Nitrification of archaeal ammonia oxidizers in acid soils is supported by hydrolysis of urea

The hydrolysis of urea as a source of ammonia has been proposed as a mechanism for the nitrification of ammonia-oxidizing bacteria (AOB) in acidic soil. The growth of Nitrososphaera viennensis on urea suggests that the ureolysis of ammonia-oxidizing archaea (AOA) might occur in natural environments. In this study, ¹⁵N isotope tracing indicates that ammonia oxidation occurred upon the addition of urea at a concentration similar to the in situ ammonium content of tea orchard soil (pH 3.75) and forest soil (pH 5.4) and was inhibited by acetylene. Nitrification activity was significantly stimulated by urea fertilization and coupled well with abundance changes in archaeal amoA genes in acidic soils. Pyrosequencing of 16S rRNA genes at whole microbial community level demonstrates the active growth of AOA in urea-amended soils. Molecular fingerprinting further shows that changes in denaturing gradient gel electrophoresis fingerprint patterns of archaeal amoA genes are paralleled by nitrification activity changes. However, bacterial amoA and 16S rRNA genes of AOB were not detected. The results strongly suggest that archaeal ammonia oxidation is supported by hydrolysis of urea and that AOA, from the marine Group 1.1a-associated lineage, dominate nitrification in two acidic soils tested.     

Ammonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidic soils

Increasing evidence demonstrated the involvement of ammonia-oxidizing archaea (AOA) in the global nitrogen cycle, but the relative contributions of AOA and ammonia-oxidizing bacteria (AOB) to ammonia oxidation are still in debate. Previous studies suggest that AOA would be more adapted to ammonia-limited oligotrophic conditions, which seems to be favored by protonation of ammonia, turning into ammonium in low-pH environments. Here, we investigated the autotrophic nitrification activity of AOA and AOB in five strongly acidic soils (pH<4.50) during microcosm incubation for 30 days. Significantly positive correlations between nitrate concentration and amoA gene abundance of AOA, but not of AOB, were observed during the active nitrification. ¹³CO₂-DNA-stable isotope probing results showed significant assimilation of ¹³C-labeled carbon source into the amoA gene of AOA, but not of AOB, in one of the selected soil samples. High levels of thaumarchaeal amoA gene abundance were observed during the active nitrification, coupled with increasing intensity of two denaturing gradient gel electrophoresis bands for specific thaumarchaeal community. Addition of the nitrification inhibitor dicyandiamide (DCD) completely inhibited the nitrification activity and CO₂ fixation by AOA, accompanied by decreasing thaumarchaeal amoA gene abundance. Bacterial amoA gene abundance decreased in all microcosms irrespective of DCD addition, and mostly showed no correlation with nitrate concentrations. Phylogenetic analysis of thaumarchaeal amoA gene and 16S rRNA gene revealed active ¹³CO₂-labeled AOA belonged to groups 1.1a-associated and 1.1b. Taken together, these results provided strong evidence that AOA have a more important role than AOB in autotrophic ammonia oxidation in strongly acidic soils.     

Autotrophic growth of nitrifying community in an agricultural soil

The two-step nitrification process is an integral part of the global nitrogen cycle, and it is accomplished by distinctly different nitrifiers. By combining DNA-based stable isotope probing (SIP) and high-throughput pyrosequencing, we present the molecular evidence for autotrophic growth of ammonia-oxidizing bacteria (AOB), ammonia-oxidizing archaea (AOA) and nitrite-oxidizing bacteria (NOB) in agricultural soil upon ammonium fertilization. Time-course incubation of SIP microcosms indicated that the amoA genes of AOB was increasingly labeled by ¹³CO₂ after incubation for 3, 7 and 28 days during active nitrification, whereas labeling of the AOA amoA gene was detected to a much lesser extent only after a 28-day incubation. Phylogenetic analysis of the ¹³C-labeled amoA and 16S rRNA genes revealed that the Nitrosospira cluster 3-like sequences dominate the active AOB community and that active AOA is affiliated with the moderately thermophilic Nitrososphaera gargensis from a hot spring. The higher relative frequency of Nitrospira-like NOB in the ¹³C-labeled DNA suggests that it may be more actively involved in nitrite oxidation than Nitrobacter-like NOB. Furthermore, the acetylene inhibition technique showed that ¹³CO₂ assimilation by AOB, AOA and NOB occurs only when ammonia oxidation is not blocked, which provides strong hints for the chemolithoautotrophy of nitrifying community in complex soil environments. These results show that the microbial community of AOB and NOB dominates the nitrification process in the agricultural soil tested.     

