施肥制度对水稻土壤nosZ型反硝化细菌群落的影响

由含氧化亚氮还原酶(NOS)的反硝化细菌驱动的氧化亚氮(N2O)还原成氮气(N2)的过程是N2O排放的重要调控途径。为探明施肥对稻田土壤nosZ型反硝化细菌群落的影响,采用荧光定量PCR和高通量测序等方法,研究了湖南省宁乡县长达30年的定位试验条件下4种施肥制度[不施肥(CK)、化肥(CF)、70%化肥+30%有机肥(CFM1)和40%化肥+60%有机肥(CFM2)]对水稻土壤nosZ型反硝化细菌数量和群落结构的影响。结果表明:不同施肥处理nosZ基因丰度为2.14×10~8～6.09×10~8copies·g~(-1)干土,施肥处理nosZ基因拷贝数比对照低47.3%～64.8%(P0.05),但不同有机肥配施比例处理间nosZ基因拷贝数差异不显著;变形菌门是优势门水平类群,占总序列的60.2%～77.5%; Bacteria_unclassified、Proteobacteria_unclassified、Betaproteobacteria_unclassified和根瘤菌目为优势目水平类群,占总序列的93.6%～95.9%。施肥显著降低了Proteobacteria_unclassified的相对丰度(P0.01),但显著提高了根瘤菌目、environmental_samples和红环菌目的相对丰度(P0.05);施肥显著改变nosZ型反硝化细菌的群落结构,但有机肥配施比例对其影响较弱;除碳氮比外,其他土壤理化性质均显著影响nosZ型反硝化细菌的数量和群落结构,其中,硝态氮和土壤p H是驱动nosZ型反硝化细菌群落变化的主要因子;施肥显著影响nosZ型反硝化细菌数量和群落结构,有机肥配施比例对nosZ型反硝化细菌群落的影响较弱,研究结果为进一步阐述施肥制度对土壤反硝化微生物的影响提供依据。     

静态好氧高温牛粪堆肥中nirK型反硝化细菌群落动态变化

【背景】堆肥过程中,不同反硝化微生物相互作用产生大量气态氮,不仅导致氮素流失使得堆肥肥效降低,而且造成环境污染。但是目前关于堆肥中反硝化细菌群落结构变化,尤其是群落结构与堆肥理化因子间相关性方面的报道较为欠缺。【目的】对堆肥中反硝化细菌进行研究,旨在揭示反硝化细菌群落动态变化,为深入理解堆肥氮循环机理提供科学数据。【方法】设计一种静态好氧高温堆肥技术处理牛粪和水稻秸秆,利用高通量测序技术研究堆肥中nirK型反硝化细菌群落组成的动态变化,并分析优势反硝化细菌菌属与理化指标之间的相关性。【结果】堆肥全程共17d,各项堆肥理化指标以及生物学指标表明堆肥已经基本腐熟。高通量测序结果表明,在堆肥的不同阶段nirK型反硝化细菌群落结构差异显著。门水平上,堆肥中反硝化细菌属于变形菌门(Proteobacteria)和一未分类门;目水平上,优势类群主要属于根瘤菌目(Rhizobiales)、红杆菌目(Rhodobacterales)和伯克氏菌目(Burkholderiales),其中根瘤菌目(Rhizobiales)的种类最多,而伯克氏菌目(Burkholderiales)的相对丰度最高。Spearman相关性分析表明未分类门的反硝化细菌和未分类科根瘤菌目的反硝化细菌与全碳、碳氮比、含水率以及pH呈显著负相关(P0.05),与凯氏氮和硝态氮呈显著正相关(P0.05);其他优势菌属与全碳、碳氮比、含水率以及pH呈显著正相关(P0.05),与凯氏氮和硝态氮呈显著负相关(P0.05);未分类科伯克氏菌目的反硝化细菌、未分类纲变形菌门的反硝化细菌、产碱杆菌科的Pusillimonas属和副球菌属(Paracoccus)与铵态氮显著相关(P0.05)。【结论】静态好氧高温堆肥技术可以缩短堆肥周期。在堆肥的不同阶段nirK型反硝化细菌群落结构差异显著,并且该菌群落结构的变化受到堆肥理化因子的显著影响。本研究有助于揭示堆肥中氮素转化规律,并为改进堆肥工艺提供理论依据。     

