Contributions of ammonia-oxidizing archaea and bacteria to nitrification in Oregon forest soils

Ammonia oxidation, the first step of nitrification, is mediated by both ammonia-oxidizing archaea (AOA) and bacteria (AOB); however, the relative contributions of AOA and AOB to soil nitrification are not well understood. In this study we used 1-octyne to discriminate between AOA- and AOB-supported nitrification determined both in soil-water slurries and in unsaturated whole soil at field moisture. Soils were collected from stands of red alder (Alnus rubra Bong.) and Douglas-fir (Pseudotsuga menziesii Mirb. Franco) at three sites (Cascade Head, the H.J. Andrews, and McDonald Forest) on acidic soils (pH 3.9–5.7) in Oregon, USA. The abundances of AOA and AOB were measured using quantitative PCR by targeting the amoA gene, which encodes subunit A of ammonia monooxygenase. Total and AOA-specific (octyne-resistant) nitrification activities in soil slurries were significantly higher at Cascade Head (the most acidic soils, pH < 5) than at either the H.J. Andrews or McDonald Forest, and greater in red alder compared with Douglas-fir soils. The fraction of octyne-resistant nitrification varied among sites (21–74%) and was highest at Cascade Head than at the other two locations. Net nitrification rates of whole soil without NH₄⁺ amendment ranged from 0.4 to 3.3 mg N kg⁻¹ soil d⁻¹. Overall, net nitrification rates of whole soil were stimulated 2- to 8-fold by addition of 140 mg NH₄⁺-N kg⁻¹ soil; this was significant for red alder at Cascade Head and the H.J. Andrews. Red alder at Cascade Head was unique in that the majority of NH₄⁺-stimulated nitrifying activity was octyne-resistant (73%). At all other sites, NH₄⁺-stimulated nitrification was octyne-sensitive (68–90%). The octyne-sensitive activity—presumably AOB—was affected more by soil pH whereas the octyne-resistant (AOA) activity was more strongly related to N availability.     

Conversion of grassland to coniferous woodland has limited effects on soil nitrogen cycle processes

In the last century, conversion of native North American grasslands to Juniperus virginiana forests or woodlands has dramatically altered ecosystem structure and significantly increased ecosystem carbon (C) stocks. We compared soils under recently established J. virginiana forests and adjacent native C₄-dominated grassland to assess changes in potential soil nitrogen (N) transformations and plant available N. Over a 2-year period, concentrations of extractable inorganic N were measured in soils from forest and grassland sites. Potential gross N ammonification, nitrification, and consumption rates were determined using ¹⁵N isotope-dilution under laboratory conditions, controlling for soil temperature and moisture content. Potential nitrification rates (V_max) and microbial biomass, as well as soil physical and chemical properties were also assessed. Extractable NH₄⁺ concentrations were significantly greater in grassland soils across the study period (P ≤ 0.01), but analysis by date indicated that differences in extractable inorganic N occurred more frequently in fall and winter, when grasses were senescent but J. virginiana was still active. Laboratory-based rates of gross N mineralization (ammonification) and nitrification were greater in grassland soils (P ≤ 0.05), but only on one of four dates. Potential nitrification rates (V_max) were an order of magnitude greater than gross nitrification rates in both ecosystems, suggesting that nitrification is highly constrained by NH₄⁺ availability. Differences in plant uptake of N, C inputs, and soil microclimate as forests replace grasslands may influence plant available N in the field, as evidenced by seasonal differences in soil extractable NH₄⁺, and total soil C and N accumulation. However, we found few differences in potential soil N transformations under laboratory conditions, suggesting that this grassland-to-forest conversion caused little change in mineralizable organic N pools or potential microbial activity.     

