Limited inhibition of nitrous oxide reduction in soil in the presence of acetylene

Anaerobic flasks with two different soils contained microorganisms which effectively reduced NO₃⁻ to N₂ in the absence of C₂H₂ and in the presence or absence of CO₂. In the presence of C₂H₂, the microorganisms reduced NO₃⁻ to N₂O and the further reduction of N₂O to N₂ was temporarily inhibited. This was shown for two partial pressures of C₂H₂ (0.1 kPa and 1.0 kPa). However. after a maximum of 168 h, microorganisms were able to reduce N₂O to N₂ in the presence of C₂H₂. This was shown in the presence of CO₂ for both partial pressures of C₂H₂ and in the absence of CO₂ for 1.0 kPa C₂H₂. The absence of CO₂ delayed the complete reduction of N₂O. Microorganisms which had reduced N₂O in the presence of C₂H₂ retained this ability for at least 3 days after the original atmosphere containing C₂H₂ had been removed.     

秸秆还田对灌溉玉米田土壤反硝化及N2O排放的影响

运用乙炔抑制技术研究了不同施氮水平下秸秆还田对灌溉玉米田土壤反硝化反应和氧化亚氮(N2O)排放的影响。结果表明,土壤反硝化速率及N2O的排放受氮肥施用、秸秆处理方式及其交互作用的显著影响。与秸秆燃烧相比,不施氮或低施氮水平时,秸秆还田可刺激培养初期反硝化反应速率及N2O排放,增加培养期间N2O平均排放通量;高施氮水平时,秸秆还田可降低反硝化反应速率及反硝化过程中的N2O排放。秸秆还田可降低反硝化中N2O/N2的比例。     

A method of selective inhibition to distinguish between nitrification and denitrification as sources of nitrous oxide in soil

Summary Nitrapyrin and C₂H₂ were evaluated as nitrification inhibitors in soil to determine the relative contributions of denitrification and nitrification to total N₂O production. In laboratory experiments nitrapyrin, or its solvent xylene, stimulated denitrification directly or indirectly and was therefore considered unsuitable. Low partial pressures of C₂H₂ (2.5–5.0 Pa) inhibited nitrification and had only a small effect on denitrification, which made it possible to estimate the contribution of denitrification. The contribution of nitrification was estimated by subtracting the denitrification value from total N₂O production (samples without C₂H₂). The critical C₂H₂ concentrations needed to achieve inhibition of nitrification, without affecting the N₂O reductase in denitrifiers, must be individually determined for each set of experimental conditions.     

Relationships between soil moisture content and nitrous oxide production during nitrification and denitrification

Summary The effect of soil water content [60%–100% water-holding capacity (WHC)] on N₂O production during autotrophic nitrification and denitrification in a loam soil was studied in a laboratory experiment by selectively inhibiting nitrification with a low C₂H₂ concentration (2.1 Pa). Nitrifiers usually produced more N₂O than denitrifiers. During an initial experimental period of 0–6 days the nitrifiers produced more N₂O than the denitrifiers by a factor ranging from 1.4 to 16.5, depending on the water content and length of incubation. The highest N₂O production rate by nitrifiers was observed at 90% WHC, when the soil had become partly anaerobic, as indicated by the high denitrification rate. At 100% WHC there were large gaseous losses from denitrification, while nitrification losses were smaller except for the first period of measurement, when there was still some O₂ remaining in the soil. The use of 10 kPa C₂H₂ to inhibit reduction of N₂O to N₂ stimulated the denitrification process during prolonged incubation over several days; thus the method is unsuitable for long-term studies.     

Nitrifier denitrification as a distinct and significant source of nitrous oxide from soil

Soils are the major source of the greenhouse gas nitrous oxide (N₂O) to our atmosphere. A thorough understanding of terrestrial N₂O production is therefore essential. N₂O can be produced by nitrifiers, denitrifiers, and by nitrifiers paradoxically denitrifying. The latter pathway, though well-known in pure culture, has only recently been demonstrated in soils. Moreover, nitrifier denitrification appeared to be much less important than classical nitrate-driven denitrification. Here we studied a poor sandy soil, and show that when moisture conditions are sub-optimal for denitrification, nitrifier denitrification can be a major contributor to N₂O emission from this soil. We conclude that the relative importance of classical and nitrifier denitrification in N₂O emitted from soil is a function of the soil moisture content, and likely of other environmental conditions as well. Accordingly, we suggest that nitrifier denitrification should be routinely considered as a major source of N₂O from soil.     