施肥对设施菜地nirK型反硝化细菌群落结构和丰度的影响

采用末端限制性片段多态性分析(T-RFLP)和实时荧光定量PCR(real-time PCR)方法,研究了甘肃武威设施菜地不同施肥条件下0~20 cm、20~40 cm土层中土壤nirK型反硝化细菌群落结构和丰度的变化.结果表明:施肥对土壤中nirK型反硝化细菌的群落结构具有明显影响,且对70、156、190 bp片段所代表设施菜地土壤优势种群影响最显著.施肥对0~20cm土层nirK型反硝化细菌丰度有明显影响,其最大值出现在全有机肥(M)处理、为每克干土2.16×107个拷贝数,分别是对照(CK)和全化肥(NPK)处理的2.04和2.02倍.设施菜地土壤0~20 cm与20~40 cm土层nirK型反硝化细菌的优势种群及其基因丰度均存在显著差异,且设施菜地土壤中nirK型反硝化细菌的群落结构和丰度与大田差异明显.土壤pH值、有机质及硝酸盐含量均影响nirK型反硝化细菌的群落结构和丰度.系统发育分析结果表明,土壤中除存在与厌氧反硝化细菌亲缘相近的nirK型反硝化微生物外,还存在与好氧反硝化菌亲缘关系相近的nirK型反硝化微生物,如根瘤菌属、苍白杆菌属、土壤杆菌属等.     

施肥对设施菜地nirK型反硝化细菌群落结构和丰度的影响

采用末端限制性片段多态性分析(T-RFLP)和实时荧光定量PCR(real-time PCR)方法,研究了甘肃武威设施菜地不同施肥条件下0～20 cm、20～40 cm土层中土壤nirK型反硝化细菌群落结构和丰度的变化.结果表明： 施肥对土壤中nirK型反硝化细菌的群落结构具有明显影响,且对70、156、190 bp片段所代表设施菜地土壤优势种群影响最显著.施肥对0～20 cm土层nirK型反硝化细菌丰度有明显影响,其最大值出现在全有机肥(M)处理、为每克干土2.16×10⁷个拷贝数,分别是对照(CK)和全化肥(NPK)处理的2.04和2.02倍.设施菜地土壤0～20 cm与20～40 cm土层nirK型反硝化细菌的优势种群及其基因丰度均存在显著差异,且设施菜地土壤中nirK型反硝化细菌的群落结构和丰度与大田差异明显.土壤pH值、有机质及硝酸盐含量均影响nirK型反硝化细菌的群落结构和丰度.系统发育分析结果表明,土壤中除存在与厌氧反硝化细菌亲缘相近的nirK型反硝化微生物外,还存在与好氧反硝化菌亲缘关系相近的nirK型反硝化微生物,如根瘤菌属、苍白杆菌属、土壤杆菌属等.
     

不同母岩发育马尾松土壤固氮菌群落结构和丰度特征

生物固氮是马尾松林地土壤氮素的重要来源,固氮微生物群落组成和数量的变化对土壤氮素供应和地力维持起重要作用。采用池栽试验,应用荧光定量PCR（聚合酶链式反应,Polymerase Chain Reaction）技术,借助Illumina Miseq高通量测序平台,以nifH基因为标靶,研究四类母岩（变余砂岩、长石石英砂岩、石英砂岩和玄武岩）发育马尾松土壤固氮菌群落结构和丰度的差异及其与土壤化学性质之间的关系。结果表明：（1）玄武岩土壤的有机碳、全氮、碱解氮、微生物量氮、马尾松株高和地径均显著高于其他母岩（P < 0.05）。（2）四类母岩土壤nifH基因丰度差异显著,玄武岩土壤nifH基因丰度分别是变余砂岩、石英砂岩和长石石英砂岩的3.75倍、7.89倍和4.41倍。（3）四类母岩土壤固氮菌群落α多样性指数（丰富度和多样性）差异显著,且玄武岩显著高于其他母岩。四类母岩发育土壤共获得有效序列159231条,分属于6个门、14个纲、41个目、69个科和122个属。门水平上,变形菌门和蓝藻门为主要优势类群。属水平上,慢生根瘤菌属、眉藻属、根瘤菌属和固氮螺菌属为主要优势属。玄武岩土壤变形菌门、慢生根瘤菌属、根瘤菌属和固氮螺菌属相对丰度显著高于其他母岩。层次聚类和非度量多维尺度（NMDS）分析结果表明,石英砂岩和长石石英砂岩土壤固氮菌群落结构相似,玄武岩与其他母岩固氮菌群落结构差异较大。（4）土壤有机碳、全氮、碱解氮和微生物量氮是固氮菌丰度、α多样性及群落结构的主要影响因子。综上,玄武岩土壤肥力高,提高了土壤固氮菌数量和群落多样性,有利于马尾松生长。研究从微生物学角度为马尾松适地造林和氮素调控提供了科学依据。     