Active Ammonia Oxidizers in an Acidic Soil Are Phylogenetically Closely Related to Neutrophilic Archaeon

All cultivated ammonia-oxidizing archaea (AOA) within the Nitrososphaera cluster (former soil group 1.1b) are neutrophilic. Molecular surveys also indicate the existence of Nitrososphaera-like phylotypes in acidic soil, but their ecological roles are poorly understood. In this study, we present molecular evidence for the chemolithoautotrophic growth of Nitrososphaera-like AOA in an acidic soil with pH 4.92 using DNA-based stable isotope probing (SIP). Soil microcosm incubations demonstrated that nitrification was stimulated by urea fertilization and accompanied by a significant increase in the abundance of AOA rather than ammonia-oxidizing bacteria (AOB). Real-time PCR analysis of amoA genes as a function of the buoyant density of the DNA gradient following the ultracentrifugation of the total DNA extracted from SIP microcosms indicated a substantial growth of soil AOA during nitrification. Pyrosequencing of the total 16S rRNA genes in the “heavy” DNA fractions suggested that archaeal communities were labeled to a much greater extent than soil AOB. Acetylene inhibition further showed that ¹³CO₂ assimilation by nitrifying communities depended solely on ammonia oxidation activity, suggesting a chemolithoautotrophic lifestyle. Phylogenetic analysis of both ¹³C-labeled amoA and 16S rRNA genes revealed that most of the active AOA were phylogenetically closely related to the neutrophilic strains Nitrososphaera viennensis EN76 and JG1 within the Nitrososphaera cluster. Our results provide strong evidence for the adaptive growth of Nitrososphaera-like AOA in acidic soil, suggesting a greater metabolic versatility of soil AOA than previously appreciated.     

Differential contributions of ammonia oxidizers and nitrite oxidizers to nitrification in four paddy soils

Rice paddy fields are characterized by regular flooding and nitrogen fertilization, but the functional importance of aerobic ammonia oxidizers and nitrite oxidizers under unique agricultural management is poorly understood. In this study, we report the differential contributions of ammonia-oxidizing archaea (AOA), bacteria (AOB) and nitrite-oxidizing bacteria (NOB) to nitrification in four paddy soils from different geographic regions (Zi-Yang (ZY), Jiang-Du (JD), Lei-Zhou (LZ) and Jia-Xing (JX)) that are representative of the rice ecosystems in China. In urea-amended microcosms, nitrification activity varied greatly with 11.9, 9.46, 3.03 and 1.43 μg NO₃⁻-N g⁻¹ dry weight of soil per day in the ZY, JD, LZ and JX soils, respectively, over the course of a 56-day incubation period. Real-time quantitative PCR of amoA genes and pyrosequencing of 16S rRNA genes revealed significant increases in the AOA population to various extents, suggesting that their relative contributions to ammonia oxidation activity decreased from ZY to JD to LZ. The opposite trend was observed for AOB, and the JX soil stimulated only the AOB populations. DNA-based stable-isotope probing further demonstrated that active AOA numerically outcompeted their bacterial counterparts by 37.0-, 10.5- and 1.91-fold in ¹³C-DNA from ZY, JD and LZ soils, respectively, whereas AOB, but not AOA, were labeled in the JX soil during active nitrification. NOB were labeled to a much greater extent than AOA and AOB, and the addition of acetylene completely abolished the assimilation of ¹³CO₂ by nitrifying populations. Phylogenetic analysis suggested that archaeal ammonia oxidation was predominantly catalyzed by soil fosmid 29i4-related AOA within the soil group 1.1b lineage. Nitrosospira cluster 3-like AOB performed most bacterial ammonia oxidation in the ZY, LZ and JX soils, whereas the majority of the ¹³C-AOB in the JD soil was affiliated with the Nitrosomona communis lineage. The ¹³C-NOB was overwhelmingly dominated by Nitrospira rather than Nitrobacter. A significant correlation was observed between the active AOA/AOB ratio and the soil oxidation capacity, implying a greater advantage of AOA over AOB under microaerophilic conditions. These results suggest the important roles of soil physiochemical properties in determining the activities of ammonia oxidizers and nitrite oxidizers.     