Microbial immobilization of ammonium and nitrate in relation to ammonification and nitrification rates in organic and conventional cropping systems

Agricultural systems that receive high or low organic matter (OM) inputs would be expected to differ in soil nitrogen (N) transformation rates and fates of ammonium (NH₄⁺) and nitrate (NO₃⁻). To compare NH₄⁺ availability, competition between nitrifiers and heterotrophic microorganisms for NH₄⁺, and microbial NO₃⁻ assimilation in an organic vs. a conventional irrigated cropping system in the California Central Valley, chemical and biological soil assays, ¹⁵N isotope pool dilution and ¹⁵N tracer techniques were used. Potentially mineralizable N (PMN) and hot minus cold KCl-extracted NH₄⁺ as indicators of soil N supplying capacity were measured five times during the tomato growing season. At mid-season, rates of gross ammonification and gross nitrification after rewetting dry soil were measured in microcosms. Microbial immobilization of NO₃⁻ and NH₄⁺ was estimated based on the uptake of ¹⁵N and gross consumption rates. Gross ammonification, PMN, and hot minus cold KCl-extracted NH₄⁺ were approximately twice as high in the organically than the conventionally managed soil. Net estimated microbial NO₃⁻ assimilation rates were between 32 and 35% of gross nitrification rates in the conventional and between 37 and 46% in the organic system. In both soils, microbes assimilated more NO₃⁻ than NH₄⁺. Heterotrophic microbes assimilated less NH₄⁺ than NO₃⁻ probably because NH₄⁺ concentrations were low and competition by nitrifiers was apparently strong. The high OM input organic system released NH₄⁺ in a gradual manner and, compared to the low OM input conventional system, supported a more active microbial biomass with greater N demand that was met mainly by NO₃⁻ immobilization.     

A method to assess whether ‘preferential use’ occurs after N ammonium addition; implication for the N isotope dilution technique

The robustness of the assumption of equilibrium between native and added N during ¹⁵N isotope dilution has recently been questioned by Watson et al. (Soil Biol Biochem 32 (2000) 2019-2030). We re-analyzed their raw data using equations that consider the added and native NH₄⁺ and NO₃⁻ pools as separate state variables. Gross mineralization rates and first-order rate constants for NH₄⁺ and NO₃⁻ consumption were obtained by combining analytical integration of the differential equations with a non-linear fitting procedure. The first-order rate constants for NH₄⁺ consumption and NO₃⁻ immobilization for the added NH₄⁺ and NO₃⁻ pool were used to estimate gross mineralization rates and first-order rate constants for nitrification of native NH₄⁺. The latter were 2-4 times lower than the first-order rate constants derived from the added N pool. This discrepancy between first-order rate constants for nitrification implies that one or more process rates estimated for the added N pools cannot be applied to the native N pools. Preferential use of the added N resulted in an overestimation of the gross mineralization by 1.5-2.5-fold, emphasizing the need for critical evaluation of the assumption of equilibrium before gross mineralization rates are calculated.     

Interpreting the temperature-induced response of ammonia oxidizing microorganisms in soil using nitrogen isotope fractionation

Purpose  

Although nitrification plays a key role in the fate of soil nitrogen (N) under global warming, little information is available for the nitrifiers’ response to changing temperatures. Nitrogen isotope fractionation associated with nitrification can be a proxy of nitrifiers’ sensitivity to changing temperature. We hypothesized that the temperature-induced balance between the transport of substrate NH₄⁺ into the microbial cell (supply) and the intracellular NH₄⁺ oxidation (consumption) governs the intracellular NH₄⁺ concentration and then affects nitrification rates and associated isotope fractionations. This study was conducted to understand the microbial response of NH₄⁺ oxidation to changing temperatures by examining the effect of changing temperature on nitrification rate and apparent isotope fractionation.     

Measuring gross nitrogen mineralization, and nitrification by 15 N isotopic pool dilution in intact soil cores