不同放牧强度下土壤氨氧化和反硝化微生物的变化特征

土壤硝化及反硝化功能微生物在氮素可利用性、硝酸盐淋溶和氧化亚氮温室气体排放等方面起着关键作用,在指示不同放牧强度对生态系统的影响及预测草地生态系统退化状况等方面具有重要意义。以内蒙古干旱半干旱草原不同放牧强度(轻度、中度和重度)的长期试验样地为对象,应用定量PCR和限制性末端片段长度多态性(Terminal restriction fragment length polymorphism,T-RFLP)的方法,研究土壤氨氧化古菌(ammonia-oxidizing archaea,AOA)、氨氧化细菌(ammonia-oxidizing bacteria,AOB)和反硝化细菌的丰度、群落结构和多样性对不同放牧强度的响应。结果表明,土壤p H和铵态氮含量分别在7.90—8.18和6.37—35.92 mg/kg之间,中度放牧处理显著增高了土壤pH(P=0.03),而铵态氮含量在重度放牧处理中最高(P=0.02)。不同放牧强度下土壤异养呼吸相比未放牧处理均显著降低(P=0.02)。土壤AOA-amoA和AOB-amoA基因丰度范围分别为每克干土(4.94—7.60)×10~9个拷贝数和(0.68—3.75)×10~6个拷贝数,放牧处理对AOA-amoA基因丰度无显著影响,中度放牧处理显著降低了AOB-amoA基因丰度(P=0.04);反硝化微生物nosZ基因丰度随在轻度放牧处理中最低(P=0.03)。土壤铵态氮含量是影响AOA-amoA和AOB-amoA基因丰度的主要因子,而nosZ基因丰度主要受反硝化底物含量及土壤通气状况的影响。冗余分析表明由放牧所引起的可利用性氮含量的变化是导致氨氧化和反硝化微生物群落结构显著变化的主要因素。     

野葛根际固氮微生物群落分布特征及其影响因素

【目的】研究野葛根际固氮微生物群落的空间分布特征,同时结合地理气候因子、土壤理化性质探讨影响野葛根际固氮微生物群落的空间分布特征的主要因素。【方法】在野葛广泛分布的广西、江西和湖南采集17份野葛根际土壤,测定土壤常规理化性质,进行基于MiSeq测序平台的nifH基因高通量测序,采用多元统计分析方法探索固氮微生物群落结构与环境因子之间的关系。【结果】不同采样区域固氮微生物α多样性无显著差异;同一采样区内各样点的固氮微生物群落结构与组成较相似,而不同采样区之间固氮微生物群落结构与组成差异较大,这表明固氮微生物群落存在明显的区域性分布特征。野葛根际土壤中固氮微生物主要归属于变形菌门(Proteobacteria)(相对丰度>60%),在目水平上,已注释出的固氮微生物主要归属于根瘤菌目(Rhizobiales)和伯克氏菌目(Burkholderiales)。根瘤菌目相对丰度在江西采样区最高,伯克氏菌目相对丰度在湖南采样区最高。pH、交换性钙、全钾、海拔和年平均气温对土壤固氮微生物群落结构的影响较大。优势种群伯克氏菌目与pH、交换性钙和全钾正相关,与C/N负相关;优势种群根瘤菌目与pH、交...     