Cultivation of a highly enriched ammonia-oxidizing archaeon of thaumarchaeotal group I.1b from an agricultural soil

Nitrification of excess ammonia in soil causes eutrophication of water resources and emission of atmospheric N₂O gas. The first step of nitrification, ammonia oxidation, is mediated by Archaea as well as Bacteria. The physiological reactions mediated by ammonia‐oxidizing archaea (AOA) and their contribution to soil nitrification are still unclear. Results of non‐culture‐based studies have shown the thaumarchaeotal group I.1b lineage of AOA to be dominant over both AOA of group I.1a and ammonia‐oxidizing bacteria in various soils. We obtained from an agricultural soil a highly enriched ammonia‐oxidizing culture dominated by a single archaeal population [c. 90% of total cells, as determined microscopically (by fluorescence in situ hybridization) and by quantitative PCR of its 16S rRNA gene]. The archaeon (termed ‘strain JG1’) fell within thaumarchaeotal group I.1b and was related to the moderately thermophilic archaeon, Candidatus Nitrososphaera gargensis, and the mesophilic archaeon, Ca. Nitrososphaera viennensis with 97.0% and 99.1% 16S rRNA gene sequence similarity respectively. Strain JG1 was neutrophilic (growth range pH 6.0–8.0) and mesophilic (growth range temperature 25–40°C). The optimum temperature of strain JG1 (35–40°C) is > 10°C higher than that of ammonia‐oxidizing bacteria (AOB). Membrane analysis showed that strain JG1 contained a glycerol dialkyl glycerol tetraether, GDGT‐4, and its regioisomer as major core lipids; this crenarchaeol regioisomer was previously detected in similar abundance in the thermophile, Ca. N. gargensis and has been frequently observed in tropical soils. Substrate uptake assays showed that the affinity of strain JG1 for ammonia and oxygen was much higher than those of AOB. These traits may give a competitive advantage to AOA related to strain JG1 in oligotrophic environments. ¹³C‐bicarbonate incorporation into archaeal lipids of strain JG1 established its ability to grow autotrophically. Strain JG1 produced a significant amount of N₂O gas – implicating AOA as a possible source of N₂O emission from soils. Sequences of archaeal amoA and 16S rRNA genes closely related to those of strain JG1 have been retrieved from various terrestrial environments in which lineage of strain JG1 is likely engaged in autotrophic nitrification.     

Quantitative analyses of ammonia-oxidizing Archaea and bacteria in the sediments of four nitrogen-rich wetlands in China

With the rapid development of ammonia-synthesizing industry, the ammonia-nitrogen pollution in wetlands acting as the sink of point and diffuse pollution has been increased dramatically. Most of ammonia-nitrogen is oxidized at least once by ammonia-oxidizing prokaryotes to complete the nitrogen cycle. Current research findings have expanded the known ammonia-oxidizing prokaryotes from the domain Bacteria to Archaea. However, in the complex wetlands environment, it remains unclear whether ammonia oxidation is exclusively or predominantly linked to Archaea or Bacteria as implied by specific high abundance. In this research, the abundance and composition of Archaea and Bacteria in sediments of four kinds of wetlands with different nitrogen concentration were investigated by using quantitative real-time polymerase chain reaction, cloning, and sequencing approaches based on amoA genes. The results indicated that AOA distributed widely in wetland sediments, and the phylogenetic tree revealed that archaeal amoA functional gene sequences from wetlands sediments cluster as two major evolutionary branches: soil/sediment and sediment/water. The bacteria functionally dominated microbial ammonia oxidation in different wetlands sediments on the basis of molecule analysis, potential nitrification rate, and soil chemistry. Moreover, the factors influencing AOA and AOB abundances with environmental indicator were also analyzed, and the results addressed the copy numbers of archaeal and bacterial amoA functional gene having the higher correlation with pH and ammonia concentration. The pH had relatively great negative impact on the abundance of AOA and AOB, while ammonia concentration showed positive impact on AOB abundance only. These findings could be fundamental to improve understanding of the importance of AOB and AOA in nitrogen and other nutrients cycle in wetland ecosystems.     