The isotope dilution method for measuring gross rates of N mineralization, immobilization, and nitrification was applied to intact soil cores so that the effects of soil mixing were avoided. Soil cores were injected with solutions of either ¹⁵NH₄⁺ or ¹⁵NO₄^?; gross mineralization rates were calculated from the decline in “N enrichment of the NH: pool during a 24-h incubation; gross nitrification rates were calculated from the decline in ¹⁵N enrichment of the NO^?₃ pool; gross rates of NH₄⁺ and NO₃^? consumption were calculated from disappearance of the ¹⁵N label. The assumptions required for application of this method to intact cores are evaluated. Sensitivity analysis revealed that homogeneous mixing of added “N with ambient pools was not a necessary assumption but that bias in distribution of added label, coincident with a non-random distribution of microbial processes, would cause significant errors in rate estimates. Rate estimates were also sensitive to errors in initial ¹⁵N and ¹⁴N pool size estimates, In a silt loam soil from a grassland site, abiotic processes consumed over 30% of the added ¹⁵NH₄⁺ within minutes of adding the label to sterilized soil. Extracting a subset of soil cores at the beginning of an incubation is recommended for obtaining initial pool size estimates. Gross immobilization is probably stimulated by addition of inorganic ¹⁵N substrate and, therefore, is overestimated by the isotope dilution method. As an alternative method, a non-linear equation is given for calculating the gross immobilization rate from the appearance of ¹⁵N in chloroform-labile microbial biomass; but incomplete extraction of biomass N may result in low estimates. Details of the isotope dilution methodology (injection rates, concentrations, experimental artefacts, etc.) are described and discussed. When care is taken to understand the underlying assumptions and sources of error, the isotope dilution method provides a powerful tool for measuring gross rates of microbial transformations of soil nitrogen in intact soil cores.     

Diffusion of NH+4 and NO−3 mineralized from organic N in soil

The mineralization of native soil organic matter and the simultaneous diffusion of zero NH⁺₄ and NO^?₃ to a solution sink of zero N concentration was analysed experimentally and theoretically for a fine sandy loam soil. Experimentally, the NH₄ and NO₃ ions produced in an incubated unsaturated soil column were allowed to diffuse through a sintered glass plate into a stirred solution sink. The distribution of NH⁺₄ and NO^?₃ in the soil column was measured after various incubation times. The rate of ammonification was measured directly during incubation and the rate of nitrification modelled from nitrifier growth kinetics. A Freundlich equation was used to describe the equilibrium between soluble and exchangeable NH⁺₄ in the soil. Terms for the microbial transformation of N and the adsorption-desorption of NH⁺₄ were combined with diffusion equations which were solved numerically using finite difference methods. The model constructed was used to predict the NH⁺₄ and NO^?₃ con-centration distributions in the soil column, and good agreement was obtained between the experimental and predicted concentration profiles. The use of the model for predicting the diffusive flux of mineral N to the outer surfaces of soil peds, where it is vulnerable to leaching, was demonstrated.     

Nitrification and ammonification in acid forest litter and humus as affected by peptone and ammonium-n amendment

Samples of strongly acid forest litter and humus from beneath Sitka spruce, heather, Scots pine and larch from two sites in north-east Scotland were incubated aerobically at 20°C in the laboratory. At the Glen Tanar site, spruce litter and larch humus showed significant nitrification and ammonification whereas spruce humus and Scots pine humus produced only NH₄⁺-N. Heather humus showed no net mineralization. At the Fetteresso site, application of fertilizer N, P and K to Sitka spruce up to 3 yr previously, significantly stimulated the production of NO₃⁻-N in both litter and humus.Amendment of the samples with organic N as peptone caused significant increases in NO₃⁻-N production in those samples that already showed nitrification. The increases in NO₃⁻-N generally represented a low proportion of the added peptone-N. Amendment with NH₄⁺-N as (NH₄)₂SO₄ either had no effect or significantly reduced NO₃⁻-N production (in larch humus). The results suggest the occurrence of heterotrophic nitrification in some of these forest samples.Net immobilization of NH₄⁺-N was typically greater in NH₄⁺-N amended than in peptone amended samples, except for heather humus which showed complete immobilization of both N sources.Total mineral N produced at the end of the aerobic incubation was correlated (P < 0.01) with NH₄⁺-N produced during a 30-day anaerobic incubation at 30°C. Net NO₃⁻-N production was greater in litter than in the corresponding humus samples and was correlated (P < 0.001) with initial organic N soluble in 1 m KCl.     

Mechanisms behind the stimulation of nitrification by N input in subtropical acid forest soil

Purpose

Input of N as NH₄ ⁺ is known to stimulate nitrification and to enhance the risk of N losses through NO₃ ^? leaching in humid subtropical soils. However, the mechanisms responsible for this stimulation effect have not been fully addressed.