转BT+ CPTI基因抗虫棉的种植对土壤根际反硝化菌丰度和多样性影响

摘要:【目的】为评价转基因棉花的种植对根际土壤反硝化细菌丰度和多样性的影响,【方法】以转Bt + CpTI 基因抗虫棉(SGK321)及非转基因亲本棉花石远321(SY321)根际土壤反硝化细菌为研究对象,分别在种植前和棉花的不同生长时期(蕾期、花期、铃期和絮期)取样。采用实时荧光定量PCR方法和末端标记限制性片段多态性(T-RFLP)技术对土壤中反硝化细菌的功能基因nosZ基因进行定量和多样性分析。【结果】结果发现随棉花不同生长时期两种棉花根际土壤中的反硝化细菌丰度均发生显著变化,但变化趋势不同。转基因棉花根际土壤反硝化细菌从种植前的3.12×106 copies/g dry soil一直增加到铃期的2.81×107 copies/g dry soil,增加了8倍。非转基因棉花根际土壤反硝化细菌丰度变化受生长周期影响更为显著,表现为蕾期增加,花期降低,铃期又增加的趋势。典范相关及部分典范相关分析反硝化细菌多样性结果表明其多样性受环境因素pH、硝酸根浓度以及棉花生长时期(蕾期和花期)影响最为显著,但此外棉花品种也起到了非常重要的作用,【结论】反硝化细菌的丰度和多样性既受棉花生长时期的影响,同时也受棉花品种的影响,转基因棉花通过调节根际土壤中的pH 和硝酸根浓度,来影响其多样性和丰度。转Bt + CpTI基因抗虫棉的种植增加了土壤pH,从而导致根际土壤反硝化细菌的多样性和丰度增加。     

亚热带森林土壤氨氧化微生物和反硝化微生物功能基因丰度对氮磷输入的响应

为探究亚热带森林土壤氨氧化微生物和反硝化微生物对氮、磷输入的响应，2015年开始在钱江源国家森林公园设置氮磷模拟添加试验，包括对照(CK)、氮(N)添加、磷(P)添加和氮磷(NP)添加4种处理，于2021年4月(湿季)和11月(干季)采集土样，采用定量PCR的方法分析亚热带森林土壤氨氧化微生物(氨氧化古菌AOA、氨氧化细菌AOB和全程氨氧化菌comammox)amoA基因和反硝化微生物功能基因(nirS、nirK和nosZ基因)的丰度变化特征。结果表明：长期N输入显著降低土壤pH,但显著提高了土壤铵态氮和硝态氮含量，而长期P输入显著提高了土壤有效磷和总磷含量。氮的输入(N和NP处理)显著提高了干湿季土壤AOB-amoA基因丰度，且在N处理中最高，达8.30×10⁷ copies·g^-1。NP处理土壤AOA-amoA基因丰度显著高于CK,达1.17×10⁹ copies·g^-1。comammox-amoA基因丰度在不同季节间差异显著，其他基因丰度在不同季节间差异均不显著。双因素方差分析表明，N输入显著...     

牛场肥水灌溉对土壤nirK、nirS型反硝化微生物群落结构的影响

以徐水县梁家营长期定位施肥试验田为研究对象,利用末端限制性片段长度多态性(T-RFLP)分析和克隆文库构建,研究了5种施肥处理(清水灌溉CK、无机肥灌溉CF、牛场肥水不同浓度、不同次数灌溉T4、T5和T11)下土壤中nirK、nirS型反硝化细菌群落多样性及其群落结构的演变。结果表明,不同施肥处理下nirK、nirS型反硝化细菌群落多样性无显著差异,但群落结构却有明显变化:nirK型反硝化细菌群落结构既受施肥种类又受施肥量影响,优势种群尤其对施肥种类和施肥量响应显著;nirS型反硝化细菌则主要受施肥种类影响,施肥量影响微弱。牛场肥水处理和无机肥处理分别促进和抑制不同的nirS型反硝化细菌,群落主成分受无机肥促进、牛场肥水抑制。系统发育分析结果表明,土壤中nirK型反硝化细菌主要与假单胞菌属(Pseudomonas)、产碱杆菌属(Alcaligenes)和根瘤菌属(Rhizobium)的反硝化细菌具有较近的亲缘关系;nirS型反硝化细菌主要与劳尔氏菌(Ralstonia)和红长命菌属(Rubrivivax)有较近的亲缘关系。试验土壤中反硝化微生物多与目前已报道的好氧反硝化细菌亲缘关系较近,这可能与微生物分析取自表层土有关。     