Influence of thiosulfate on nitrification,denitrification, and production of nitric oxide and nitrous oxide in soil

Thiosulfate and CS₂ inhibit nitrification. The effect of the addition of thiosulfate on the turnover of inorganic N compounds was tested in an Egyptian and a German arable soil under nitrifying and denitrifying conditions. For nitrification, the soils were amended with NH _inf4 ^sup+ and incubated under aerobic conditions. For denitrification, the soils were amended with NO _inf3 ^sup- and incubated under anaerobic conditions. In both cases, the thiosulfate decreased with time while tetrathionate accumulated to an intermediate extent. Both compounds disappeared completely after <25 days. Production of CS₂ was not observed. Carbonyl sulfide was produced only in the Egyptian soil, but production decreased with increasing amounts of added thiosulfate. Under nitrifying conditions, the addition of increasing amounts of thiosulfate (25, 50, and 100 g S g^-1 dry weight) resulted in decreasing rates of NH _inf4 ^sup+ oxidation to NO _inf3 ^sup- ; it also resulted in an increasing intermediate accumulation of NO _inf2 ^sup- and NO, and in an increasing production of N₂O. Under denitrifying conditions, the addition of increasing amounts of thiosulfate did not significantly affect the rate of NO _inf3 ^sup- reduction, and resulted in an increasing intermediate accumulation of NO _inf2 ^sup- and of NO only in the German soil in which the production of N₂O was slightly inhibited by thiosulfate. These results demonstrate that the nitrification of NH _inf4 ^sup+ and NO _inf2 ^sup- was inhibited by increasing concentrations of thiosulfate and/or tetrathionate without involving the formation of volatile S compounds as potential nitrification inhibitors. Denitrification was not affected by the addition of thiosulfate.     

Oxygen inhibition of acetylene reduction (nitrogen fixation) in soil: Effect of glucose and oxygen concentrations

Sandy loam soil was amended with different concentrations of glucose and was incubated at different pO₂ levels. Under many conditions of incubation time and treatment, N₂ ase activity as determined by 1-h aerobic C₂H₂ reduction assay (flushed with Ar:O₂, 4:1 before assay) was significantly less than that determined by means of ambient assay (carried out at the pO₂ of incubation without flushing with Ar:O₂, 4:1 before assay). The difference between the N₂ase activity in aerobic assay and that in ambient assay increased with decreasing glucose and O₂ concentrations imposed during incubation. The inhition in aerobic assays was analogous to O₂-induced shut-off of N₂ase and amounted to 75 per cent inhibition after incubation at 0.06 atm pO₂ of samples amended with 0.75% glucose (w/w). Similar O₂ inhibition was observed after amendment with mannitol and with lactate. Times of incubation were chosen such that development of anaerobic N₂ase activity was either absent or too low to account for the observed effects of O₂ during assay. It was shown that 0.05 atm pC₂H₂ was adequate for routine 1-h assays of the soil system employed. Individual soil samples could be subjected to repeated 1-h assays (with removal of C₂H₂ and C₂H₄ by evacuation after each assay) thus avoiding side-effects of long exposure to C₂H₂.     

Carbon dioxide concentration in soil. Effects on nitrification,denitrification and associated nitrous oxide production

Experiments were conducted to study the effects of a range of CO₂ concentrations (ambient to 100%) on nitrification, denitrification and associated nitrous oxide (N₂O) production in a silt loam soil. It was found that increase in CO₂ concentration from 0.3 to 100% CO₂ increasingly retarded the rate of nitrification. No nitrification occurred at 100% CO₂. Nitrous oxide production associated with nitrification increased as CO₂ increased from 0.3 to 2.6% and tended to be greater as CO₂ concentration increased to 73%. At 100% CO₂, no N₂O was produced during 7 days at 25°C.Carbon dioxide did not affect N₂O production or reduction in a saturated NO₃^?amended soil or the rate of N₂O reduction in anaerobic environments.     

Nitrification and denitrification as sources of atmospheric nitrous oxide – role of oxidizable carbon and applied nitrogen

Laboratory incubation experiments were conducted to study the influence of easily oxidizable C (glucose) and mineral N (NH₄⁺ and NO₃^-) on N₂O emission, evolution of CO₂ and consumption of O₂. A flush of N₂O was always observed during the first few hours after the start of soil incubation, which was significantly higher with NH₄⁺ compared to NO₃^- applications. The increase in N₂O emission was attributed mainly to enhanced soil respiration and subsequent O₂ limitation at the microsite level. Application of NH₄⁺ helped to develop denitrifying populations since subsequent additions of NO₃^- and a C source significantly enhanced N₂O emissions. In soils treated with NH₄⁺, N₂O emissions declined rapidly, which was related to decreasing concentrations of easily oxidizable C. Addition of glucose in different amounts and pre-incubation of soil for different lengths of time (to create variation in the amount of easily oxidizable C) changed the pattern of N₂O emissions, which was ascribed to changes in soil respiration.     

