Chronic Nitrogen Fertilization Modulates Competitive Interactions Among Microbial Ammonia Oxidizers in a Loess Soil

Nitrogen(N) application may lead to niche segregation of soil ammonia-oxidizing archaea(AOA) and bacteria(AOB), thereby reducing the competitive interactions between AOA and AOB due to higher ammonium substrate availability. However, the adaptive mechanisms of AOA and AOB under N enrichment remain poorly understood. Stable isotope probing(SIP) microcosm incubation was employed to reveal community changes of active AOA and AOB in a loess soil from a field experiment growing potatoes that received no N(control, CK), low N(LN, 75 kg N ha~(-1)), and high N(HN, 375 kg N ha~(-1)). The results showed that the soil potential nitrification rate(PNR) was measured by culturing of the soil samples from the field experiment. Soil PNR was significantly increased in HN by87.5% and 67.5% compared with CK and LN, respectively. Compared with CK, the~(13)C-amoA genes of soil AOA and AOB in HN had 2.58 × 10~4 and 1.55 × 10~6 copies, representing 1.6-and 16.2-fold increase respectively. It was indicated that AOB dominated soil ammonia oxidation. A phylogenetic analysis of the~(13)C-amoA gene showed that N application significantly increased the proportion of54 d9-like AOA up to 90% in HN, while the Nitrososphaera gargensis-like and Nitrososphaera viennensis-like AOA were inhibited and completely disappeared. Nitrogen application also resulted in the community shift of active AOB-dominant group from Nitrosospira briensis-like to Nitrosospira sp. TCH711-like. Our study provides compelling evidence for the emergence and maintenance of active nitrifying communities under the intensified N input to an agricultural ecosystem.     

两种农田土壤中的氨氧化细菌和氨氧化古菌对硝化抑制剂DCD和DMPP的响应-盆栽试验

Taking two important agricultural soils with different pH, brown soil （Hap-Udic Luvisol） and cinnamon soil （Hap-Ustic Luvisol）, from Northeast China, a pot culture experiment with spring maize （Zea mays L.） was conducted to study the dynamic changes in the abundance and diversity of soil ammonia-oxidizing bacteria （AOB） and ammonia-oxidizing archaea （AOA） populations during maize growth period in response to the additions of nitrification inhibitors dicyandiamide （DCD） and 3,4-dimethylpyrazole phosphate （DMPP） by the methods of real-time polymerase chain reaction （PCR） assay, PCR-denaturing gradient gel electrophoresis （DGGE）, and construction of clone library targeting the amoA gene. Four treatments were established, i.e., no urea （control）, urea, urea plus DCD, and urea plus DMPP. Both DCD and DMPP inhibited growth of AOB significantly, compared to applying urea alone. Soil bacterial amoA gene copies had a significant positive linear correlation with soil nitrate content, but soil archaeal amoA gene copies did not. In both soils, all AOB sequences fell within Nitrosospira or Nitrosospira-like groups, and all AOA sequences belonged to group 1.1b crenaxchaea. With the application of DCD or DMPP, community composition of AOB and AOA in the two soils had less change except that the AOB community composition in Hap-Udic Luvisol changed at the last two growth stages of maize under the application of DCD. AOB rather than AOA likely dominated soil ammonia oxidation in these two agricultural soils.     

长期施肥黑土微生物区系及功能多样性的变化

Black soil （Mollisol） is one of the main soil types in northeastern China. Biolog and polymerase chain reactiondenaturing gradient gel electrophoresis （PCR-DGGE） methods were used to examine the influence of various fertilizer combinations on the structure and function of the bacterial community in a black soil collected from Harbin, Heilongjiang Province. Biolog results showed that substrate richness and catabolic diversity of the soil bacterial community were the greatest in the chemical fertilizer and chemical fertilizer＋manure treatments. The metabolic ability of the bacterial community in the manure treatment was similar to the control. DGGE fingerprinting indicated similarity in the distribution of most 16S rDNA bands among all treatments, suggesting that microorganisms with those bands were stable and not influenced by fertilization. However, chemical fertilizer increased the diversity of soil bacterial community. Principal com- ponent analysis of Biolog and DGGE data revealed that the structure and function of the bacterial community were similar in the control and manure treatments, suggesting that the application of manure increased the soil microbial population, but had no effect on the bacterial community structure. Catabolic function was similar in the chemical fertilizer and chemical fertilizer＋manure treatments, but the composition structure of the soil microbes differed between them. The use of chemical fertilizers could result in a decline in the catabolic activity of fast-growing or eutrophic bacteria.     