不同季节艾比湖湿地盐角草根际nirS-型与nirK-型反硝化细菌群落组成分析

旨在了解艾比湖湿地盐生植物盐角草根际与非根际中不同类型反硝化细菌的分布及其随季节变化情况,为温带干旱地区荒漠盐化生态系统的代表-艾比湖湿地在生态植被恢复过程中,由微生物推动的土壤氮素循环过程提供数据支撑。采集了艾比湖湿地夏、秋、春三个季节的盐角草根际和非根际土壤样本,通过高通量测序技术,比较分析了nirS-型和nirK-型两种类型的反硝化细菌的多样性和群落结构特点;利用RDA (redundancy analysis)探究了土壤理化因素对反硝化细菌多样性及群落结构的影响。艾比湖湿地盐角草根际与非根际中,nirS-型和nirK-型反硝化细菌多样性最高的为秋季根际土壤样本;各土壤样本中的反硝化细菌多样性均呈现根际>非根际。盐角草各土壤样本中的nirS-型反硝化细菌在门分类水平上隶属于变形菌门(Proteobacteria),厚壁菌门(Firmicutes)和放线菌门(Actinobacteria),而nirK-型反硝化细菌在门水平上分类仅包括了Proteobacteria和Firmicutes。Proteobacteria在各土壤样本中的占比均较高;其中Gamma-Proteobacteria的盐单胞菌属(Halomonas)和假单胞菌属(Pseudomonas)是各土壤样本所共有的nirS-型反硝化菌的优势菌属,但它们在每个土壤样本中的相对丰度各有差异。Alpha-Proteobacteria的根瘤菌属(Rhizobium)是盐角草各土壤样本中较为广泛存在的nirK-型反硝化细菌。艾比湖湿地盐角草各土壤样本中的反硝化细菌群落结构存在着一定的差异。RDA结果显示含水量、有机质、全氮和铵态氮等对各土壤样本中的nirS-型反硝化细菌的多样性影响较大,含水量、有机质、全氮、碱解氮等是nirK-型反硝化细菌多样性的主要影响因素。土壤电导率、全磷、全钾、全氮和碱解氮协同影响nirS-型反硝化细菌的群落结构,有机质、速效钾、速效磷、pH和硝态氮是nirK-型反硝化细菌群落结构组成的主要影响因素。艾比湖湿地反硝化细菌呈现季节性变化,nirS-型和nirK-型反硝化细菌以不同的主要菌属,共同推进湿地反硝化作用。而对于湿地生态系统的保护,则需要进行长期而广泛的土壤状态评估和土壤反硝化微生物菌群的动态监测。     

TGGE analysis of the diversity of ammonia-oxidizing and denitrifying bacteria in submerged filter biofilms for the treatment of urban wastewater

The spatial and temporal diversity of the bacterial community-forming biofilms in a pilot-scale submerged biofilter used for the treatment of urban wastewater was analyzed by a temperature-gradient gel electrophoresis (TGGE) approach. TGGE profiles based on partial sequence of the 16S rRNA gene showed that the community composition of the biofilms remained fairly stable along the column system and during the whole time of operation of the biofilter (more than 1 year). Community-profiling based on the amplification and separation of partial ammonia monooxygenase (amoA) and nitrous oxide reductase (nosZ) genes demonstrated that ammonia-oxidizing and denitrifying bacteria coexisted in both the anoxic and the aerated parts of the system. Several amoA and nosZ bands separated by TGGE were reamplified and sequenced, in order to further analyze the composition of these microbial communities in the biofilm. Phylogeny inferred from amoA/AmoA revealed the prevalence of Nitrosomonas species with five sequences affiliated to Nitrosomonas oligotropha, six sequences affiliated to Nitrosomonas europaea, and three sequences that showed only 75.7–76.1% identity of the DNA sequence with the closest described species (Nitrosomonas nitrosa). According to literature, this low identity value is indicative of previously undiscovered species. Eighteen new partial nosZ sequences were obtained which were mostly related to nosZ of gamma-proteobacteria (Pseudomonas) or clustered in the periphery of previously known denitrifying alpha-proteobacteria (Bradyrhizobium and Azospirillum).     

