不同酸碱性紫色土的硝化活性及微生物群落组成

为研究紫色土中的硝化作用、总微生物和硝化微生物的群落结构,以重庆永川的酸性紫色土(p H=5.3)和中性紫色土(p H=7.2)以及四川盐亭的石灰性紫色土(p H=8.5)为研究对象,采用稳定性同位素标记技术进行培养实验,并通过Miseq测序对三种紫色土微生物群落结构进行分析。每种土样共设有三种处理,包括~(13)CO_2标记处理、~(12)CO_2对照处理和~(13)CO_2+C_2H_2对照处理。结果表明,中性紫色土(p H=7.2)和石灰性紫色土(p H=8.5)经过56 d的培养后发生了强烈的硝化作用,而酸性紫色土(p H=5.3)中未发生明显的硝化作用,并且硝化作用类型都以自养硝化为主。三种紫色土中都存在着变形菌门(Proteobacteria)、酸杆菌门(Acidobacteria)、厚壁菌门(Firmicutes)、绿弯菌门(Chloroflexi)、放线菌门(Actinobacteria)、拟杆菌门(Bacteroidetes)和芽单胞菌门(Gemmatimonadetes),其中变形菌门在三种紫色土中都大约占有20%的比例。中性紫色土和石灰性紫色土各处理中硝化螺旋菌门(Nitrospirae)百分比大于酸性紫色土各处理。三种紫色土各处理中的氨氧化细菌(Ammonia-oxidizing bacteria,AOB)主要以亚硝化螺菌属(Nitrosospira)为主,亚硝酸氧化细菌(Nitrite-oxidizing bacteria,NOB)主要以硝化螺菌属(Nitrospira)为主。且NOB/AOB的值在三种紫色土各处理中最高可达到13,这意味着全程氨氧化细菌(Comammox)可能在紫色土的硝化作用中占据重要贡献。     

模拟条件下土壤硝化作用及硝化微生物对不同水分梯度的响应

以江苏滨海县一植稻土壤为研究对象,在微宇宙培养条件下设置了不同水分处理(最大持水量的30%、60%、90%和淹水2 cm深),研究了硝化作用及硝化微生物对水分变化的响应特征。结果表明:淹水处理显著降低了土壤的氧化还原电位(Eh),但所有处理土壤Eh变化范围为330~500 m V,土壤整体处于氧化态。在每7天向土壤加入10 mg kg-1NH+4-N的连续培养过程中,各个水分处理均观察到明显的NH+4-N降低和NO-3-N累积的现象,60%WHC处理下土壤硝态氮累积最显著和迅速,90%WHC处理次之,随培养时间延长,30%WHC和淹水处理也观察到明显的硝化作用。淹水处理中氨氧化细菌(Ammonia-oxidizing bacteria,AOB)的数量显著高于非淹水处理,且淹水处理中AOB在DGGE图谱上的条带更加清晰明亮,而氨氧化古菌(Ammonia-oxidizing archaea,AOA)的群落组成和数量在不同水分处理间无明显变化。表明该土壤中AOB对水分条件变化响应灵敏,是该土壤的硝化作用、尤其是淹水条件下硝化作用发生的主要原因。     

O_3浓度升高对麦田土壤氨氧化细菌、氨氧化古菌和硝化细菌数量的影响

硝化作用在氮循环过程中至关重要,包括氨氧化作用和亚硝酸盐氧化作用,通过氨氧化反应和亚硝酸盐氧化反应将N素转化为植物可利用的NO;形态。利用开顶式臭氧气室（OTCs,open-topchambers）试验平台,通过大田模拟熏气试验,结合Real-timePCR探讨大气O,浓度升高对麦田土壤氨氧化细菌（AOB）、氨氧化古菌（AOA）及硝化细菌（NOB）数量的影响。结果表明,AOB、AOA和NOB对O,胁迫的反应不一样,AOB基因拷贝数基本上随着O,浓度的升高呈现出降低的趋势,而AOA和NOB基因拷贝数随O_3浓度的升高变化不明显。冬小麦拔节期,当O_3浓度为40、80、120nmol·mol。时,20-40em土层的AOB基因拷贝数分别比对照处理降低39．8％、51．2％和59．4％。AOB和NOB基因拷贝数灌浆期多于收获期,0-10cm土层多于10-20em。AOA基因拷贝数随季节的变化不大。O_3胁迫可通过影响AOB、AOA和NOB的数量和活性来影响土壤的硝化反应,从而影响土壤的氮素循环过程。     