不同细菌饲喂细菌线虫对土壤细菌数量、活性以及群落结构的影响

The effects of bacterial-feeding nematodes on bacterial number, activity, and community composition were studied through a microcosm experiment using sterilized soil inoculated with soil bacteria （soil suspension） and with bacteria and three species of bacterial-feeding nematodes （ Cephalobus persegnis, Protorhabditis filiformis, and Caenorhabditis elegans）. Catalyzed reporter deposition-fluorescence in situ hybridization, CO2 evolution, and denaturing gradient gel electrophoresis （DGGE） of PCR ampli- fied 16S rRNA gene fragments were used to investigate bacterial numbers, antivity, and community composition, respectively. Our results showed that bacterial numbers and activity significantly increased in the presence of bacterial-feeding nematodes, which indicated that bacterial-feeding nematodes had a significant positive effect on soil bacteria. The different nematode species had different effects on bacterial numbers and activity. C. persegnis and P. filiformis, isolated from native soil, increased the bacterial number and activity more than C. elegans. The DGGE analysis results showed that dominant bacterial species significantly differed among the treatments, which suggested that bacterial-feeding nematode species modified the bacterial community composition in soil. Further gene sequence analysis results showed that the dominant bacterial species in this study were gram-negative bacteria. Given the completely same conditions except nematode species, the varied selective feeding behavior of different nematode species was the most likely reason for the altered bacterial community composition. Overall, the alteration of bacterial numbers, activity and community composition resulting from the bacterial-feeding nematodes may ult!mately affect soil ecological functioning and processes.     

利用PCR-DGGE技术研究长期自然恢复、种植作物和裸地条件下黑土土壤细菌群落结构变化

Soil microbial biomass and community structures are commonly used as indicators for soil quality and fertility. A investigation was performed to study the effects of long-term natural restoration, cropping, and bare fallow managements on the soil microbial biomass and bacterial community structures in depths of 0--10, 20--30, and 40--50 cm in a black soil (Mollisol). Microbial biomass was estimated from chloroform fumigation-extraction, and bacterial community structures were determined by analysis of 16S rDNA using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). Experimental results showed that microbial biomass significantly declined with soil depth in the managements of restoration and cropping, but not in the bare fallow. DGGE profiles indicated that the band number in top 0--10 cm soils was less than that in depth of 20--30 or 40--50 cm. These suggested that the microbial population was high but the bacterial community structure was simple in the topsoil. Cluster and principle component analysis based on DGGE banding patterns showed that the bacterial community structure was affected by soil depth more primarily than by managements, and the succession of bacterial community as increase of soil depth has a similar tendency in the three managements. Fourteen predominating DGGE bands were excised and sequenced, in which 6 bands were identified as the taxa of Verrucomicrobia, 2 bands as Actinobacteria, 2 bands as α-Proteobacteria, and the other 4 bands as δ-Proteobacteria, Acidobacteria, Nitrospira, and unclassified bacteria. In addition, the sequences of 11 DGGE bands were closely related to uncultured bacteria. Thus, the bacterial community structure in black soil was stable, and the predominating bacterial groups were uncultured.     

Responses of Soil Enzyme Activities and Microbial Community Composition to Moisture Regimes in Paddy Soils Under Long-Term Fertilization Practices