广东省本地油茶和引种油茶根际土壤微生物群落特征

【背景】近年来,油茶低效林面积较大,根际土壤微生物影响林木抗性和生长,对林业可持续发展具有重要意义。【目的】了解广东省本地油茶和引种油茶根际土壤微生物群落特征。【方法】利用高通量测序分析油茶根际土壤微生物群落组成。【结果】油茶根际土壤细菌有26门77纲201目377科593属676种,真菌有14门50纲121目266科502属631种。油茶根际土壤中的优势细菌为酸杆菌门和变形菌门,优势真菌为子囊菌门和担子菌门。两种油茶根际土壤微生物组成差异显著,本地油茶根际土壤的细菌多样性显著高于引种油茶。在门水平上,脱硫杆菌门细菌和罗兹菌门、被孢霉门真菌的相对丰度在两种油茶间差异显著,Amorphotheca在本地油茶根际土壤中特异性富集。两种油茶根际土壤细菌碳代谢相对丰度差异显著,真菌以腐生营养型为主,其次为病理营养型和共生营养型。本地油茶根际土壤中显著富集土壤腐生菌,而共生营养型真菌(尤其是丛枝菌根真菌)相对丰度(6.43%)显著低于引种油茶中(21.83%)。此外,有机质和养分含量是影响油茶根际土壤微生物群落的关键因子。【结论】本地油茶和引种油茶根际土壤微生物群落组成和结构差异显著,Amorp...     

Differentiated Response of Denitrifying Communities to Fertilization Regime in Paddy Soil

The impact of fertilization regimes on sequential denitrifying communities was investigated in a rice paddy field with 17 years continuous fertilization, located in Taoyuan Agro-ecosystem Research Station (110°72″ E, 28°52″ N), China. The diversity, community composition, and size of denitrifying genes of narG, qnorB, and nosZ were determined using molecular tools including terminal restriction fragment length polymorphism, quantitative polymerase chain reaction (qPCR), cloning, and sequencing analysis. Soil samples were collected from the plots with no fertilizer (NF), urea (UR), balanced mineral fertilizers (BM), and BM combined with rice straw (BMR). UR and BM caused marked increase in the community size of the denitrifying genes; however, BMR resulted in the highest abundance. The community size of narG was the most affected by the fertilization regimes, while qnorB was the least. Fertilization also induced some shifts in the composition of denitrifying genes, but the responses of different genes varied. However, fertilization regimes caused no significant changes to the diversity of the denitrifying genes. Potential denitrification activity (PDA) was significantly correlated with the abundance of narG and nosZ rather than qnorB, but there were no such correlations between PDA and the composition and diversity of denitrifying communities. Conclusively, long-term fertilization significantly affected denitrifying community size and composition, but not diversity. Among the sequential denitrifying genes, narG was the most, while qnorB was the least sensitive communities to fertilization regimes.     

Long-term fertilization and tillage regimes have limited effects on structuring bacterial and denitrifier communities in a sandy loam UK soil

Denitrification causes loss of available nitrogen from soil systems, thereby reducing crop productivity and increasing reliance on agrochemicals. The dynamics of denitrification and denitrifying communities are thought to be altered by land management practices, which affect the physicochemical properties of the soil. In this study, we look at the effects of long-term tillage and fertilization regimes on arable soils following 16 years of treatment in a factorial field trial. By studying the bacterial community composition based on 16S rRNA amplicons, absolute bacterial abundance and diversity of denitrification functional genes (nirK, nirS and nosZ), under conditions of minimum/conventional tillage and organic/synthetic mineral fertilizer, we tested how specific land management histories affect the diversity and distribution of both bacteria and denitrification genes. Bacterial and denitrifier communities were largely unaffected by land management history and clustered predominantly by spatial location, indicating that the variability in bacterial community composition in these arable soils is governed by innate environmental differences and Euclidean distance rather than agricultural management intervention.     