Materials and methods

In this study, an acid subtropical forest soil amended with urea at rates of 0, 20, 50, 100 mg N kg^?1 was pre-incubated at 25 °C and 60 % water-holding capacity (WHC) for 60 days. Gross N transformation rates were then measured using a ¹⁵N tracing methodology.

Results and discussion

Gross rates of mineralization and nitrification of NH₄ ⁺-N increased (P?<?0.05), while gross rate of NO₃ ^? immobilization significantly decreased with increasing N input rates (P?<?0.001). A significant relationship was established between the gross nitrification rate of NH₄ ⁺ and the gross mineralization rate (R ²?=?0.991, P?<?0.01), so was between net nitrification rate of NH₄ ⁺ and the net mineralization rate (R ²?=?0.973, P?<?0.05).

Conclusions

Stimulation effect of N input on the gross rate of nitrification of NH₄ ⁺-N in the acid soil, partially, resulted from stimulation effect of N input on organic N mineralization, which provides pH-favorable microsites for the nitrification of NH₄ ⁺ in acid soils (De Boer et al., Soil Biol Biochem 20:845–850, 1988; Prosser, Advan Microb Physiol 30:125–181, 1989). The stimulated gross nitrification rate with the decreased gross NO₃ ^? immobilization rate under the elevated N inputs could lead to accumulation of NO₃ ^? and to enhance the risk of NO₃ ^? loss from humid forest soils.

     

Nitrogen mineralization and assimilation at millimeter scales

This study used inoculated, artificial soil microcosms containing sand, clay, cellulose, and localized “hotspots” of highly labile, organic N-containing dead bacteria to study N mineralization and assimilation at millimeter scales. Labeling with ¹⁵NH₄⁺ along with measurement of its assimilation into microbial biomass at the bulk scale allowed estimation of gross rates of ammonification and N assimilation using isotope dilution. Time-of-flight secondary ion mass spectrometry (ToF-SIMS) analyses of transects of organic-¹⁵N across Si wafers in contact with the microcosms indicated strong gradients of ¹⁵NH₄⁺ assimilation as a function of proximity to the hotspots that were not apparent using bulk analyses. This combination of bulk and ToF-SIMS analyses represents a powerful approach to explore the physical and biochemical factors that affect N process heterogeneities in soils.     

Gross nitrogen mineralization-, immobilization-, and nitrification rates as a function of soil C/N ratio and microbial activity

A laboratory experiment was designed to challenge the idea that the C/N ratio of forest soils may control gross N immobilization, mineralization, and nitrification rates. Soils were collected from three deciduous forests sites varying in C/N ratio between 15 and 27. They were air-dried and rewetted to induce a burst of microbial activity. The N transformation rates were calculated from an isotope dilution and enrichment procedure, in which ¹⁵NH₄Cl or Na¹⁵NO₃ was repeatedly added to the soils during 7 days of incubation. The experiments suggested that differences in gross nitrogen immobilization and mineralization rates between the soils were more related to the respiration rate and ATP content than to the C/N ratio. Peaks of respiration and ATP content were followed by high rates of mineralization and immobilization, with 1-2 days of delay. The gross immobilization of NH₄⁺ was dependent on the gross mineralization and one to two orders of magnitude larger than the gross NO₃⁻ immobilization. The gross nitrification rates were negatively related to the ATP content and the C/N ratio and greatly exceeding the net nitrification rates. Taken together, the observations suggest that leaching of nitrate from forest soils may be largely dependent on the density and activity of the microbial community.     