The effects of fertilization on activity and composition of soil microbial community depend on nutrient and water availability;however,the combination of these factors on the response of microorganisms was seldom studied.This study investigated the responses of soil microbial community and enzyme activities to changes in moisture along a gradient of soil fertility formed within a long-term(24 years)field experiment.Soils(0–20 cm)were sampled from the plots under four fertilizer treatments:i)unfertilized control(CK),ii)organic manure(M),iii)nitrogen,phosphorus,and potassium fertilizers(NPK),and iv)NPK plus M(NPK+M).The soils were incubated at three moisture levels:constant submergence,five submerging-draining cycles(S-D cycles),and constant moisture content at 40%water-holding capacity(low moisture).Compared with CK,fertilization increased soil organic carbon(SOC) by 30.1%–36.3%,total N by 27.3%–38.4%,available N by 35.9%–56.4%,available P by 61.4%–440.9%,and total P by 28.6%–102.9%.Soil fertility buffered the negative effects of moisture on enzyme activities and microbial community composition.Enzyme activities decreased in response to submergence and S-D cycles versus low moisture.Compared with low moisture,S-D cycles increased total phospholipid fatty acids(PLFAs)and actinomycete,fungal,and bacterial PLFAs.The increased level of PLFAs in the unfertilized soil after five S-D cycles was greater than that in the fertilized soil.Variations in soil microbial properties responding to moisture separated CK from the long-term fertilization treatments.The coefficients of variation of microbial properties were negatively correlated with SOC,total P,and available N.Soils with higher fertility maintained the original microbial properties more stable in response to changes in moisture compared to low-fertility soil.     

北美中部大平原半干旱农业生态系统中覆盖作物和灌溉对土壤微生物群落和酶活性的影响

Cover crops can have beneficial effects on soil microbiology by increasing carbon (C) supply, but these beneficial effects can be modulated by precipitation conditions. The objective of this study was to compare a fallow-winter wheat (Triticum aestivum L.) rotation to several cover crop-winter wheat rotations under rainfed and irrigated conditions in the semiarid US High Plains. Experiments were carried out at two sites, Sidney in Nebraska and Akron in Colorado, USA, with three times of soil sampling in 2012--2013 at cover crop termination, wheat planting, and wheat maturity. The experiments included four single-species cover crops, a 10-species mixture, and a fallow treatment. The variables measured were soil C and nitrogen (N), soil community structure by fatty acid methyl ester (FAME) profiles, and soil β-glucosidase, β-glucosaminidase, and phosphodiesterase activities. The fallow treatment, devoid of living plants, reduced the concentrations of most FAMEs at cover crop termination. The total FAME concentration was correlated with cover crop biomass (R = 0.62 at Sidney and 0.44 at Akron). By the time of wheat planting, there was a beneficial effect of irrigation, which caused an increase in mycorrhizal and protozoan markers. At wheat maturity, the cover crop and irrigation effects on soil FAMEs had subsided, but irrigation had a positive effect on the β-glucosidase and phosphodiesterase activities at Akron, which was the drier of the two sites. Cover crops and irrigation were slow to impact soil C concentration. Our results show that cover crops had a short-lived effect on soil microbial communities in semiarid wheat-based rotations and irrigation could enhance soil enzyme activity. In the semiarid environment, longer time spans may have been needed to see beneficial effects of cover crops on soil microbial community structure, soil enzyme activities, and soil C sequestration.     

稻田系统管理措施对微生物生物量C、N、P的长期影响

A 12-year field experiment was conducted to investigate the effect of different tillage methods and fertil-ization systems on microbial biomass C,N and P of a gray fluvo-aguic soil in rice-based cropping system .Five fertilization treatments were designed under conventional tillae(CT) or on tillage(NT) system:no fertilizer(CK) ; chemical fertilizer only(CF) ; combining chemical fertilizer with pig manure(PM); combining chemical fertilizer with crop straw (CS) and fallow (F). The results showed that biomass C,N and P were enriched in the surface layer of no-tilled soil,whereas they distributed relatively evenly in the tilled soil,which might result from enrichment of crop resdue,organic manure and mineral fertilzer,and surficial developent of root systems under NT.Under the cultivation system NT had slightly greater biomass C,N and P at 0-5 cm depth ,significantly less biomass C,N and P at 5-15 cm depth ,less microbial biomass C,N and equivalent biomass P at 15-30 cm depth as compared to CT,indicating hat tillage was beneficial for the multiplication of organims in the plowed layer of soil.Under the fallow system,biomass C,N and P in the surface layer were significantly greater for NT than CT while their differences between the two tillage methods were neligible in the deeper layers.In the surface layer,biomass C,N and P in the soils amended with oranic manure combined with mineral fertilizers were significantly greater than those of the treatments only with mineral fertilizers and the control.Soils without fertilzer had the least biomass nutrient contents among the five fertilization treatments.Obviously,the long-term application of organic manure could maintain the higher activity of microorganisms in soils.The amounts of biomass C,N and P in the fallowed soils varied with the tillage methods;they were much greater under NT than under CT,especially in the surface layer,suggesting that the frequent plowing could decrease the content of organic matter in the surface layer of the fallowed soil.     