有机碳氮添加对酸性森林土壤氨氧化过程的影响

以亚热带酸性森林土壤为研究对象,开展了微宇宙室内培养实验,设置了有机碳和有机氮添加处理,分析了土壤硝化活性和氨氧化古菌(Ammonia-oxidizing archaea,AOA)、氨氧化细菌(Ammonia-oxidizing bacteria,AOB)的功能基因丰度,研究了外源有机碳和有机氮对酸性森林土壤氨氧化过程的影响规律。结果表明:外源有机氮添加显著刺激了酸性森林土壤硝化活性,乙炔抑制实验表明自养氨氧化对酸性森林土壤硝化过程的贡献率90%。有机碳添加对土壤硝化活性未有显著影响,同时添加有机碳和无机铵态氮也未显著提高土壤硝化活性,而外源有机氮添加提高了土壤矿化速率并导致土壤NH3浓度升高,可能是土壤硝化活性、AOA和AOB数量显著增加的主要原因。     

长期施用含氯化肥对棕壤硝化作用及氨氧化微生物的影响

【目的】氨氧化微生物是氨氧化过程的主要驱动者,氨氧化过程作为硝化作用的限速步骤对氮循环具有重要作用。本研究以沈阳农业大学棕壤含氯化肥长期定位试验的土壤为研究对象,探讨了连续34年施用高氯和低氯化肥对棕壤硝化作用及氨氧化微生物的影响。【方法】该长期试验在等量氮、磷、钾条件下,设置高氯和低氯处理,共8个处理:T1(不施肥);T2(单施尿素);T3(尿素+氯化钾);T4(尿素+过磷酸钙);T5(尿素+过磷酸钙+氯化钾);T6(尿素+磷酸一铵+氯化钾);T7(尿素+氯磷铵+氯化钾);T8(硝酸磷肥+过磷酸钙+氯化钾),T7为高氯处理。采集0—20cm土壤样品,利用荧光定量PCR技术测定氨氧化细菌(AOB)和古菌(AOA)丰度,并结合土壤硝化潜势和基本化学性质,分析长期施用含氯化肥对棕壤硝化作用及氨氧化微生物丰度的影响及影响氨氧化微生物丰度的主要环境因素。【结果】长期施肥降低了土壤pH值,高氯处理降低得最多,显著低于其他处理;高氯处理的土壤硝化潜势也显著低于其他处理,且除高氯处理外,配施磷肥的处理土壤硝化潜势显著高于不施磷处理。各处理土壤中AOA丰度均显著高于AOB,高氯处理土壤中AOA、AOB丰度均显著低于其他处理,土壤硝化潜势与AOA和AOB均呈显著正相关关系。【结论】连续施用高氯化肥34年显著降低了棕壤AOA和AOB丰度,抑制了硝化潜势。该结果可为通过含氯化肥的合理施用来调节土壤AOA和AOB,进而调控土壤氮素循环提供参考。     

