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...     

不同林龄杉木人工林土壤硝化和反硝化作用

对不同林龄杉木人工林(5、8、21、27和40年生)土壤硝化与反硝化过程及功能微生物丰度进行研究。结果表明: 土壤净硝化速率随林龄的增加波动变化,8、27年生杉木人工林土壤净硝化速率显著低于5、21和40年生。27年生杉木人工林土壤氨氧化古菌(AOA) amoA基因丰度显著低于40年生,其他林龄AOA amoA基因丰度之间无显著差异。不同林龄杉木人工林的氨氧化细菌(AOB) amoA基因丰度、反硝化功能基因丰度以及反硝化潜势均无显著差异。逐步回归分析表明,土壤氨氧化微生物AOA amoA基因丰度受土壤理化性质的影响不显著,土壤总碳和土壤pH是影响AOB丰度的重要因子。反硝化功能基因narG、nirK及nosZ随土壤pH的增加而增加,编码亚硝酸盐还原酶(NIR)的功能基因(nirK、nirS)受土壤总碳的影响。林龄可通过影响AOA amoA基因丰度影响土壤净硝化速率。林龄直接作用于反硝化潜势,或间接影响土壤微生物生物量碳、土壤pH及反硝化功能基因丰度(narG和nirK),进而影响反硝化潜势。相较于反硝化过程,土壤硝化作用及AOA amoA基因丰度对杉木林分发育更加敏感,可适当延长轮伐期以降低土壤硝化作用造成的氮流失风险。     

尕海湿地不同退化阶段土壤氮转化的关键微生物功能基因

明确湿地退化对土壤氮素转化微生物基因丰度的影响,为认识湿地的退化机理以及科学治理高寒退化湿地提供重要依据。以尕海湿地未退化、轻度退化、中度退化和重度退化4种不同退化程度土壤为研究对象,用实时荧光定量PCR技术检测固氮菌(nifH)、氨氧化古菌(AOA-amoA)、氨氧化细菌(AOB-amoA)和反硝化菌(nirK)基因丰度,分析功能基因丰度在不同退化阶段的变化特征。结果表明：0～10 cm土层4种功能基因丰度均显著高于10～20 cm土层,nifH、AOA、AOB和nirK基因丰度的变化范围为10⁶～10⁸、10⁷～10⁸、10⁵～10⁶和10⁷～10⁸ copies·g^-1 soil,且AOA丰度显著高于AOB丰度;4种基因丰度均在轻度退化阶段达到最大值,中度和重度退化阶段显著降低;土壤理化性质对微生物功能基因的解释率达到了87.9%和91.2%,但不同月份关键驱动因子不同;在6月,土壤含...     

秸秆还田对玉米根际氨氧化微生物群落及红壤硝化潜势的影响

为探讨酸性红壤根际氨氧化微生物群落以及硝化作用对不同秸秆还田处理的响应,基于中国科学院鹰潭红壤生态实验站设置的秸秆还田长期试验平台(9年),采用荧光定量PCR和高通量测序技术,研究不同秸秆还田处理(不施肥(CK);氮磷钾肥(NPK);氮磷钾肥+秸秆(NPKS);氮磷钾肥+秸秆猪粪配施(NPKSM);氮磷钾肥+秸秆生物炭(NPKB))下玉米根际土壤氨氧化古菌(ammonia-oxidizing archaea, AOA)和细菌(ammonia-oxidizing bacteria, AOB)丰度和群落结构的变化,揭示了秸秆还田对根际氨氧化微生物群落结构和硝化潜势(potential nitrification activity, PNA)的影响机制。结果发现：相比CK和NPK处理,秸秆还田显著提高了土壤养分含量和硝化潜势,其中有机碳(SOC)、全氮(TN)、全磷(TP)、速效磷(AP)、速效钾(AK)、硝态氮(NO~-₃-N)和铵态氮(NH~+₄-N)含量显著增加,NPKSM处理对土壤肥力提升效果最佳。AOA的硝化潜势显著高于AOB,表明AOA...     