不同施肥管理对红壤性水稻土有机碳、氮形态的影响

A long-term experiment beginning in 1981 in Jinxian County of Jiangxi Province, subtropical China, was conducted in a paddy field under a double rice cropping system with four different fertilization regimes, including 1) no fertilizer as control (CK), 2) balanced chemical N, P, and K fertilizers (NPK), 3) organic manure using milk vetch and pig manure in the early and late rice growing season, respectively (OM), and 4) balanced chemical fertilizers combined with organic manure (NPKM). Samples (0-17 cm) of the paddy field soil, which was derived from Quaternary red clay, were collected after the late rice harvest in November 2003 for determination of total organic carbon (TOC) and total nitrogen (TN) and fractions of organic C and N. Results showed that TOC and TN in the NPKM and OM treatments were significantly higher than those in other two treatments (CK and NPK). Application of organic manure with or without chemical fertilizers significantly increased the contents of all fractions of organic C and N, whereas chemical fertilizer application only increased the contents of occluded particulate organic C (oPOC) and amino acid N. In addition, application of organic manure significantly enhanced the proportions of free particulate organic carbon (fPOC) and oPOC in total C, and those of amino sugar N and amino acid N (P < 0.01) in total N. In contrast, chemical fertilizer application only increased the proportions of oPOC and amino acid N (P < 0.05). There were no significant differences in either contents or proportions of soil organic C and organic N fractions between the NPKM and OM treatments. These indicated that organic manure application with or without chemical fertilizers played the most significant role in enhancing soil organic C and N quantity and quality in the paddy field studied.     

Dynamic Relationship Between Biologically Active Soil Organic Carbon and Aggregate Stability in Long-Term Organically Fertilized Soils

Biologically active soil organic carbon (BASOC) is an important fraction of soil organic carbon (SOC), but our understanding of the correlation between BASOC and soil aggregate stability is limited. At an ecological experimental station (28°04 -28°37 N, 116° 41 -117° 09 E) in Yujiang County, Jiangxi Province, China, we analyzed the dynamic relationship between soil aggregate stability and BASOC content over time in the red soil (Udic Ferrosols) fertilized with a nitrogen-phosphorus-potassium chemical fertilizer (NPK) without manure or with NPK plus livestock manure or green manure. The dynamics of BASOC was evaluated using CO2 effux, and soil aggregates were separated according to size using a wet-sieving technique. The soils fertilized with NPK plus livestock manure had a significantly higher content of BASOC and an improved aggregate stability compared to the soils fertilized with NPK plus green manure or NPK alone. The BASOC contents in all fertilized soils decreased over time. The contents of large aggregates (800-2000 μm) dramatically decreased over the first 7 d of incubation, but the contents of small aggregates (< 800 μm) either remained the same or increased, depending on the incubation time and specific aggregate sizes. The aggregate stability did not differ significantly at the beginning and end of incubation, but the lowest stability in all fertilized soils occurred in the middle of the incubation, which implied that the soils had a strong resilience for aggregate stability. The change in BASOC content was only correlated with aggregate stability during the first 27 d of incubation.     

茶树种植对中国东部黄棕壤酸化的影响

Soil acidification is an important process in land degradation around the world as well as in China. Acidification of Alfisols was investigated in the tea gardens with various years of tea cultivation in the eastern China. Cultivation of tea plants caused soil acidification and soil acidity increased with the increase of tea cultivation period. Soil pH of composite samples from cultivated layers decreased by 1.37, 1.62 and 1.85, respectively, after 13, 34 and 54 years of tea plantation, as compared to the surface soil obtained from the unused land. Soil acidification rates at early stages of tea cultivation were found to be higher than those at the later stages. The acidification rate for the period of 0-13 years was as high as 4.40 kmol H⁺ ha^-1 year^-1 for the cultivated layer samples. Soil acidification induced the decrease of soil exchangeable base cations and base cation saturation and thus increased the soil exchangeable acidity. Soil acidification also caused the decrease of soil cation exchange capacity, especially for the 54-year-old tea garden. Soil acidification induced by tea plantation also led to the increase of soil exchangeable Al and soluble Al, which was responsible for the Al toxicity to plants.     