Comparison of bacterial community characteristics between complete and shortcut denitrification systems for quinoline degradation

For quinoline-denitrifying degradation, very few researches focused on shortcut denitrification process and its bacterial community characteristics. In this study, complete and shortcut denitrification systems were constructed simultaneously for quinoline degradation. By calculation, specific quinoline removal rates were 0.905 and 1.123 g/(gVSS d), respectively, in the complete and shortcut systems, and the latter was 1.24 times of the former. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), high-throughput sequencing, and quantitative PCR (qPCR) techniques based on 16S rRNA were jointly applied to compare microbial community structures of two systems. Many denitrifying bacteria phyla, classes, and genera were detected in the two systems. Phylum Proteobacteria, Class Gammaproteobacteria, and Genus Alicycliphilus denitrificans were the dominant contributors for quinoline-denitrifying degradation. In the shortcut denitrification system, main and specific strains playing crucial roles were more; the species richness and the total abundance of functional genes (narG, nirS, nirK, and nosZ) were higher compared with the complete denitrification system. It could be supposed that inorganic-nitrogen reductase activity of bacterial community was stronger in the shortcut denitrification system, which was the intrinsic reason to result in higher denitrification rate.

     

Nitrous Oxide Reductase (nosZ) Gene Fragments Differ between Native and Cultivated Michigan Soils

The effect of standard agricultural management on the genetic heterogeneity of nitrous oxide reductase (nosZ) fragments from denitrifying prokaryotes in native and cultivated soil was explored. Thirty-six soil cores were composited from each of the two soil management conditions. nosZ gene fragments were amplified from triplicate samples, and PCR products were cloned and screened by restriction fragment length polymorphism (RFLP). The total nosZ RFLP profiles increased in similarity with soil sample size until triplicate 3-g samples produced visually identical RFLP profiles for each treatment. Large differences in total nosZ profiles were observed between the native and cultivated soils. The fragments representing major groups of clones encountered at least twice and four randomly selected clones with unique RFLP patterns were sequenced to verify nosZ identity. The sequence diversity of nosZ clones from the cultivated field was higher, and only eight patterns were found in clone libraries from both soils among the 182 distinct nosZ RFLP patterns identified from the two soils. A group of clones that comprised 32% of all clones dominated the gene library of native soil, whereas many minor groups were observed in the gene library of cultivated soil. The 95% confidence intervals of the Chao1 nonparametric richness estimator for nosZ RFLP data did not overlap, indicating that the levels of species richness are significantly different in the two soils, the cultivated soil having higher diversity. Phylogenetic analysis of deduced amino acid sequences grouped the majority of nosZ clones into an interleaved Michigan soil cluster whose cultured members are α-Proteobacteria. Only four nosZ sequences from cultivated soil and one from the native soil were related to sequences found in γ-Proteobacteria. Sequences from the native field formed a distinct, closely related cluster (D_mean = 0.16) containing 91.6% of the native clones. Clones from the cultivated field were more distantly related to each other (D_mean = 0.26), and 65% were found outside of the cluster from the native soil, further indicating a difference in the two communities. Overall, there appears to be a relationship between use and richness, diversity, and the phylogenetic position of nosZ sequences, indicating that agricultural use of soil caused a shift to a more diverse denitrifying community.     

不同土地利用方式土壤氨氧化微生物和反硝化微生物时空分布特征

研究不同土地利用方式下氮循环相关微生物在不同土壤剖面的分布,可为认识和理解土壤氮转化过程提供科学依据。土壤氨氧化微生物和反硝化微生物在调节氮肥利用率、硝态氮淋溶和氧化亚氮(N₂O)排放等方面有着重要作用。以北京郊区农田和林地两种土地利用方式为研究对象,分析土壤氨氧化潜势和亚硝酸盐氧化潜势在0—100 cm土壤剖面上的季节分布(春季和秋季),并通过实时荧光定量PCR方法表征土壤氨氧化和反硝化微生物的时空分布特征。结果表明,农田土壤氨氧化潜势、亚硝酸盐氧化潜势、氨氧化微生物和反硝化微生物丰度均显著高于林地土壤,且随土壤深度增加而显著降低。除氨氧化古菌amoA基因丰度在不同季节间无显著差异外,春季土壤氨氧化细菌(amoA基因)、反硝化微生物nirS、nirK和典型nosZ I基因的丰度均显著高于秋季。土壤有机质、总氮、NH~+₄-N、NO~-₃-N含量与氨氧化微生物和反硝化微生物的功能基因丰度显著相关。综上,不同土地利用方式下土壤氮循环相关微生物的丰度与土壤氮素的可利用性和转化过程紧密相关,研究结果对土壤氮素利用和养分管理提供...     