RETRACTED ARTICLE: Archaeal <Emphasis Type="Italic">amoA</Emphasis> Genes Outnumber Bacterial <Emphasis Type="Italic">amoA</Emphasis> Genes in Municipal Wastewater Treatment Plants in Bangkok

The contribution of ammonia-oxidizing archaea (AOA) to nitrogen removal in wastewater treatment plants (WWTPs) remains unknown. This study investigated the abundance of archaeal (AOA) and bacterial (ammonia-oxidizing bacteria (AOB)) amoA genes in eight of Bangkok’s municipal WWTPs. AOA amoA genes (3.28 × 10⁷ ± 1.74 × 10⁷–2.23 × 10¹¹ ± 1.92 × 10¹¹ copies l⁻¹ sludge) outnumbered AOB amoA genes in most of the WWTPs even though the plants’ treatment processes, influent and effluent characteristics, removal efficiencies, and operation varied. An estimation of the ammonia-oxidizing activity of AOA and AOB suggests that AOA involved in autotrophic ammonia oxidation in the WWTPs. Statistical analysis shows that the numbers of AOA amoA genes correlated negatively to the ammonium levels in effluent wastewater, while no correlation was found between the AOA amoA gene numbers and the oxygen concentrations in aeration tanks. An analysis of the AOB sequences shows that AOB found in the WWTPs limited to only two AOB clusters which exhibit high or moderate affinity to ammonia. In contrast to AOB, AOA sequences of various clusters were retrieved, and they were previously recovered from a variety of environments, such as thermal and marine environments.     

Do ammonia-oxidizing archaea respond to soil Cu contamination similarly asammonia-oxidizing bacteria?

Inhibitory experiments were conducted to investigate the responses of the population sizes of ammonia-oxidizing archaea (AOA) and bacteria (AOB) and the potential nitrification rates (PNRs) to Cu contamination in four Chinese soils. PNR was determined using a substrate-induced nitrification (SIN) assay, and the population size of the nitrifiers represented by amoA gene abundances was quantified using a real-time polymerase chain reaction (qPCR) assay. Both population size and PNR of the ammonia oxidizers reduced considerably at high Cu concentrations in all the soils. Bacterial amoA gene abundance was reduced by from 107-fold (Hailun soil) to more than 232-fold (Hangzhou soil) at the highest Cu concentrations (2,400 mg kg^?1 Cu for Hailun, Langfang and Guangzhou soils and 1,600 mg kg^?1 Cu for Hangzhou soil), while reduction in archaeal amoA gene abundance was from 10-fold (Langfang soil) to 89-fold (Hangzhou soil). AOA seemed more tolerant to Cu contamination than AOB. Nitrification rates were inhibited by more than 50% at a Cu concentration of 600 mg kg^?1, and by more than 90% at the highest Cu concentrations in all soils. These results indicated that both AOA and AOB can be inhibited by toxic metals, highlighting the need to consider the role of AOA in nitrification in soils.     

Population ecology of nitrifying Archaea and Bacteria in the Southern California Bight

Marine Crenarchaeota are among the most abundant microbial groups in the ocean, and although relatively little is currently known about their biogeochemical roles in marine ecosystems, recognition that Crenarchaeota posses ammonia monooxygenase (amoA) genes and may act as ammonia‐oxidizing archaea (AOA) offers another means of probing the ecology of these microorganisms. Here we use a time series approach combining quantification of archaeal and bacterial ammonia oxidizers with bacterial community fingerprints and biogeochemistry, to explore the population and community ecology of nitrification. At multiple depths (150, 500 and 890 m) in the Southern California Bight sampled monthly from 2003 to 2006, AOA were enumerated via quantitative PCR of archaeal amoA and marine group 1 Crenarchaeota 16S rRNA genes. Based on amoA genes, AOA were highly variable in time – a consistent feature of marine Crenarchaeota– however, average values were similar at different depths and ranged from 2.20 to 2.76 × 10⁴amoA copies ml^?1. Archaeal amoA genes were correlated with Crenarchaeota 16S rRNA genes (r² = 0.79) and the slope of this relationship was 1.02, demonstrating that the majority of marine group 1 Crenarchaeota present over the dates and depths sampled possessed amoA. Two AOA clades were specifically quantified and compared with betaproteobacterial ammonia‐oxidizing bacteria (β‐AOB) amoA genes at 150 m; these AOA groups were found to strongly co‐vary in time (r² = 0.70, P < 0.001) whereas AOA : β‐AOB ratios ranged from 13 to 5630. Increases in the AOA : β‐AOB ratio correlated with the accumulation of nitrite (r² = 0.87, P < 0.001), and may be indicative of differences in substrate affinities and activities leading to periodic decoupling between ammonia and nitrite oxidation. These data capture a dynamic nitrogen cycle in which multiple microbial groups appear to be active participants.     