Abundance and community structure of ammonia-oxidizing archaea and bacteria in an acid paddy soil

Nitrification is essential to the nitrogen cycle in paddy soils. However, it is still not clear which group of ammonia-oxidizing microorganisms plays more important roles in nitrification in the paddy soils. The changes in the abundance and composition of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) were investigated by real-time PCR, terminal restriction fragment length polymorphism, and clone library approaches in an acid red paddy soil subjected to long-term fertilization treatments, including treatment without fertilizers (CT); chemical fertilizer nitrogen (N); N and potassium (NK); N and phosphorus (NP); N, P, and K (NPK); and NPK plus recycled crop residues (NPK+C). The AOA population size in NPK+C was higher than those in CT, while minor changes in AOB population sizes were detected among the treatments. There were also some changes in AOA community composition responding to different fertilization treatments. Still few differences were detected in AOB community composition among the treatments. Phylogenetic analysis showed that the AOA sequences fell into two main clusters: cluster A and cluster soil/sediment. The AOB composition in this paddy soil was dominated by Nitrosospira cluster 12. These results suggested that the AOA were more sensitive than AOB to different fertilization treatments in the acid red paddy soil.     

Influence of bacterial-feeding nematodes on nitrification and the ammonia-oxidizing bacteria (AOB) community composition

The effects of bacterial-feeding nematodes on nitrification and the ammonia-oxidizing bacteria (AOB) community composition were studied in soil microcosms. Sterilized soils were inoculated with mixed soil bacteria (obtained by filtering) or with bacteria and bacterial-feeding nematodes, after which the dynamic inorganic nitrogen concentration was measured weekly. After 28 days of incubation, denaturing gradient gel electrophoresis (DGGE) based on PCR amplification of the amoA gene was used to analyze the AOB community composition. In addition, a clone library from the amoA gene fragments was established using clones randomly selected and sequenced from the two treatments. The results showed that the presence of bacterial-feeding nematodes led to significantly greater NH₄⁺ and NO₃⁻ contents over the entire incubation period, indicating that bacterial-feeding nematodes promoted both N mineralization and nitrification. The results of DGGE showed that the AOB community composition was significantly changed in the presence of bacterial-feeding nematodes. Furthermore, the sequencing results suggested that Nitrosospira sp. was the dominant species in the treatment without nematodes, while Nitrosomonas sp. and Nitrosospira sp. were the dominant species in the treatment with nematodes. Such changes in the AOB community may be one of explanation of the important role that nematodes play in promoting nitrification.     

长期施肥对水稻土剖面氨氧化古菌和细菌丰度及组成的影响

长期施肥影响稻田土壤理化性质和硝化微生物群落,但长期施肥对稻田不同土层氨氧化古菌(AOA)和氨氧化细菌(AOB)群落结构的影响尚不明确.以湖南宁乡稻田不同施肥制度长期定位试验为平台,选取不施肥(CK)、施秸秆有机肥(ST)、有机-无机肥配施(OM)和施全量化肥(NPK)4个处理,采用实时荧光定量PCR和Illumina...     