农田土壤中氮同位素分级与硝化能力的关系研究

A laboratory-based aerobic incubation was conducted to investigate nitrogen(N) isotopic fractionation related to nitrification in five agricultural soils after application of ammonium sulfate((NH4)2SO4). The soil samples were collected from a subtropical barren land soil derived from granite(RGB),three subtropical upland soils derived from granite(RQU),Quaternary red earth(RGU),Quaternary Xiashu loess(YQU) and a temperate upland soil generated from alluvial deposit(FAU). The five soils varied in nitrification potential,being in the order of FAU YQU RGU RQU RGB. Significant N isotopic fractionation accompanied nitrification of NH+4. δ15N values of NH+4 increased with enhanced nitrification over time in the four upland soils with NH+4 addition,while those of NO-3 decreased consistently to the minimum and thereafter increased. δ15N values of NH+4 showed a significantly negative linear relationship with NH+4-N concentration,but a positive linear relationship with NO-3-N concentration. The apparent isotopic fractionation factor calculated based on the loss of NH+4 was 1.036 for RQU,1.022 for RGU,1.016 for YQU,and 1.020 for FAU,respectively. Zero- and first-order reaction kinetics seemed to have their limitations in describing the nitrification process affected by NH+4 input in the studied soils. In contrast,N kinetic isotope fractionation was closely related to the nitrifying activity,and might serve as an alternative tool for estimating the nitrification capacity of agricultural soils.     

好氧条件下两种水稻土的氮素总转化速率比较

Although to date individual gross N transformations could be quantified by ~(15)N tracing method and models,studies are still limited in paddy soil.An incubation experiment was conducted using topsoil(0-20 cm) and subsoil(20-60 cm) of two paddy soils,alkaline and clay(AC) soil and neutral and silt loam(NSL) soil,to investigate gross N transformation rates.Soil samples were labeled with either ~(15)NH4_NO_3 or NH_4~(15)NO_3,and then incubated at 25 °C for 168 h at 60%water-holding capacity.The gross N mineralization(recalcitrant and labile organic N mineralization) rates in AC soil were 1.6 to 3.3 times higher than that in NSL soil,and the gross N nitrification(autotrophic and heterotrophic nitrification) rates in AC soil were 2.4 to 4.4 times higher than those in NSL soil.Although gross NO_3~- consumption(i.e.,NO_3~- immobilization and dissimilatory NO_3~- reduction to NH_4~+ rates increased with increasing gross nitrification rates,the measured net nitrification rate in AC soil was approximately 2.0 to 5.1 times higher than that in NSL soil.These showed that high NO_3~- production capacity of alkaline paddy soil should be a cause for concern because an accumulation of NO_3~- can increase the risk of NO_3~- loss through leaching and denitrification.     

Release of mineral N from soils: Influence of inhibitors of nitrification

Nitrification inhibitors (N-Serve, ATC, and CS₂) were added to soils without N fertilizers. While the amount of nitrification of NH₄⁺-N was reduced, so was the amount of ammonification of soil N. This effect was greater with ATC and CS₂ than with N-Serve. In three field experiments, the application in the fall of ATC at 22 kg ha^?1 mixed into the soil reduced the loss of soil mineral N in early spring. Apparently, the inhibition suppressed both ammonification and nitrification of soil N during the winter, and consequently there was less NO^?₃ in soil when the wet period occurred in the spring.     

Comparison of NH4 pool dilution techniques to measure gross N fluxes in a coarse textured soil

We compared gross N fluxes by ¹⁵N pool dilution in a coarse-textured agricultural soil when ¹⁵N was applied to the soil NH₄⁺ pool by either: (i) mixing a ¹⁵NH₄NO₃ solution into disturbed soil or (ii) injection of ¹⁵NH₃ gas into intact soil cores. The two techniques produced similar results for gross N mineralization rates indicating that NH₄⁺ production in soil was not altered by soil disturbance, method of application (gas vs. solution), or amount of N applied. This was not the case for immobilization rates, which were twofold higher when ¹⁵N label was applied to the soil NH₄⁺ pool with the mixing technique compared to the injection technique. This was attributed to the fact that more NH₄⁺ was applied with the mixing technique. Estimates of gross nitrification were accompanied by large error terms meaning differences between ¹⁵N labeling methods could not be accurately assessed for this process rate.     

Regulation of Nitrification by Benzotriazole,o-Nitrophenol,m-Nitroaniline and Dicyandiamide and Pattern of NH3 Emissions from Citronella Field Fertilized with Urea

Nitrification inhibitors areuseful for reducing fertilizer related environmentalpollution. Use of such nitrification inhibitors as,benzotriazole, o-nitrophenol, m-nitroaniline anddicyandiamide has effectively regulated nitrification in acitronella (Cymbopogon winterianus Jowitt.) fieldfertilized with urea. At 450 kg N ha^-1 yr^-1, there wassubstantially higher accumulation of NH⁺ ₄-N in thesoil. Proper placement (5 cm below soil surface) offertilizers have minimized NH₃ emissions even fromnitrification inhibitor treated urea plots. Thus, thenitrification inhibitors can potentially reduceenvironmental pollution connected to NO^- ₃ in soilwhile maintaining low NH₃ gas emissions, if thefertilizer is properly placed.     