间作对马铃薯种植土壤硝化作用和硝态氮供应的影响

土壤硝态氮供应对满足作物氮素需求至关重要,但间作如何影响土壤硝态氮供应及其作用机制尚不清楚。本研究基于4个氮水平(N₀, 0 kg·hm^-2; N₁, 62.5 kg·hm^-2; N₂, 125 kg·hm^-2; N₃, 187.5 kg·hm^-2)的马铃薯单作、马铃薯与玉米间作小区试验,分析土壤硝态氮含量与强度、硝化势和氨氧化功能基因丰度的差异,探讨间作影响土壤硝态氮供应和氮调控的机理。结果表明: 土壤硝态氮含量和强度随施氮量增加而升高,但同一施氮水平下间作均低于单作。施氮提高了土壤硝化势,且单作的响应高于间作。土壤中氨氧化细菌(AOB)的amoA基因丰度大于氨氧化古菌(AOA),二者在间作时均随施氮量增加呈现先增加后降低的趋势;相同施氮量下,间作的AOA和AOB基因丰度(除N₂外)均低于单作。相关分析、回归分析和主成分分析显示,马铃薯间作后,土壤AOB、AOA的amoA基因丰度下降,硝化势减弱,导致土壤硝态氮含量和强度降低。因此,间作导致土壤硝态氮供应降低与土壤氮转化的微生物过程有关,间作条件下的马铃薯种植应注意保障土壤氮素供应。     

生物炭与炭基肥对采煤塌陷复垦区土壤硝化和反硝化微生物群落的影响

作为一种新型土壤改良剂,生物炭对土壤微生物群落的影响已有报道,但在采煤塌陷复垦区土壤氮循环微生物群落对生物炭添加的响应鲜有报道。以生物炭和炭基肥为添加材料,以淮北地区塌陷复垦土为供试土壤,通过室外盆栽试验,采用荧光定量PCR(qPCR)和末端限制性片段长度多态性(T-RFLP)技术,研究不同生物炭处理的土壤硝化和反硝化微生物的菌群变化。试验共设5个处理:对照(CK)、常规化肥(CF)、炭基肥(BF)、2%生物炭配施化肥(LB)和4%生物炭配施化肥(HB)。结果表明: 与CK处理相比,各施肥处理均显著提高了土壤氨氧化古菌(AOA)、氨氧化细菌(AOB)、反硝化细菌nirK和nirS基因丰度。与CF处理相比,生物炭和炭基肥处理显著提高了AOB和nirK基因丰度,增幅分别达到42.9%～82.1%和33.5%～62.7%。冗余分析表明,土壤有机碳、pH、NH₄⁺-N和速效钾是显著影响AOB群落结构的主要因子,而土壤有机碳、pH和NO₃^--N含量是影响nirK型反硝化细菌群落结构的关键因子。因此,施用生物炭与炭基肥能改良采煤塌陷复垦区土壤质量,提高硝化和反硝化微生物丰度,并改变AOB和nirK型反硝化细菌群落结构。     

亚热带米槠天然林土壤氨氧化微生物对模拟氮沉降的响应

我国亚热带地区是全球氮沉降的热点区域。氮沉降会影响氨氧化微生物的丰度和群落结构,进而改变土壤微生物驱动的养分循环。目前对新近发现的完全氨氧化菌认识不足,极大地制约了对森林土壤氨氧化微生物响应氮沉降的整体认识。本研究以福建省三明市辛口镇格氏栲自然保护区长期模拟氮沉降处理土壤为研究对象,利用实时定量PCR方法,研究氨氧化微生物(包括氨氧化细菌AOB、氨氧化古菌AOA和完全氨氧化菌comammox Nitrospira),尤其是完全氨氧化菌的amoA基因丰度。模拟氮沉降处理包括:不添加N(CK)、低氮(添加40 kg N·hm^-2·a^-1,LN)和高氮(添加80 kg N·hm^-2·a^-1,HN)。结果表明: 8年的氮添加降低了土壤pH值和有机碳含量,提高了土壤硝态氮含量。供试土壤的AOB丰度低于检测限,无法获得目的片段。高氮处理显著提高了AOA丰度,但对完全氨氧化菌clade A和clade B丰度无显著影响。两种氮添加处理均降低了完全氨氧化菌/AOA值,表明氮添加降低了完全氨氧化菌在亚热带森林土壤氨氧化微生物类群中的相对竞争力。针对完全氨氧化菌clade A和clade B的扩增都存在非特异性产物,表明针对森林土壤的高特异性和覆盖度设计引物的必要性。Clade A和clade B丰度与总氮和铵态氮含量呈显著正相关,clade B丰度还与有机碳含量呈显著正相关。总之,模拟氮沉降提高了AOA在亚热带米槠天然林土壤硝化过程中的相对重要性,这些发现可为该地区应对全球变化和氮沉降的风险评估提供理论依据。     