不同施肥模式对土壤氮循环功能微生物的影响

  【目的】   微生物在土壤氮循环过程中发挥着重要作用。通过研究农田土壤氮循环过程中不同功能微生物群落基因丰度对施肥模式的响应及其关键影响因素,探讨不同施肥模式调控下氮素转化的微生物学机制,为改善农业生产中氮素的管理策略提供理论依据。   【方法】   田间试验始于2011年,试验地点位于江苏省常州市溧阳市南渡镇,供试土壤为白土型水稻土,种植制度为稻麦轮作。试验包括单施化肥 (NPK)、化肥+畜禽有机肥 (NPKM)、化肥+秸秆还田 (NPKS) 以及相邻江苏省耕地质量监测点不施肥对照 (CK),共 4个处理。于2014年水稻成熟期采集土壤样品,采用实时荧光定量PCR法分析了土壤硝化 (amoA)、反硝化 (narG、nirS、nirK、norB、nosZ)、固氮 (nifH)、硝酸盐异化还原 (napA) 等氮循环过程的相关功能微生物基因丰度的变化。以氨氧化微生物为模式微生物,测定添加与不添加1-辛炔情况下的土壤硝化潜势,分析氨氧化古菌 (AOA) 与氨氧化细菌 (AOB) 功能基因丰度与土壤硝化功能的内在联系。   【结果】   与CK相比,NPK处理显著增加了土壤中AOB-amoA、narG、nosZ和nifH基因的丰度。与NPK处理相比,NPKS处理进一步提高了土壤中AOB-amoA、narG、nosZ、nifH以及nirK基因的丰度。与CK相比,除AOA-amoA、nirS、napA基因以外,NPKM处理显著提高了土壤中所有氮循环功能基因的丰度。AOB-amoA基因丰度的变化对土壤氮循环功能基因丰度的整体变异影响最大。AOB主导了施肥土壤的硝化过程 (81.90%～84.42%)。土壤总硝化潜势与AOB-amoA基因丰度显著相关,但与AOA-amoA基因丰度相关性不显著。氮循环功能微生物基因丰度主要受到土壤pH、土壤有机碳(SOC)和NO₃–含量的影响。   【结论】   畜禽有机肥与秸秆的施用能够进一步刺激氮循环功能基因丰度的增长,促进土壤氮循环。土壤pH、SOC和NO₃–含量是影响土壤氮循环功能微生物丰度的关键因素。施肥主要通过提高土壤AOB-amoA功能基因的丰度,进而提高土壤硝化潜势,因此在控制土壤硝化作用时应重点关注AOB微生物群落。     

Diversity–function relationship of ammonia-oxidizing bacteria in soils among functional groups of grassland species under climate warming

Although warming and plant diversity losses have important effects on aboveground ecosystem functioning, their belowground effects remain largely unknown. We studied the impact of a 3 °C warming and of three plant functional groups (forbs, grasses, legumes) on ammonia-oxidizing bacteria (AOB) diversity (polymerase chain reaction-denaturing gradient gel electrophoresis, PCR-DGGE) and their function (potential nitrification) in artificial grasslands. Warming did not influence AOB diversity and function. Sequencing of 16S rRNA gene fragments retrieved from DGGE gel revealed that they were all related to Nitrosospira-like sequences. Clustering analysis of DGGE profiles resulted in two nodes, separating AOB community structure under legumes from all other samples. Decreased AOB richness (number of DGGE bands) and concurrent increased potential nitrification were also observed under legumes. We hypothesized that ammonium availability was the driving force regulating the link between aboveground and belowground communities, as well as the AOB diversity and function link. The results document that the physiology of AOB might be an important regulator of AOB community structure and function under plant functional groups. This study highlights the major role of the microbial community composition in soil process responses to changes in the functional composition of plant communities.     

Net nitrification rate and presence of Nitrosospira cluster 2 in acid coniferous forest soils appear to be tree species specific

The impact of four coniferous tree species and their corresponding soil factors on N transformation rates and presence of ammonia-oxidising bacteria (AOB) was studied in an acid pine forest soil (Appelscha, The Netherlands). Pine soil had a relatively low net nitrification rate, while spruce, fir and larch soils showed high net nitrification rates. 16S rRNA and amoA sequences were only found in soils with high nitrification rates and belonged solely to Nitrosospira cluster 2. We conclude that tree species, possibly through their effects on soil C/N ratios, determines the presence of Nitrosospira cluster 2. Whenever AOB are present, however, the AOB community composition appears to be similar.     