长期定位施肥对中性紫色土硝化作用及氨氧化微生物的影响

为了研究长期不同施肥措施对中性紫色土氨氧化微生物及其硝化作用的影响,以国家紫色土肥力与肥料效益监测基地的中性紫色土为研究对象,进行土壤氨氧化细菌和氨氧化古菌amo A基因的Real-time PCR分析,比较长期不同定位施肥对土壤氨氧化潜势和硝化强度的影响,并分析不同施肥制度对功能微生物丰度与功能的作用。数据显示,土壤中氨氧化古菌amo A基因拷贝数(Log值6.21～7.14)远大于氨氧化细菌(Log值3.65～5.73),相对于对氨氧化细菌丰度的影响,施肥对土壤氨氧化古菌丰度影响较小。施用氮肥与磷肥都显著提高了土壤氨氧化细菌丰度,1.5NPK+M处理氨氧化细菌丰度最高(Log值5.73),有机无机肥配施可以显著提高土壤氨氧化微生物丰度;而含氯化肥的施用在一定程度上降低了土壤氨氧化细菌丰度与硝化细菌生长,与施用不含氯的肥料处理相比,含氯肥料处理的土壤氨氧化细菌丰度与硝化细菌数分别降低了3.74%和88.12%。研究表明,长期施肥能影响中性紫色土中氨氧化细菌的丰度,有机无机肥配施能够提高土壤的氨氧化潜力与土壤的硝化能力。     

青藏高原三种典型生境中硝化微生物分布和群落结构

以青藏高原3种典型生境(荒漠草地、湿地和盐碱地)为研究对象，通过amoA功能基因qPCR和16SrRNA基因扩增子测序，研究了其氨氧化古菌(ammonia-oxidizingarchaea,AOA)、氨氧化细菌(ammonia-oxidizingbacteria,AOB)、亚硝酸盐氧化菌(nitrite-oxidizing bacteria,NOB)和完全硝化细菌(complete ammonia oxidizer,CMX)的分布与群落结构特征。qPCR结果表明，荒漠草地和盐碱地中3种氨氧化微生物，其丰度顺序为AOA>CMX>AOB，而在湿地中则为CMX≥AOA>AOB。各生境中AOA主要类群为土壤类群(即group 1.1b)，且其中约半数属于Nitrosocosmicus属分支，该分支在北极冻土中也有分布。荒漠草地和湿地中AOB主要为亚硝化螺菌属(Nitrosospira spp.,71.21%～100%)，而盐碱地主要为亚硝化单胞菌属(Nitrosomonas spp.,75.51%～88.71%)。3种生境中NOB主要类群均为硝化螺菌属(Nitrospira ...     

中性紫色水稻土硝化作用中细菌和古菌的相对贡献

不同的硝化抑制剂对硝化作用的强度以及氨氧化古菌(ammonia-oxidizing archeae,AOA)和氨氧化细菌(ammonia-oxidizing bacteria,AOB)丰度可以产生不同的影响,因此可推算AOA和AOB在硝化作用中的相对贡献。本研究选取硝化过程中AOA和AOB共同起作用的中性紫色土为研究对象,分析不同硝化抑制剂对土壤硝化势以及AOB和AOA的丰度的影响。结果发现,烯丙基硫脲(Allylthiourea,ATU)、2-苯基-4,4,5,5,-四甲基咪唑烷-1-氧-3-氧化物(2-carboxyphenyl-4,4,5,5-tetramethylimidazoline-1-oxyl3-oxide,PTIO)、辛炔(Octyne、C_8)均可显著降低影响土壤的硝化作用。PTIO显著降低AOA的丰度(约40%),对AOB没有显著影响;ATU和C_8显著降低AOB的丰度(约50%),对AOA没有显著影响。通过计算,该土中AOA与AOB对硝化作用的相对贡献率分别为7.5%和55.6%。结果表明,在中性紫色土中AOA与AOB共同推动硝化作用,其中AOB起主导作用。     