长期施加氮肥及氧化钙调节对酸性土壤硝化作用及氨氧化微生物的影响

以长期施加氮肥及添加氧化钙调节的酸性土壤为研究对象,运用定量PCR和DGGE技术,探讨了土壤氨氧化微生物及硝化作用对不同施肥处理及氧化钙调节的响应。长期施化学氮肥导致酸性土壤p H(KCl)值(3.35—3.47)和硝化潜势(0.02—0.14μg NO-2-N g-1土壤h-1)进一步降低,而添加Ca O后土壤酸化得以缓解(p H值4.10—4.46),土壤硝化潜势(0.22—0.34μg NO-2-N g-1土h-1)显著增加。同时,添加Ca O处理对氨氧化古菌(AOA)的群落结构无明显影响,但明显提高了各施肥处理土壤中氨氧化细菌(AOB)的群落多样性,加Ca O处理的土壤中,AOA的数量降低而AOB的数量增加。这些结果表明虽然酸性土壤中AOA在数量和活性上占主导优势,AOB在功能上冗余,但当添加Ca O后,AOA和AOB对环境变化迅速作出响应,并根据其不同的生态位需求重新分配优势地位,二者交替作用共同驱动酸性土壤硝化作用。     

牦牛排泄物输入对滇西北高寒泥炭沼泽湿地土壤氮转化的影响

牦牛放牧对滇西北高寒湿地土壤环境产生严重影响，改变土壤氮的迁移转化过程，影响湿地生态系统初级生产。然而，关于牦牛排泄物输入对滇西北高寒泥炭沼泽湿地土壤氮转化过程的影响尚不清楚。本研究以滇西北高原典型泥炭沼泽湿地为对象，采用原位土芯室内控制实验方法，研究牦牛排泄物输入对泥炭沼泽湿地土壤氮转化的影响。结果表明，粪便和尿液输入初期促进土壤铵态氮(NH₄⁺-N)积累，但整个培养期则表现为消耗NH₄⁺-N,积累硝态氮(NO₃^--N),表明该过程以硝化作用为主。粪便和尿液输入提高土壤脲酶活性(P<0.05),降低反硝化酶活性(P<0.05)。粪便输入提高过氧化氢酶活性(P<0.05)和N-乙酰氨基葡萄糖苷酶活性(P<0.05),尿液输入降低N-乙酰氨基葡萄糖苷酶活性(P<0.05)。粪便输入对土壤的矿化和硝化作用无显著性影响，尿液输入对土壤硝化作用影响显著。粪便输入抑制土壤反硝化作用，而尿液输入促进反硝化作用。牦牛排泄物输入通过影响泥炭沼...     

小兴安岭酸性泥炭土硝化作用的类型及其驱动因子

以小兴安岭酸性森林泥炭土为研究对象,通过加入10 mL·L^-1乙炔及不同浓度的外源硫酸铵(0、1.2、6.0 mmol N·kg^-1)进行硝化培养试验,探究酸性泥炭土中硝化作用类型及主要驱动因子.结果表明:无论有无外加氮源,酸性泥炭土均存在较强的矿化作用(0.9～1.4 mg N·kg^-1·d^-1),经过2周的培养均发生了硝化作用(0.4～0.6 mg N·kg^-1·d^-1),且不同浓度硫酸铵处理之间无显著差异;而乙炔处理虽有较强的矿化作用(0.8 mg N·kg^-1·d^-1),但未发生明显的硝化作用(0 mg N·kg^-1·d^-1),说明该酸性泥炭土以自养硝化为主,外源无机氮源浓度对硝化作用无显著影响,硝化底物NH₃的主要来源不是外源硫酸铵,更可能来源于土壤中有机氮的矿化.培养0～14 d,无论有无外加氮源,酸性泥炭土氨氧化细菌(AOB)和古菌(AOA)丰度均显著增加,但不同浓度硫酸铵处理间无显著差异,表明外源无机氮浓度对氨氧化微生物的生长无显著促进作用.与不加乙炔的对照相比,乙炔处理AOB和AOA丰度随时间均无显著变化,推测AOA与AOB在该酸性泥炭土的硝化过程中都可能起一定的作用.     

