Antifungal effect of a heterotrophic nitrifier Alcaligenes faecalis

Alcaligenes faecalis suppressed the growth of 11 strains of fungal plant pathogens in vitro. When it was cultivated on a synthetic medium containing (NH4)2 SO4 as the sole nitrogen source, NH2OH, NO2- and NO3- were produced, indicating that heterotrophic nitrification was occurring. The suppressive effect of A. faecalis on plant pathogens was due to its NH2OH produced. © Rapid Science Ltd. 1998     

Nitrosospira spp. can produce nitrous oxide via a nitrifier denitrification pathway

Nitrous oxide (N(2)O) emission from soils is a major contributor to the atmospheric loading of this potent greenhouse gas. It is thought that autotrophic ammonia oxidizing bacteria (AOB) are a significant source of soil-derived N(2)O and a denitrification pathway (i.e. reduction of NO(2) (-) to NO and N(2)O), so-called nitrifier denitrification, has been demonstrated as a N(2)O production mechanism in Nitrosomonas europaea. It is thought that Nitrosospira spp. are the dominant AOB in soil, but little information is available on their ability to produce N(2)O or on the existence of a nitrifier denitrification pathway in this lineage. This study aims to characterize N(2)O production and nitrifier denitrification in seven strains of AOB representative of clusters 0, 2 and 3 in the cultured Nitrosospira lineage. Nitrosomonas europaea ATCC 19718 and ATCC 25978 were analysed for comparison. The aerobically incubated test strains produced significant (P < 0.001) amounts of N(2)O and total N(2)O production rates ranged from 2.0 amol cell(-1) h(-1), in Nitrosospira tenuis strain NV12, to 58.0 amol cell(-1) h(-1), in N. europaea ATCC 19718. Nitrosomonas europaea ATCC 19718 was atypical in that it produced four times more N(2)O than the next highest producing strain. All AOB tested were able to carry out nitrifier denitrification under aerobic conditions, as determined by production of (15)N-N(2)O from applied (15)N-NO(2) (-). Up to 13.5% of the N(2)O produced was derived from the exogenously applied (15)N-NO(2) (-). The results suggest that nitrifier denitrification could be a universal trait in the betaproteobacterial AOB and its potential ecological significance is discussed.     

Contributions of Autotrophic and Heterotrophic Nitrifiers to Soil NO and N(2)O Emissions

Soil emission of gaseous N oxides during nitrification of ammonium represents loss of an available plant nutrient and has an important impact on the chemistry of the atmosphere. We used selective inhibitors and a glucose amendment in a factorial design to determine the relative contributions of autotrophic ammonium oxidizers, autotrophic nitrite oxidizers, and heterotrophic nitrifiers to nitric oxide (NO) and nitrous oxide (N(2)O) emissions from aerobically incubated soil following the addition of 160 mg of N as ammonium sulfate kg. Without added C, peak NO emissions of 4 mug of N kg h were increased to 15 mug of N kg h by the addition of sodium chlorate, a nitrite oxidation inhibitor, but were reduced to 0.01 mug of N kg h in the presence of nitrapyrin [2-chloro-6-(trichloromethyl)-pyridine], an inhibitor of autotrophic ammonium oxidation. Carbon-amended soils had somewhat higher NO emission rates from these three treatments (6, 18, and 0.1 mug of N kg h after treatment with glucose, sodium chlorate, or nitrapyrin, respectively) until the glucose was exhausted but lower rates during the remainder of the incubation. Nitrous oxide emission levels exhibited trends similar to those observed for NO but were about 20 times lower. Periodic soil chemical analyses showed no increase in the nitrate concentration of soil treated with sodium chlorate until after the period of peak NO and N(2)O emissions; the nitrate concentration of soil treated with nitrapyrin remained unchanged throughout the incubation. These results suggest that chemoautotrophic ammonium-oxidizing bacteria are the predominant source of NO and N(2)O produced during nitrification in soil.     

Stimulation of NO and N2O emissions from soils by SO2 deposition

Sulfur dioxide (SO₂) in the atmosphere has been demonstrated to have many adverse impacts on the environment and human health. In this study, deposition of SO₂ ranging from 9.0 to 127.8 mg kg^?1 with an average of 35.7 mg S kg^?1 was found to substantially stimulate NO and N₂O emissions from soils in the humid subtropical areas of Hainan, Fujian, Jiangxi, and Yunnan provinces of China under field conditions. Laboratory tests indicated that the stimulations were mediated biologically as the effects were not observed in sterilized soils. Acidification of soil resulting from SO₂ deposition was not responsible for the stimulated NO and N₂O emissions alone as the stimulation did not occur by acidifying soil with HNO₃ treatment. By using the ¹⁵N tracing method, we found that the N₂O emissions stimulated by SO₂ deposition were from either denitrification, heterotrophic nitrification or both, but not from autotrophic nitrification. Therefore, atmospheric SO₂ deposition would most likely stimulate NO and N₂O emissions in acidic soils in which heterotrophic nitrification dominates NO and N₂O production and waterlogged soils in which denitrification dominates NO and N₂O production.     