秸秆直接还田与炭化还田对潮土硝化微生物的影响

为比较秸秆直接还田和炭化还田对黄淮海平原小麦/玉米轮作体系典型潮土硝化作用及硝化微生物群落的影响,设置4个处理:全量小麦秸秆还田(S)、全量秸秆炭化还田(B)、半量秸秆半量生物质炭还田(SB)和不进行秸秆或生物质炭还田的对照(CK),连续进行3 a田间试验。对小麦、玉米两个生长季土壤理化性质进行分析,用末端限制性片段长度多态性(Terminal-restriction length polymorphism,T-RFLP)技术和克隆文库技术对氨氧化古菌(Ammonia-oxidizing archaea,AOA)和氨氧化细菌(Ammonia-oxidizing bacteria,AOB)群落结构和多样性进行分析。结果表明,在小麦季,与S处理相比,B处理显著降低了土壤容重,提高了土壤pH、有机碳(SOC)和速效钾(AK)含量(P0.05),但并未显著影响土壤水分、铵态氮(NH_4~+-N)和硝态氮(NO_3~--N)含量;B和SB处理的硝化潜势(Potential nitrification rate,PNR)分别为0.58、0.49μg·h~(-1)·g~(-1)(以NO_2~-计,下同),显著高于CK,与S处理(0.40μg·h~(-1)·g~(-1))差异不显著。玉米季,B处理显著提高了土壤水分、SOC和AK(P0.05),各处理玉米季的PNR整体低于小麦季,B处理最高(0.27μg·h~(-1)·g~(-1)),显著高于CK和S处理(P0.05)。小麦季PNR分别与AK、NH_4~+浓度和土壤容重显著相关(P0.05),与AOA和AOB群落组成均无显著关系;玉米季PNR仅与理化因子SOC显著相关,但该季节PNR与AOB群落结构显著相关。冗余分析(RDA)表明,土壤SOC、容重、pH和AK是显著影响硝化微生物群落结构的主要因子,对AOA和AOB群落结构总变异的解释量分别为76.4%和75.5%。系统发育树分析表明,AOA大部分属于土壤古菌Group1.1b,AOB多属于亚硝化螺菌Nitrosospira簇3。综上,与秸秆直接还田相比,炭化还田提高土壤硝化活性,改善部分土壤理化性质,引起土壤硝化微生物群落结构变化。     

水氮耦合对设施农田土壤自养和异养硝化作用的影响

水氮措施影响设施土壤氮素的转化及硝化微生物活性,但水氮耦合对设施土壤自养和异养硝化作用差异的影响尚不明确。以连续8年设施水氮耦合田间定位试验土壤为研究对象,控制不同土壤田间持水量(WHC)(40%WHC、60%WHC和80% WHC)进行室内微宇宙培养试验,通过添加乙炔抑制剂抑制自养硝化途径,研究水氮耦合对设施土壤自养和异养硝化速率及参与自养硝化的氨氧化微生物的影响,分析氨氧化微生物氨氧化古细菌(AOA)和氨氧化细菌(AOB)对自养硝化作用的贡献。结果表明,水氮耦合下,不同硝化途径NH4+-N、NO^3--N含量以及参与自养硝化的AOA amoA和AOB amoA基因拷贝数均有显著差异。无乙炔培养7 d后,NO^3--N含量显著增加,而NH4+-N含量显著降低,AOA amoA和AOB amoA的基因丰度显著增加。添加乙炔后,NO^3--N、NH4+-N含量基本保持恒定,AOA amoA和AOB amoA基因丰度显著减少。水氮耦合显著影响自养和异养硝化速率,冗余分析(RDA)表明,NH4+-N含量、AOB amoA、NO^3--N-C2H2、AOA amoA可分别解释自养和异养硝化速率变异的68.9%、34.9%、32.8%和24.4%。设施土壤存在自养硝化和异养硝化两种途径,60%～80%WHC各施氮处理均以自养硝化为主,占总硝化速率的65%～86%;仅40%WHC下,氮纯养分量300和525 kg·hm-2处理以异养硝化为主,占总硝化速率的61%～77%。AOB和AOA共同驱动自养硝化,且AOB贡献更大。     

The role of ammonium oxidizing communities in mediating effects of an invasive plant on soil nitrification