Different roles of rhizosphere effect and long-term fertilization in the activity and community structure of ammonia oxidizers in a calcareous fluvo-aquic soil

Ammonia oxidation is a critical step in the soil nitrogen (N) cycle and can be affected by the application of mineral fertilizers or organic manure. However, little is known about the rhizosphere effect on the function and structure of ammonia-oxidizing bacterial (AOB) and archaeal (AOA) communities, the most important organisms responsible for ammonia oxidation in agricultural ecosystems. Here, the potential nitrification activity (PNA), population size and composition of AOB and AOA communities in both the rhizosphere and bulk soil from a long-term (31-year) fertilizer field experiment conducted during two seasons (wheat and maize) were investigated using the shaken slurry method, quantitative real-time polymerase chain reaction and denaturing gradient gel electrophoresis. N fertilization greatly enhanced PNA and AOB abundance, while manure application increased AOA abundance. The community structure of AOB exhibited more obvious shifts than that of AOA after long-term fertilization, resulting in more abundant AOB phylotypes similar to Nitrosospira clusters 3 and 4 in the N-fertilized treatments. Moreover, PNA was closely correlated with the abundance and community structure of AOB rather than that of AOA among soils during both seasons, indicating that AOB play an active role in ammonia oxidation. Conversely, the PNA and population sizes of AOB and AOA were typically higher in the rhizosphere than the bulk soil, implying a significant rhizosphere effect on ammonia oxidation. Cluster and redundancy analyses further showed that this rhizosphere effect played a more important role in shaping AOA community structure than long-term fertilization. Overall, the results indicate that AOB rather than AOA functionally dominate ammonia oxidation in the calcareous fluvo-aquic soil, and that rhizosphere effect and fertilization regime play different roles in the activity and community structures of AOB and AOA.     

Liming induces growth of a diverse flora of ammonia-oxidising bacteria in acid spruce forest soil as determined by SSCP and DGGE

The autotrophic ammonia-oxidising bacterial (AOB) community composition was studied in acid coniferous forest soil profiles at a site in southwestern Sweden 6 years after liming. Liming caused a significant increase in pH in the organic horizons, while the mineral soil was unaffected. The AOB communities were studied by single-strand conformation polymorphism (SSCP) in parallel with denaturing gradient gel electrophoresis (DGGE) analysis of partial 16S rRNA genes amplified by PCR using primers reported to be specific for β-Proteobacteria AOB, followed by nucleotide sequencing. High genetic diversity of Nitrosospira-like sequences was found in the limed soil profiles, whereas no AOB-like sequences were detected in the control soil at any depth, according to both the SSCP and DGGE analyses. This clearly showed that liming induced growth of a diverse flora of AOB at this forest site. Both Nitrosospira cluster 2 and cluster 4 sequences were present in the limed soil profiles, regardless of soil pH, but we found a higher number of sequences affiliated with cluster 4. The high lime dose seemed to affect the AOB community more than the low dose, and its effects reached deeper into the soil profile. Seven different Nitrosospira-like sequences were found 10 cm under the litter layer in the soil limed with the high dose, but only two in the soil amended with the low lime dose.     

Long-term fertilization effects on active ammonia oxidizers in an acidic upland soil in China

The effects of long-term fertilization of acidic soils on ammonia-oxidizing archaea (AOA) and bacteria (AOB) communities and its ecological implications remain poorly understood. We chose an acidic upland soil site under long-term (27-year) fertilization to investigate ammonia oxidizer communities under four different regimes: mineral N fertilizer (N), mineral NPK fertilizer (NPK), organic manure (OM) and an unfertilized control (CK). Soil net nitrification rates were significantly higher in OM soils than in CK, N or NPK soils. Quantitative analysis of the distribution of amoA genes by DNA-based stable isotope probing revealed that AOA dominate in CK, N and NPK soils, while AOB dominate in OM soils. Denaturing gradient gel electrophoresis and clone library analyses of amoA genes revealed that Group 1.1a-associated AOA (also referred to as Nitrosotalea) were the most dominant active AOA population (>92%), while Nitrosospira Cluster 3 and Cluster 9 were predominant among active AOB communities. The functional diversity of active ammonia oxidizers in acidic soils is affected by long-term fertilization practices, and the responses of active ammonia oxidizers to mineral fertilizer and organic manure are clearly different. Our results provide strong evidence that AOA are more highly adapted to growth at low pH and low substrate availability than AOB, and they suggest that the niche differentiation and metabolic diversity of ammonia oxidizers in acidic soils are more complex than previously thought.