Production of NO and N(inf2)O by Pure Cultures of Nitrifying and Denitrifying Bacteria during Changes in Aeration

Peak emissions of NO and N(inf2)O are often observed after wetting of soil. The reactions to sudden changes in the aeration of cultures of nitrifying and denitrifying bacteria with respect to NO and N(inf2)O emissions were compared to obtain more information about the microbiological aspects of peak emissions. In continuous culture, the nitrifier Nitrosomonas europaea and the denitrifiers Alcaligenes eutrophus and Pseudomonas stutzeri were cultured at different levels of aeration (80 to 0% air saturation) and subjected to changes in aeration. The relative production of NO and N(inf2)O by N. europaea, as a percentage of the ammonium conversion, increased from 0.87 and 0.17%, respectively, at 80% air saturation to 2.32 and 0.78%, respectively, at 1% air saturation. At 0% air saturation, ammonium oxidation and N(inf2)O production ceased but NO production was enhanced. Coculturing of N. europaea with the nitrite oxidizer Nitrobacter winogradskyi strongly reduced the relative levels of NO and N(inf2)O production, probably as an effect of the lowered nitrite concentration. After lowering the aeration, N. europaea produced large short-lasting peaks of NO and N(inf2)O emissions in the presence but not in the absence of nitrite. A. eutrophus and P. stutzeri began to denitrify below 1% air saturation, with the former accumulating nitrite and N(inf2)O and the latter reducing nitrate almost completely to N(inf2). Transition of A. eutrophus and P. stutzeri from 80 to 0% air saturation resulted in transient maxima of denitrification intermediates. Such transient maxima were not observed after transition from 1 to 0%. Reduction of nitrate by A. eutrophus continued 48 h after the onset of the aeration, whereas N(inf2)O emission by P. stutzeri increased for only a short period. It was concluded that only in the presence of nitrite are nitrifiers able to dominate the NO and N(inf2)O emissions of soils shortly after a rainfall event.     

N2O production from hydroxylamine oxidation and corresponding hydroxylamine oxidoreductase involved in a heterotrophic nitrifier A. faecalis strain NR

Bioprocess and Biosystems Engineering - N2O production from NH2OH oxidation involved in a heterotrophic nitrifier Alcaligenes faecalis strain NR was studied. 15N-labeling experiments showed that...     

Selective inhibition of ammonium oxidation and nitrification-linked n(2)o formation by methyl fluoride and dimethyl ether

Methyl fluoride (CH(3)F) and dimethyl ether (DME) inhibited nitrification in washed-cell suspensions of Nitrosomonas europaea and in a variety of oxygenated soils and sediments. Headspace additions of CH(3)F (10% [vol/vol]) and DME (25% [vol/vol]) fully inhibited NO(2) and N(2)O production from NH(4) in incubations of N. europaea, while lower concentrations of these gases resulted in partial inhibition. Oxidation of hydroxylamine (NH(2)OH) by N. europaea and oxidation of NO(2) by a Nitrobacter sp. were unaffected by CH(3)F or DME. In nitrifying soils, CH(3)F and DME inhibited N(2)O production. In field experiments with surface flux chambers and intact cores, CH(3)F reduced the release of N(2)O from soils to the atmosphere by 20- to 30-fold. Inhibition by CH(3)F also resulted in decreased NO(3) + NO(2) levels and increased NH(4) levels in soils. CH(3)F did not affect patterns of dissimilatory nitrate reduction to ammonia in cell suspensions of a nitrate-respiring bacterium, nor did it affect N(2)O metabolism in denitrifying soils. CH(3)F and DME will be useful in discriminating N(2)O production via nitrification and denitrification when both processes occur and in decoupling these processes by blocking NO(2) and NO(3) production.     