Invasive plants often benefit from changes that they impose on soil microbes via positive plant–soil feedback, but the mechanisms that underlie these changes, and the legacy of their effects, remain poorly quantified. We investigated the impacts of an invasive annual grass, Microstegium vimineum, on the structure and functioning of soil microbial communities in a multi-year, field-based common garden experiment. Given previous reports that M. vimineum can both elevate nitrification rates in soil and benefit from enhanced nitrate availability, we sought to answer the following questions: 1) Does M. vimineum alter the abundance or composition of soil nitrifying microbial communities (ammonia oxidizing archaea and bacteria, AOA and AOB, respectively)? 2) Are such effects reversible or do soil legacy effects persist after M. vimineum is no longer present? After three years, invaded plots had greater AOA abundances than uninvaded native dominated plots, as well as different AOA community structure. However, after seven years, and following a period of M. vimineum replacement by native plants in the invaded plots, AOA abundances and nitrification rates declined toward levels found in uninvaded plots. Collectively, our results suggest that while the impacts of M. vimineum invasions on nitrogen cycling likely relate to their association with AOA, these effects may not persist if M. vimineum declines over time and native plants and their associated microbes are able to re-establish.     

Effects of inorganic fertilizer and manure on soil archaeal abundance at two experimental farms during three consecutive rotation-cropping seasons

Soil archaeal population dynamics at two experimental sites of the same clay-loam type in Ottawa and Woodslee, Ontario, were investigated to determine fertilizer and manure effects following their different long-term crop rotation and fertilization schemes. Phylogenetic analysis of cloned soil archaeal 16S rRNA gene libraries of both sites identified them with group 1.1b of Thaumarchaeota. The gene population dynamics subtly varied in the order of 10⁷ copies g⁻¹ soil when monitored by quantitative real-time PCR during three growing seasons (2007–2009). In Ottawa, where plots were amended with dairy-farm manure, soil thaumarchaeal gene abundance was double of the unamended plots. At the Woodslee N-P-K-fertilized plots, it remained at least 30% fewer than that of the unfertilized ones. These cultivated plots showed soil carbon limitation while the fertilized ones were low in soil pH (ca. 5.5). Surface soils from an unfertilized sod plot and an adjacent deciduous forest had higher total carbon content (C:N ratio of 9 and 11, respectively). Their thaumarchaeal gene abundance varied up to 4.8 × 10⁷ and 7.0 × 10⁷ copies g⁻¹ soil, respectively. The former value was also attained at the manure-amended plots in Ottawa, where the C:N ratio was just below 10. Where soil pH was above 6.0, there was a weak and positive correlation between soil total C and the estimated gene abundance. Such gene population dynamics consistently demonstrated the stimulating and suppressive effects of dairy-farm manure (Ottawa site) and inorganic fertilizers (Woodslee site), respectively, on soil thaumarchaea. At both sites archaeal amoA and 16S rRNA gene abundance were similarly affected. Archaeal amoA gene abundance also outnumbered bacterial amoA abundance, suggesting that ammonia-oxidizing archaea might be dominant in these soils. Only minor crop effects on gene population dynamics were detected.     

Ammonia oxidizer abundance in paddy soil profile with different fertilizer regimes

Studies about ammonia-oxidizing bacteria (AOB) and archaea (AOA) are often focused on topsoil, but little is known about their activity and distribution in subsoil. A long-term fertilizer experiment was conducted to assess the effects of different fertilizer treatments on AOB and AOA in vertical soil profiles of paddy soil plots that received no nitrogen fertilizer control (CK), NPK chemical fertilizers (CF), organic–inorganic mixed fertilizer (OIMF) and organic fertilizer (OF). Soil properties, potential nitrification rate (PNR) and amoA gene abundance of AOB and AOA were measured and analyzed by two-way ANOVA and correlation analysis. Quantitative PCR analysis of amoA genes showed that AOA were more abundant than AOB in all the soil samples. AOB declined sharply with soil depth. Compared with CK and OF treatments, CF and OIMF treatments had higher abundance of AOB throughout the soil profiles. However, AOA tend less responsive to soil depth and fertilizers compared to AOB. This caused the AOA/AOB ratios in subsoil higher than in topsoil, and in CK and OF higher than in CF and OIMF treatments. These results suggest that AOA are more abundant and can be better adapted to nutrient-poor subsoils than AOB, and autotrophic nitrification could likely be determined by a complex suite of environmental factors in vertical profiles of the paddy soil tested.     