Simazine application inhibits nitrification and changes the ammonia-oxidizing bacterial communities in a fertilized agricultural soil

s-Triazine herbicides are widely used for weed control, and are persistent in soils. Nitrification is an essential process in the global nitrogen cycle in soil, and involves ammonia-oxidizing Bacteria (AOB) and ammonia-oxidizing Archaea (AOA). In this study, we evaluated the effect of the s-triazine herbicide simazine on the nitrification and on the structure of ammonia-oxidizing microbial communities in a fertilized agricultural soil. The effect of simazine on AOB and AOA were studied by PCR-amplification of amoA genes of nitrifying Bacteria and Archaea in soil microcosms and denaturing gradient gel electrophoresis (DGGE) analyses. Simazine [50?μg g(-1) dry weight soil (d.w.s)] completely inhibited the nitrification processes in the fertilized agricultural soil. The inhibition by simazine of ammonia oxidation observed was similar to the reduction of ammonia oxidation by the nitrification inhibitor acetylene. The application of simazine-affected AOB community DGGE patterns in the agricultural soil amended with ammonium, whereas no significant changes in the AOA community were observed. The DGGE analyses strongly suggest that simazine inhibited Nitrosobacteria and specifically Nitrosospira species. In conclusion, our results suggest that the s-triazine herbicide not only inhibits the target susceptible plants but also inhibits the ammonia oxidation and the AOB in fertilized soils.     

Nanosilver inhibits nitrification and reduces ammonia‐oxidising bacterial but not archaeal amoA gene abundance in estuarine sediments

Silver nanoparticles (AgNPs) enter estuaries via wastewater treatment effluents, where they can inhibit microorganisms, because of their antimicrobial properties. Ammonia‐oxidising bacteria (AOB) and archaea (AOA) are involved in the first step of nitrification and are important to ecosystem function, especially where effluent discharge results in high nitrogen inputs. Here, we investigated the effect of a pulse addition of AgNPs on AOB and AOA ammonia monooxygenase (amoA) gene abundances and benthic nitrification potential rates (NPR) in low‐salinity and mesohaline estuarine sediments. Whilst exposure to 0.5 mg L^?1 AgNPs had no significant effect on amoA gene abundances or NPR, 50 mg L^?1 AgNPs significantly decreased AOB amoA gene abundance (up to 76% over 14 days), and significantly decreased NPR by 20‐fold in low‐salinity sediments and by twofold in mesohaline sediments, after one day. AgNP behaviour differed between sites, whereby greater aggregation occurred in mesohaline waters (possibly due to higher salinity), which may have reduced toxicity. In conclusion, AgNPs have the potential to reduce ammonia oxidation in estuarine sediments, particularly where AgNPs accumulate over time and reach high concentrations. This could lead to long‐term risks to nitrification, especially in polyhaline estuaries where ammonia‐oxidation is largely driven by AOB.     

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...     

重庆紫色水稻土中“全程”和“半程”氨氧化微生物的垂直分异

【目的】系统评估全程氨氧化细菌(complete ammonia oxidizing bacteria, Comammox bacteria)、半程氨氧化细菌(AOB)和古菌(AOA)在典型水稻土剖面的垂直分异规律。2015年发现的"全程"氨氧化细菌(Comammox Nitrospira)可将氨分子一步氧化为硝酸盐,实现硝化作用。而经典的"半程"氨氧化细菌(AOB)或古菌(AOA)将氨分子氧化为亚硝酸盐后,再由系统发育完全不同的硝化细菌将其氧化为硝酸盐。全程氨氧化细菌实现了一步硝化全过程,根本改变了学术界对2类微生物分步硝化的经典认知,但相关研究仍处于初步阶段。【方法】选择重庆北碚地区2017年典型水稻土并采集5、10、20和40 cm不同深度土壤(剖面采样点的上下误差不超过1cm),提取水稻土总DNA后,利用标靶功能基因amoA,通过实时荧光定量PCR技术分析全程氨氧化细菌(Comammox)、半程氨氧化细菌(AOB)和古菌(AOA)在水稻土不同深度的数量变异规律。【结果】半程氨氧化细菌AOB和古菌AOA均随土壤深度增加呈显著下降趋势。然而,全程氨氧化细菌的两大类微生物则表现出相反的规律,Comammox Clade A的丰度随着土壤剖面的加深而显著增加(P0.05),但Clade B并未有类似规律。Clade A在水稻土不同层次的土层中均比Clade B高出1个数量级,在5 cm和40 cm处的最低和最高值分别为3.42×10~7、8.46×10~7 copies/g。AOA与AOB的丰度大致相当,5cm剖面处数量最高分别为1.23×10~7、1.83×10~5copies/g,但其平均丰度远低于全程氨氧化细菌,Comammox与AOA、AOB amoA功能基因拷贝数之比为10–2000。【结论】全程氨氧化细菌(Comammox bacteria)广泛分布于水稻土不同土层中,且数量远高于"半程"氨氧化细菌和古菌,意味着Comammox可能在水稻土硝化作用中起重要作用。     