The contribution of nitrogen transformation processes to total N2O emissions from soils used for intensive vegetable cultivation

The rapid expansion of intensively farmed vegetable fields has substantially contributed to the total N₂O emissions from croplands in China. However, to date, the mechanisms underlying this phenomenon have not been completely understood. To quantify the contributions of autotrophic nitrification, heterotrophic nitrification, and denitrification to N₂O production from the intensive vegetable fields and to identify the affecting factors, a ¹⁵N tracing experiment was conducted using five soil samples collected from adjacent fields used for rice-wheat rotation system (WF), or for consecutive vegetable cultivation (VF) for 0.5 (VF1), 6 (VF2), 8 (VF3), and 10 (VF4) years. Soil was incubated under 50% water holding capacity (WHC) at 25°C for 96 h after being labeled with ¹⁵NH₄NO₃ or NH ₄ ¹⁵ NO₃. The average N₂O emission rate was 24.2 ng N?kg^?1 h^?1 in WF soil, but it ranged from 69.6 to 507 ng N?kg^?1 h^?1 in VF soils. Autotrophic nitrification, heterotrophic nitrification and denitrification accounted for 0.3–31.4%, 25.4–54.4% and 22.5–57.7% of the N₂O emissions, respectively. When vegetable soils were moderately acidified (pH, 6.2 to ?≥?5.7), the increased N₂O emissions resulted from the increase of both the gross autotrophic and heterotrophic nitrification rates and the N₂O product ratio of autotrophic nitrification. However, once severe acidification occurred (as in VF4, pH?≤?4.3) and salt stress increased, both autotrophic and heterotrophic nitrification rates were inhibited to levels similar to those of WF soil. The enhanced N₂O product ratios of heterotrophic nitrification (4.84‰), autotrophic nitrification (0.93‰) and denitrification processes were the most important factors explaining high N₂O emission in VF4 soil. Data from this study showed that various soil conditions (e.g., soil salinity and concentration of NO ₃ ^- or NH ₄ ⁺ ) could also significantly affect the sources and rates of N₂O emission.     

Contributions of Autotrophic and Heterotrophic Nitrifiers to Soil NO and N2O Emissions

Soil emission of gaseous N oxides during nitrification of ammonium represents loss of an available plant nutrient and has an important impact on the chemistry of the atmosphere. We used selective inhibitors and a glucose amendment in a factorial design to determine the relative contributions of autotrophic ammonium oxidizers, autotrophic nitrite oxidizers, and heterotrophic nitrifiers to nitric oxide (NO) and nitrous oxide (N₂O) emissions from aerobically incubated soil following the addition of 160 mg of N as ammonium sulfate kg⁻¹. Without added C, peak NO emissions of 4 μg of N kg⁻¹ h⁻¹ were increased to 15 μg of N kg⁻¹ h⁻¹ by the addition of sodium chlorate, a nitrite oxidation inhibitor, but were reduced to 0.01 μg of N kg⁻¹ h⁻¹ in the presence of nitrapyrin [2-chloro-6-(trichloromethyl)-pyridine], an inhibitor of autotrophic ammonium oxidation. Carbon-amended soils had somewhat higher NO emission rates from these three treatments (6, 18, and 0.1 μg of N kg⁻¹ h⁻¹ after treatment with glucose, sodium chlorate, or nitrapyrin, respectively) until the glucose was exhausted but lower rates during the remainder of the incubation. Nitrous oxide emission levels exhibited trends similar to those observed for NO but were about 20 times lower. Periodic soil chemical analyses showed no increase in the nitrate concentration of soil treated with sodium chlorate until after the period of peak NO and N₂O emissions; the nitrate concentration of soil treated with nitrapyrin remained unchanged throughout the incubation. These results suggest that chemoautotrophic ammonium-oxidizing bacteria are the predominant source of NO and N₂O produced during nitrification in soil.     

Heterotrophic nitrification by Alcaligenes faecalis: NO2-, NO3-, N2O, and NO production in exponentially growing cultures

Heterotrophic nitrification by Alcaligenes faecalis DSM 30030 was not restricted to media containing organic forms of nitrogen. In both peptone-meat extract and defined media with ammonium and citrate as the sole nitrogen and carbon sources, respectively, NO2-, NO3-, NO, and N2O were produced under aerobic growth conditions. Heterotrophic nitrification was not attributable to old or dying cell populations. Production of NO2-, NO3-, NO, and N2O was detectable shortly after cultures started growth and proceeded exponentially during the logarithmic growth phase. NO2- and NO3- production rates were higher for cultures inoculated in media with pH values below 7 than for those in media at alkaline pH. Neither assimilatory nor dissimilatory nitrate or nitrite reductase activities were detectable in aerobic cultures.     