pH regulates key players of nitrification in paddy soils

Increasing lines of evidence have suggested the functional importance of ammonia-oxidizing archaea (AOA) rather than bacteria (AOB) for nitrification in upland soils with low pH. However, it remains unclear whether niche specialization of AOA and AOB occurs in rice paddy wetlands constrained by oxygen availability. Using DNA-based stable isotope probing, we conclude that AOA dominated nitrification activity in acidic paddy soils (pH 5.6) while AOB dominated in alkaline soils (pH 8.2). Nitrification activity was stimulated by urea fertilization and accompanied by a significant increase of AOA in acid soils and AOB in alkaline soils. DNA-based stable isotope probing indicated significant assimilation of ¹³CO₂ for AOA only in acidic paddy soil, while AOB was the solely responsible for ammonia oxidation in the alkaline paddy soil. Phylogenetic analysis further indicated that AOA members within the soil group 1.1b lineage dominated nitrification in acid soils. Ammonia oxidation in the alkaline soil was catalyzed by Nitrosospira cluster 3-like AOB, suggesting that the physiological diversity of AOA is more complicated than previously thought, and soil pH plays important roles in shaping the community structures of ammonia oxidizers in paddy field.     

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。     

土壤亚硝酸气体(HONO)排放过程及其驱动机制

气态亚硝酸(HONO)是大气中氢氧自由基(OH·)的重要来源,直接影响到大气氧化能力和空气质量。通过比较外场测定和模型计算的HONO浓度,发现白天时存在未知的大气HONO来源。研究表明,土壤可以向大气中排放HONO。其机理可能是土壤亚硝态氮和氢离子的化学平衡作用;或土壤夜间吸附和白天解吸附的动态物理化学过程;或氨氧化细菌等微生物的直接排放;也可能是硝化过程中产生的羟胺,在土壤颗粒物等表面的化学反应。因此,土壤HONO排放通量与土壤亚硝态氮浓度、pH、氨氧化细菌丰度、土壤矿物、土壤湿度及C/N值等相关。目前对于土壤HONO排放的研究尚在起步阶段,国内亦少见相关成果报道。本文综述了土壤HONO排放的研究背景、探讨了土壤HONO排放的机理及影响因素,以期为减少氮素损失、提高氮肥利用率、评估氮肥的环境效应及城市空气质量等提供理论依据和科学指导。     

海水稻根际效应对滨海盐碱地土壤氨氧化微生物的影响

滨海盐碱地的特殊环境严重限制了土壤氮素转化和利用。微生物介导的水稻根际氨氧化过程是盐碱稻田土壤氮循环的关键过程,但限于研究盲点和技术不足,海水稻根际效应对滨海盐碱地土壤氨氧化微生物群落结构的影响仍少有报道。据此,本研究以“海稻86”为研究对象,分别设置低盐浓度(2 g·kg^–1)和高盐浓度(6 g·kg–1)两组处理进行盆栽试验。结果显示：种植海水稻70 d后,高盐和低盐处理根际土壤的pH分别下降了0.82和0.70个单位,土壤有机质(SOM)含量下降了6.41和4.46 g·kg^–1,腐殖质(HU)含量提高了5.76和4.45 g·kg^–1,全氮(TN)含量减少0.46和0.37 g·kg^–1,表明海水稻可通过降低盐碱地土壤pH,加速有机质分解转化,提高土壤氮循环速率。水稻根际作用可显著提高土壤微生物生物量碳(MBC)、微生物生物量氮(MBN)和微生物呼吸强度,并在种植第55天达到最高,在高盐处理中分别达到850.0 mg·kg^–1、72.2 mg·kg^–1