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.     

Aggregate Size Distribution of Ammonia-Oxidizing Bacteria and Archaea at Different Landscape Positions

To quantify the spatial distribution of ammonia-oxidizing bacteria (AOB) and archaea (AOA) and to determine nitrification activity in soil aggregates along a landscape, soil samples were collected from three landscape positions (shoulder, backslope, and toeslope) at two pasture sites with contrasting climatic conditions. The abundance of AOB and AOA was estimated by quantifying their respective bacterial and archaeal amoA gene copies using real-time polymerase chain reaction. Soil organic C (SOC), total N (TN), and the potential nitrification rate (PNR) were measured in aggregate size ranges (4–1, 1–0.25, and 0.25–0.05 mm). At site 1, a decreasing trend in PNR was observed as the size of aggregates decreased. Both bacterial and archaeal amoA genes were higher in the macroaggregates (4–1 and 1–0.25 mm) than in the microaggregates (0.25–0.05 mm) along the landscape. At site 2, PNR was higher in the smallest size of aggregates. In the 0.25–0.05-mm fraction, the abundance of bacterial and archaeal amoA genes was equal to, or greater than, those found in larger aggregate sizes. The relative abundance of archaeal amoA gene and the PNR correlated with relative SOC and TN contents along the landscapes. The positive relationship between relative archaeal amoA gene abundance and PNR suggests that nitrification in the studied pastures is probably driven by ammonia-oxidizing Thaumarchaeota.     

Abundance and Diversity of Ammonia-Oxidizing Prokaryotes in the Root–Rhizosphere Complex of Miscanthus × giganteus Grown in Heavy Metal-Contaminated Soils

Mine wastes have been considered as a source of heavy metal (HM) contamination in the environment and negatively impact many important ecosystem services provided by soils. Plants like Miscanthus, which tolerate high HM concentrations in soil, are often used for phytoremediation and provide the possibility to use these soils at least for the production of energy crops. However, it is not clear if plant growth at these sites is limited by the availability of nutrients, mainly nitrogen, as microbes in soil might be affected by the contaminant. Therefore, in this study, we investigated in a greenhouse experiment the response of ammonia-oxidizing microbes in the root–rhizosphere complex of Miscanthus × giganteus grown in soils with different levels of long-term arsenic (As) and lead (Pb) contamination. Quantitative PCR of the ammonia monooxigenease gene (amoA) was performed to assess the abundance of ammonia-oxidizing bacteria (AOB) and archaea (AOA) at two different points of plant growth. Furthermore, bulk soil samples before planting were analyzed. In addition, terminal restriction fragment length polymorphism (T-RFLP) analysis was used to investigate the diversity of archaeal amoA amplicons. Whereas high concentrations of As and Pb in soil (83 and 15?g/kg, respectively) resulted independent from plant growth in a clear reduction of AOA and AOB compared to the control soils with lower HM contents, in soils with contamination levels of 10?g/kg As and 0.2?g/kg Pb, only AOB were negatively affected in bulk soil samples. Diversity analysis of archaeal amoA genes revealed clear differences in T-RFLP patterns in response to the degree of HM contamination. Therefore, our results could clearly prove the different response patterns of AOA and AOB in HM-contaminated soils and the development of archaeal amoA phylotypes which are more tolerant towards HMs in soil samples from the areas that were impacted the most by mining waste, which could contribute to functional redundancy of ammonia-oxidizing microbes in soils and stability of nitrification pattern.