Impact of nitrogen and phosphorus additions on soil gross nitrogen transformations in a temperate desert steppe

Nutrient addition has a significant impact on plant growth and nutrient cycling. Yet, the understanding of how the addition of nitrogen (N) or phosphorus (P) significantly affects soil gross N transformations and N availability in temperate desert steppes is still limited. Therefore, a ¹⁵N tracing experiment was conducted to study these processes and their underlying mechanism in a desert steppe soil that had been supplemented with N and P for 4 years in northwestern China. Soil N mineralization was increased significantly by P addition, and N and P additions significantly promoted soil autotrophic nitrification, rather than NH₄⁺-N immobilization. The addition of N promoted dissimilatory NO₃⁻ reduction to NH₄⁺, while that of P inhibited it. Soil NO₃⁻-N production was greatly increased by N added alone and by that of N and P combined, while net NH₄⁺-N production was decreased by these treatments. Soil N mineralization was primarily mediated by pH, P content or organic carbon, while soil NH₄⁺-N content regulated autotrophic nitrification mainly, and this process was mainly controlled by ammonia-oxidizing bacteria rather than archaea and comammox. NH₄⁺-N immobilization was mainly affected by functional microorganisms, the abundance of narG gene and comammox Ntsp-amoA. In conclusion, gross N transformations in the temperate desert steppe largely depended on soil inorganic N, P contents and related functional microorganisms. Soil acidification plays a more key role in N mineralization than other environmental factors or functional microorganisms.     

Variation in the relationship between nitrification and acidification of subtropical soils as affected by the addition of urea or ammonium sulfate

The effects on nitrification and acidification in three subtropical soils to which (NH₄)₂SO₄ or urea had been added at rate of 250 mg N kg⁻¹ was studied using laboratory-based incubations. The results indicated that NH₄⁺ input did not stimulate nitrification in a red forest soil, nor was there any soil acidification. Unlike red forest soil, (NH₄)₂SO₄ enhanced nitrification of an upland soil, whilst urea was more effective in stimulating nitrification, and here the soil was slightly acidified. For another upland soil, NH₄⁺ input greatly enhanced nitrification and as a result, this soil was significantly acidified. We conclude that the effects of NH₄⁺ addition on nitrification and acidification in cultivated soils would be quite different from in forest soils. During the incubation, N isotope fractionation was closely related to the nitrifying capacity of the soils.     

Gross N mineralization–immobilization rates in heterogeneous intact soil cores can be estimated without marked error

Gross rates of N mineralization–immobilization turnover (MIT) and gross nitrification in soil can be determined by use of a ¹⁵NH₄⁺ pool dilution technique, under the assumption that native ¹⁴NH₄⁺, applied ¹⁴NH₄⁺ and microbial processes are uniformly distributed in the soil. In a laboratory investigation we compared gross N fluxes obtained from two labelling techniques applied to an arable sandy loam: (1) injection of ¹⁵NH₄⁺ solution into intact soil cores, and (2) mixing ¹⁵NH₄⁺ solution into disturbed soil. It was assumed that MIT obtained with the mixing technique reflected the true rates, since the assumption of uniformity was thought to be satisfied by this technique. MIT from the two techniques were not significantly different, thus non-uniform distribution of native ¹⁴NH₄⁺, injected ¹⁵NH₄⁺ and microbial processes in the intact-core technique did not cause a marked error in the MIT rates. In contrast the gross nitrification rates were twofold higher with the mixing technique than with the intact-core technique. Gross nitrification rates are likely to increase with the mixing technique because mixing redistributes nitrifiers and added ¹⁵NH₄⁺, and thereby increases the contact between NH₄⁺ and nitrifiers.