Changes in N-transforming archaea and bacteria in soil during the establishment of bioenergy crops

Widespread adaptation of biomass production for bioenergy may influence important biogeochemical functions in the landscape, which are mainly carried out by soil microbes. Here we explore the impact of four potential bioenergy feedstock crops (maize, switchgrass, Miscanthus X giganteus, and mixed tallgrass prairie) on nitrogen cycling microorganisms in the soil by monitoring the changes in the quantity (real-time PCR) and diversity (barcoded pyrosequencing) of key functional genes (nifH, bacterial/archaeal amoA and nosZ) and 16S rRNA genes over two years after bioenergy crop establishment. The quantities of these N-cycling genes were relatively stable in all four crops, except maize (the only fertilized crop), in which the population size of AOB doubled in less than 3 months. The nitrification rate was significantly correlated with the quantity of ammonia-oxidizing archaea (AOA) not bacteria (AOB), indicating that archaea were the major ammonia oxidizers. Deep sequencing revealed high diversity of nifH, archaeal amoA, bacterial amoA, nosZ and 16S rRNA genes, with 229, 309, 330, 331 and 8989 OTUs observed, respectively. Rarefaction analysis revealed the diversity of archaeal amoA in maize markedly decreased in the second year. Ordination analysis of T-RFLP and pyrosequencing results showed that the N-transforming microbial community structures in the soil under these crops gradually differentiated. Thus far, our two-year study has shown that specific N-transforming microbial communities develop in the soil in response to planting different bioenergy crops, and each functional group responded in a different way. Our results also suggest that cultivation of maize with N-fertilization increases the abundance of AOB and denitrifiers, reduces the diversity of AOA, and results in significant changes in the structure of denitrification community.     

Community composition of ammonia-oxidizing bacteria and archaea in soils under stands of red alder and Douglas fir in Oregon

This study determined nitrification activity and nitrifier community composition in soils under stands of red alder (Alnus rubra) and Douglas fir (Pseudotsuga menziesii) at two sites in Oregon. The H.J. Andrews Experimental Forest, located in the Cascade Mountains of Oregon, has low net N mineralization and gross nitrification rates. Cascade Head Experimental Forest, in the Coast Range, has higher net N mineralization and nitrification rates and soil pH is lower. Communities of putative bacterial [ammonia-oxidizing bacteria (AOB)] and archaeal [ammonia-oxidizing archaea (AOA)] ammonia oxidizers were examined by targeting the gene amoA, which codes for subunit A of ammonia monooxygenase. Nitrification potential was significantly higher in red alder compared with Douglas-fir soil and greater at Cascade Head than H.J. Andrews. Ammonia-oxidizing bacteria amoA genes were amplified from all soils, but AOA amoA genes could only be amplified at Cascade Head. Gene copy numbers of AOB and AOA amoA were similar at Cascade Head regardless of tree type (2.3-6.0 x 10(6)amoA gene copies g(-1) of soil). DNA sequences of amoA revealed that AOB were members of Nitrosospira clusters 1, 2 and 4. Ammonia-oxidizing bacteria community composition, determined by terminal restriction fragment length polymorphism (T-RFLP) profiles, varied among sites and between tree types. Many of the AOA amoA sequences clustered with environmental clones previously obtained from soil; however, several sequences were more similar to clones previously recovered from marine and estuarine sediments. As with AOB, the AOA community composition differed between red alder and Douglas-fir soils.     

Quantitative analyses of the abundance and composition of ammonia-oxidizing bacteria and ammonia-oxidizing archaea of a Chinese upland red soil under long-term fertilization practices

The abundance and composition of soil ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) were investigated by using quantitative real-time polymerase chain reaction, cloning and sequencing approaches based on amoA genes. The soil, classified as agri-udic ferrosols with pH (H(2)O) ranging from 3.7 to 6.0, was sampled in summer and winter from long-term field experimental plots which had received 16 years continuous fertilization treatments, including fallow (CK0), control without fertilizers (CK) and those with combinations of fertilizer nitrogen (N), phosphorus (P) and potassium (K): N, NP, NK, PK, NPK and NPK plus organic manure (OM). Population sizes of AOB and AOA changed greatly in response to the different fertilization treatments. The NPK + OM treatment had the highest copy numbers of AOB and AOA amoA genes among the treatments that received mineral fertilizers, whereas the lowest copy numbers were recorded in the N treatment. Ammonia-oxidizing archaea were more abundant than AOB in all the corresponding treatments, with AOA to AOB ratios ranging from 1.02 to 12.36. Significant positive correlations were observed among the population sizes of AOB and AOA, soil pH and potential nitrification rates, indicating that both AOB and AOA played an important role in ammonia oxidation in the soil. Phylogenetic analyses of the amoA gene fragments showed that all AOB sequences from different treatments were affiliated with Nitrosospira or Nitrosospira-like species and grouped into cluster 3, and little difference in AOB community composition was recorded among different treatments. All AOA sequences fell within cluster S (soil origin) and cluster M (marine and sediment origin). Cluster M dominated exclusively in the N, NP, NK and PK treatments, indicating a pronounced difference in the community composition of AOA in response to the long-term fertilization treatments. These findings could be fundamental to improve our understanding of the importance of both AOB and AOA in the cycling of nitrogen and other nutrients in terrestrial ecosystems.     

