK, and nosZ genes. Potential N₂O emissions and ammonium and nitrate concentrations were related to the nirS and nirK gene abundance. Compared with the NPK treatments, the addition of a Si fertilizer decreased N₂O emission rates and denitrification potential by 32.4–66.6 and 22.0–59.2%, respectively, which were probably related to increased rice productivity, soil Fe availability, and soil N depletion. The abundances of nirS and nirK genes were decreased by 17.7–35.8% and 21.1–43.5% with addition of silicate fertilizers, respectively. Rates of total N₂O and N₂O from denitrification (DeN₂O) emission were positively correlated with the nirS and nirK gene abundance. Nitrate, exchangeable NH₄ ⁺, and Fe concentrations were the main factors regulating the nirS and nirK gene abundance. Silicate fertilization during rice growth may serve as an effective approach to decreasing N₂O emissions.     

Carbon-iron coupling reduced N2O emissions via promoting the conversion to N2 in paddy soils

The influence of redox reactions involving carbon-iron coupling (organic carbon and iron oxides) on nitrous oxide (N₂O) production in paddy soils remains poorly understood. In this study, two microcosm experiments were conducted to investigate the effects of carbon-iron coupling on N₂O emissions, and the underlying mechanisms were verified using quantitative denitrification functional genes (nirS, nirK, nosZI and nosZII) and high-throughput sequencing. The results showed that ferrihydrite (iron) significantly promoted N₂O-N emissions (p < 0.05) after adding ammonium nitrogen, while glucose (carbon) significantly inhibited N₂O-N emissions (p < 0.05). Carbon-iron coupling significantly decreased N₂O-N emissions (p < 0.05) but did not affect soil total nitrogen loss and increased nitrogen (N₂) emissions. After adding high concentrations of acetylene (10% C₂H₂), the N₂O-N emissions from carbon-iron coupling treatment increased significantly from 6.4 to 11.9 mg N kg⁻¹ (p < 0.05), which confirmed that the carbon-iron coupling reduced the N₂O emissions by promoting the conversion of N₂O to N₂. The mechanisms behind carbon-iron coupling promoting complete denitrification and reducing N₂O emissions were attributed to glucose promoting iron reduction and carbon-iron coupling enhancing the abundance of nosZI (42.7%) and nosZII (16.6%).     

Elevated carbon dioxide stimulates nitrous oxide emission in agricultural soils: A global meta-analysis

Elevated carbon dioxide (CO₂)(e CO₂) has been shown to affect the nitrous oxide (N₂O) emission from terrestrial ecosystems by altering the interaction of plants,soils,and microorganisms.However,the impact of e CO₂ on the N₂O emission from agricultural soils remains poorly understood.This meta-analysis summarizes the effect of e CO₂ on N₂O emission in agricultural ecosystems and soil physiochemical and biological characteristics using 50 publications selected.The e CO₂ effect values,which equal to the percentage changes of N₂O emission under e CO₂,were calculated based on the natural logarithm of the response ratio to e CO₂.We found that e CO₂ significantly increased N₂O emission (by 44%),which varied depending on experimental conditions,agricultural practices,and soil properties.In addition,e CO₂ significantly increased soil water-filled pore space (by 6%),dissolved organic carbon content (by11%),and nitrate nitrogen content (by 13%),but significantly reduced soil p H (by 1%).Moreover,e CO₂ significantly increased soil microbial biomass carbon(by 28%) and soil microbial biomass nitrogen (by 7%) contents.Additionally,e CO₂ significantly increased the abundances of ammonia-oxidizing bacteria(AOB) amo A (by 21%),nir K (by 15%),and nir S (by 15%),but did not affect the abundances of ammonia-oxidizing archaea (AOA) amo A and nos Z.Our findings indicate that e CO₂ substantially stimulates N₂O emission in agroecosystems and highlight that optimization of nitrogen management and agronomic options might suppress this stimulation and aid in reducing greenhouse effect.