区分土壤中硝化与反硝化对N2O产生贡献的方法

土壤是产生N2O的最主要来源之一.硝化和反硝化反应是产生N2O的主要机理,由于硝化和反硝化微生物同时存在于土壤中,因而硝化和反硝化作用能同时产生N2O.N2O的来源可通过使用选择性抑制剂,杀菌剂以及加入的标记底物确定.通过对生成N2O反应的每一步分析,主要从抑制反应发生的催化酶和细菌着手,总结了测量区分硝化、反硝化和DNRA反应对N2O产生的贡献方法.并对15N标记底物法,乙炔抑制法和环境因子抑制法作了详细介绍.     

区分土壤中硝化与反硝化对N2O产生贡献的方法

土壤是产生N₂O的最主要来源之一。硝化和反硝化反应是产生N₂O的主要机理，由于硝化和反硝化微生物同时存在于土壤中，因而硝化和反硝化作用能同时产生N₂O。N₂O的来源可通过使用选择性抑制剂，杀菌剂以及加入的标记底物确定。通过对生成N₂O反应的每一步分析，主要从抑制反应发生的催化酶和细菌着手，总结了测量区分硝化、反硝化和DNRA反应对N₂O产生的贡献方法。并对¹⁵N标记底物法，乙炔抑制法和环境因子抑制法作了详细介绍。     

区分土壤中硝化与反硝化对N2O产生贡献的方法

土壤是产生N2O的最主要来源之一。硝化和反硝化反应是产生N2O的主要机理,由于硝化和反硝化微生物同时存在于土壤中,因而硝化和反硝化作用能同时产生N2O。N2O的来源可通过使用选择性抑制剂,杀菌剂以及加入的标记底物确定。通过对生成N2O反应的每一步分析,主要从抑制反应发生的催化酶和细菌着手,总结了测量区分硝化、反硝化和DNRA反应对N2O产生的贡献方法。并对15N标记底物法,乙炔抑制法和环境因子抑制法作了详细介绍。     

Role of nitrifier denitrification in the production of nitrous oxide

Nitrifier denitrification is the pathway of nitrification in which ammonia (NH₃) is oxidized to nitrite (NO₂⁻) followed by the reduction of NO₂⁻ to nitric oxide (NO), nitrous oxide (N₂O) and molecular nitrogen (N₂). The transformations are carried out by autotrophic nitrifiers. Thus, nitrifier denitrification differs from coupled nitrification–denitrification, where denitrifiers reduce NO₂⁻ or nitrate (NO₃⁻) that was produced by nitrifiers. Nitrifier denitrification contributes to the development of the greenhouse gas N₂O and also causes losses of fertilizer nitrogen in agricultural soils. In this review article, present knowledge about nitrifier denitrification is summarized in order to give an exact definition, to spread awareness of its pathway and controlling factors and to identify areas of research needed to improve global N₂O budgets. Due to experimental difficulties and a lack of awareness of nitrifier denitrification, not much is known about this mechanism of N₂O production. The few measurements carried out so far attribute up to 30% of the total N₂O production to nitrifier denitrification. Low oxygen conditions coupled with low organic carbon contents of soils favour this pathway as might low pH. As nitrifier denitrification can lead to substantial N₂O emissions, there is a need to quantify this pathway in different soils under different conditions. New insights attained through quantification experiments should be used in the improvement of computer models to define sets of conditions that show where and when nitrifier denitrification is a significant source of N₂O. This may subsequently render the development of guidelines for low-emission farming practices necessary.     