长期施肥对土壤氨氧化微生物的影响

长期施肥可改变土壤碳氮等养分供应,进而影响微生物数量与群落组成。本研究基于棕壤长期定位实验站,分析不同施肥方式下(不施肥,CK;低量无机氮肥,N₂;高量无机氮肥,N₄;有机无机氮肥配施,M₂N₂)土壤氨氧化古菌(AOA)和细菌(AOB)的变化,为土壤氮素转化的微生物学机制和培肥土壤提供依据。结果表明:不同施肥方式下,土壤AOA与AOB的数量比值为2.28～61.95。与CK相比,施肥后土壤AOA数量降低了1.6%～13.6%。N₄处理AOB数量随土壤深度增加呈先降低后升高的趋势,其他处理则相反。土壤AOB群落Shannon多样性指数、均匀度指数和Simpson指数均高于AOA。M₂N₂处理0～20 cm土层土壤AOB多样性增加,但AOA多样性降低。土壤AOB主要因土壤深度发生聚类,AOB和AOA均未因施肥方式发生聚类。综上,长期施肥改变了土壤AOA和AOB的构成状况,AOA对环境变化较为敏感,AOB较为丰富和稳定。     

Rapid and dissimilar response of ammonia oxidizing archaea and bacteria to nitrogen and water amendment in two temperate forest soils

Biochemical processes relevant to soil nitrogen (N) cycling are performed by soil microorganisms affiliated with diverse phylogenetic groups. For example, the oxidation of ammonia, representing the first step of nitrification, can be performed by ammonia oxidizing bacteria (AOB) and, as recently reported, also by ammonia oxidizing archaea (AOA). However, the contribution to ammonia oxidation of the phylogenetically separated AOA versus AOB and their respective responsiveness to environmental factors are still poorly understood. The present study aims at comparing the capacity of AOA and AOB to momentarily respond to N input and increased soil moisture in two contrasting forest soils. Soils from the pristine Rothwald forest and the managed Schottenwald forest were amended with either NH(4)(+)-N or NO(3)(-)-N and were incubated at 40% and 70% water-filled pore space (WFPS) for four days. Nitrification rates were measured and AOA and AOB abundance and community composition were determined via quantitative PCR (qPCR) and terminal restriction length fragment polymorphism (T-RFLP) analysis of bacterial and archaeal amoA genes. Our study reports rapid and distinct changes in AOA and AOB abundances in the two forest soils in response to N input and increased soil moisture but no significant effects on net nitrification rates. Functional microbial communities differed significantly in the two soils and responded specifically to the treatments during the short-term incubation. In the Rothwald soil the abundance and community composition of AOA were affected by the water content, whereas AOB communities responded to N amendment. In the Schottenwald soil, by contrast, AOA responded to N addition. These results suggest that AOA and AOB may be selectively influenced by soil and management factors.     

施用生物质炭对采煤塌陷区土壤氨氧化微生物丰度和群落结构的影响

生物质炭作为一种新型土壤改良剂,施入土壤不仅能提高肥力,改善土壤结构,还能够影响土壤氮素的转化.本文利用培养试验研究施用生物质炭对采煤塌陷区土壤性质及氨氧化菌丰度和群落结构的影响.结果表明: 生物质炭显著提高土壤铵氮(NH4⁺-N)、全氮、有效磷和速效钾含量.生物质炭施用量对氨氧化古菌(AOA)丰度没有显著影响,但是增加施用量显著提高了氨氧化细菌(AOB)丰度.对T-RFLP数据进行分析发现,生物质炭提高了AOA和AOB多样性,并在一定程度上改变了AOA和AOB群落结构.施用生物质炭提高了采煤塌陷区土壤养分含量,并在一定程度上提高了氨氧化菌的丰度和多样性,表明生物质炭对塌陷区复垦土壤具有培肥改良的潜能.