生物炭对褐土旱地玉米季氮转化功能基因、丛枝菌根真菌及N2O释放的影响

以豫西旱地玉米农田为研究对象,设置不同生物炭施用量处理（T0：不施用生物炭;T1：施用生物炭20 t/hm²;T2：施用生物炭40 t/hm²）,采用密闭式静态箱法测定N₂O排放通量和荧光定量PCR法分析丛枝菌根（arbuscular mycorrhizal,AM）真菌、氨单加氧酶（amoA）、亚硝酸盐还原酶（nirS、nirK）以及氧化亚氮还原酶（nosZ）的基因丰度,同时测定土壤理化性状的变化。研究结果表明,随着生物炭施用量的增加,土壤pH和含水量呈增加趋势,土壤有机碳、全氮和铵态氮含量显著提高,土壤容重和硝态氮含量显著降低。T1和T2处理土壤有机碳含量分别较T0显著提高38.44%和71.01%;T1和T2处理土壤铵态氮含量分别较T0显著增加15.89%和30.46%;T2处理土壤全氮含量较T0处理显著提高14.87%;T1和T2处理土壤硝态氮含量分别较T0减少10.57%和21.40%。随着生物炭施用量的增加,AM真菌侵染率显著增加,T1和T2处理分别较T0处理提高71.88%和115.88%;AOA、AOB、nirK和nirS基因丰度显著降低;nosZ基因丰度增加。施加生物炭处理的N₂O排放通量和累积排放量均低于不施生物炭处理,具体表现为：T0 > T1 > T2。相关分析表明,生物炭施用量与AM真菌基因丰度呈显著正相关;与nosZ基因丰度呈正相关;与AOA、AOB、nirK、nirS基因丰度呈极显著负相关。N₂O排放通量与AOA、nirK、nirS基因丰度呈极显著正相关;与土壤含水量和土壤硝态氮含量呈显著正相关;与AM真菌、nosZ基因丰度、易提取球囊霉素含量、铵态氮含量呈极显著负相关。集成推进树（ABT）分析表明,AOA对N₂O排放的影响最大,其次是AM真菌和nirS。总之,生物炭处理改善土壤理化性质、提高土壤AM真菌侵染率、调节硝化、反硝化相关功能基因的丰度,减少N₂O气体排放,为旱地农田合理施用生物炭减少N₂O气体排放提供理论依据。     

Abundance and Structure Composition of nirK and nosZ Genes as Well as Denitrifying Activity in Heavy Metal-polluted Paddy Soils

Denitrification is an important microbial process in soils and leads to the emission of nitrous oxide (N₂O). However, studies about the microbial community involved in denitrification processes in polluted paddy fields are scarce. Here, we studied two rice paddies which had been polluted for more than three decades by metal mining and smelter activities. Abundance and community composition were determined using real-time polymerase chain reaction (PCR) assay and denaturing gradient gel electrophoresis of nitrite reductase and nitrous oxide reductase gene amplicons (nirK and nosZ), while denitrifying activities were assessed by measuring potential denitrifier enzyme activity. We found that the community structure of both nirK and nosZ containing denitrifiers shifted under pollution in the two rice paddies. All the retrieved nirK sequences did not group into either α- or β-proteobacteria, while most of the nosZ species were affiliated with α-proteobacteria. While the abundance of both nirK and nosZ was significantly reduced in the polluted soils at “Dexing” (with relatively higher Cu levels), these parameters did not change significantly at “Dabaoshan” (polluted with Cd, Pb, Cu, and Zn). Furthermore, total denitrifying activity and N₂O production and reduction rates also only decreased under pollution at “Dexing.” These findings suggest that nirK and nosZ containing denitrifier populations and their activities could be sensitive to considerable Cu pollution, which could potentially affect N₂O release from polluted paddy soils.