Mechanistic model for nitrous oxide emission via nitrification and denitrification

Seasonal and annual N₂O fluxes from urine-affected pasture were approximated with a mechanistic model based on Michaelis-Menten kinetics. The model combined the effects of soil nitrate-N, soil ammonium-N, soil temperature and soil moisture (all from the top 5cm) to calculate N₂O emissions from nitrification (F _nit ) and denitrification (F _den ), with total N₂O emission being the sum of the two (F _tot =F _nit +F _den ). Best results were obtained when different kinetic parameters were used for periods of constant soil moisture conditions and after heavy rainfalls when a rapid change of the soil moisture status occurred. Modelled N₂O emissions over a year were within the range of uncertainties of measured N₂O emissions. Results indicate that the spatial variability of N₂O emissions at times when all the model inupt variables were constant may be related to microorganism growth dynamics or enzyme production rates. Received: 2 October 1995     

Effect of increasing oxygen concentration on total denitrification and nitrous oxide release from soil by different bacteria

Summary The effect of increasing oxygen concentrations (0, 5, 10 and 20 Vol% O₂) on total denitrification and N₂0 release was studied in model experiments using a neutral pH loamy soil relatively rich in easily decomposable organic matter and supplied with nitrate (300 g nitrate N/g dry soil). The sterilized soil was inoculated with three different denitrifying bacteria (Bacillus licheniformis,Aeromonas denitrificans andAzospirillum lipoferum) and incubated (80% WHC, 30°C). The gas volume was analysed for O₂, CO₂, N₂O, NO and N₂ by gas chromatography and the soil investigated for changes in ammonium, nitrite, nitrate, pH, total N and C as well as water-extractable C. WithB. licheniformis andAeromonas denitrificans total denitrification increased remarkably with increasing pO₂ as the result of intensified mineralization.Azospirillum lipoferum, however, showed the highest activity at 5 vol% O₂. WithB. licheniformis N₂O was released only in anaerobic conditions and at 5 Vol% O₂ (maximum) or 10 Vol% 02, but not at 20 Vol%, whereasAeromonas denitrificans produced N₂O only in the presence of He gas (maximum) or at 5 Vol% O₂. In contrast to these bacteria, N₂O production withAzospirillum lipoferum was restricted to 10 Vol% O₂ (maximum) and to 20 Vol% 02, with some traces at 5 vol% O₂. With a certain set of conditions, total denitrification and N₂O formation seem to be governed by the mineralization rate of the organisms in question. The increased demand for electron acceptors by a high turnover rate rather than the presence of anaerobic conditions seems to have determined the rate of denitrification.     

Respiration of nitrous oxide in suboxic soil

Reduction of nitrous oxide (N₂O) is an autonomous respiratory pathway. Nitrous oxide is an alternative electron acceptor to O₂ when intensive biological activity and reduced diffusivity result in an O₂ deficit. Hypoxic or anoxic micro sites may form even in well-aerated soils, and provide a sink for N₂O diffusing through the gas-filled pore space. We reproduced similar in vitro conditions in suboxic (0.15% O₂) flow-through incubation experiments with samples from a Stagnosol and from a Histosol. Apparent half-saturation constants ( k _m) for N₂O reduction were similar for both soils and were, on average, 3.8 μmol mol⁻¹ at 5°C, 5.1 μmol mol⁻¹ at 10°C, and 6.9 μmol mol⁻¹ at 20°C. Respiration of N₂O was estimated to contribute a maximum proportion of 1.7% to total respiration in the Stagnosol (pH 7.0) and 0.9% in the Histosol (pH 2.9).     

Influence of soil properties on the turnover of nitric oxide and nitrous oxide by nitrification and denitrification at constant temperature and moisture

 Soils are a major source of atmospheric NO and N₂O. Since the soil properties that regulate the production and consumption of NO and N₂O are still largely unknown, we studied N trace gas turnover by nitrification and denitrification in 20 soils as a function of various soil variables. Since fertilizer treatment, temperature and moisture are already known to affect N trace gas turnover, we avoided the masking effect of these soil variables by conducting the experiments in non-fertilized soils at constant temperature and moisture. In all soils nitrification was the dominant process of NO production, and in 50% of the soils nitrification was also the dominant process of N₂O production. Factor analysis extracted three factors which together explained 71% of the variance and identified three different soil groups. Group I contained acidic soils, which showed only low rates of microbial respiration and low contents of total and inorganic nitrogen. Group II mainly contained acidic forest soils, which showed relatively high respiration rates and high contents of total N and NH₄ ⁺. Group III mainly contained neutral agricultural soils with high potential rates of nitrification. The soils of group I produced the lowest amounts of NO and N₂O. The results of linear multiple regression conducted separately for each soil group explained between 44–100% of the variance. The soil variables that regulated consumption of NO, total production of NO and N₂O, and production of NO and N₂O by either nitrification or denitrification differed among the different soil groups. The soil pH, the contents of NH₄ ⁺, NO₂ ^– and NO₃ ^–, the texture, and the rates of microbial respiration and nitrification were among the important variables. Received: 28 October 1999