Heterotrophic nitrification by Alcaligenes faecalis: NO2-, NO3-, N2O, and NO production in exponentially growing cultures.

Heterotrophic nitrification by Alcaligenes faecalis DSM 30030 was not restricted to media containing organic forms of nitrogen. In both peptone-meat extract and defined media with ammonium and citrate as the sole nitrogen and carbon sources, respectively, NO2-, NO3-, NO, and N2O were produced under aerobic growth conditions. Heterotrophic nitrification was not attributable to old or dying cell populations. Production of NO2-, NO3-, NO, and N2O was detectable shortly after cultures started growth and proceeded exponentially during the logarithmic growth phase. NO2- and NO3- production rates were higher for cultures inoculated in media with pH values below 7 than for those in media at alkaline pH. Neither assimilatory nor dissimilatory nitrate or nitrite reductase activities were detectable in aerobic cultures.     

Relative Rates of Nitric Oxide and Nitrous Oxide Production by Nitrifiers, Denitrifiers, and Nitrate Respirers

Biogenic emissions of nitric and nitrous oxides have important impacts on the photochemistry and chemistry of the atmosphere. Although biogenic production appears to be the overwhelming source of N₂O, the magnitude of the biogenic emission of NO is very uncertain. In soils, possible sources of NO and N₂O include nitrification by autotrophic and heterotrophic nitrifiers, denitrification by nitrifiers and denitrifiers, nitrate respiration by fermenters, and chemodenitrification. The availability of oxygen determines to a large extent the relative activities of these various groups of organisms. To better understand this influence, we investigated the effect of the partial pressure of oxygen (pO₂) on the production of NO and N₂O by a wide variety of common soil nitrifying, denitrifying, and nitrate-respiring bacteria under laboratory conditions. The production of NO per cell was highest by autotrophic nitrifiers and was independent of pO₂ in the range tested (0.5 to 10%), whereas N₂O production was inversely proportional to pO₂. Nitrous oxide production was highest in the denitrifier Pseudomonas fluorescens, but only under anaerobic conditions. The molar ratio of NO/N₂O produced was usually greater than unity for nitrifiers and much less than unity for denitrifiers. Chemodenitrification was the major source of both the NO and N₂O produced by the nitrate respirer Serratia marcescens. Chemodenitrification was also a possible source of NO and N₂O in nitrifier cultures but only when high concentrations of nitrite had accumulated or were added to the medium. Although most of the denitrifiers produced NO and N₂O only under anaerobic conditions, chemostat cultures of Alcaligenes faecalis continued to emit these gases even when the cultures were sparged with air. Based upon these results, we predict that aerobic soils are primary sources of NO and that N₂O is produced only when there is sufficient soil moisture to provide the anaerobic microsites necessary for denitrification by either denitrifiers or nitrifiers.     

温带草甸草原土壤N2O产生过程研究

以大量的室内模拟培养实验,以内蒙古温带草甸草原土壤为研究对象,利用AIM乙炔抑制法,模拟野外条件对原样土壤样品进行N₂O产生过程进行研究。实验结果表明:内蒙古温带草甸草原土壤N₂O产生过程以硝化作用为主。其中异养硝化作用起主导作用,自养硝化潜势和反硝化潜势在草原植物不同生长季节变化不同,总体上异养硝化潜势>自养硝化潜势>反硝化潜势。由于自养硝化作用在不同季节的发生,使得草甸草原土壤N₂O的产生潜势也高、低起伏变化。从而揭示了内蒙古温带草原土壤以异养硝化作用过程为主产生N₂O和N₂O排放通量较低的微生物学机理。     

A mathematical description of the behaviour of mixed chemostat cultures of an autotrophic nitrifier and a heterotrophic nitrifier/aerobic denitrifier; a comparison with experimental data

Abstract A general, unstructed mathematical model has been used to describe the behaviour of nutrient-limited growth of two bacteria in a continuous co-culture. The experimental system consisted of a two-membered mixed culture of the heterotrophic nitrifier/aerobic denitrifier, Thiosphaera pantotropha , and the autotrophic nitrifier, Nitrosomonas europaea , competing for ammonia in chemostat culture. The outcome of competition was only dependent on the Monod constants and the growth yields of the two bacteria. The model shows that both bacteria will oxidize equal amounts of ammonia when the cell ratio of T. pantotropha/N. europaea is 260.     

Nitrous oxide emissions from denitrifying activated sludge of urban wastewater treatment plants, under anoxia and low oxygenation

Batch experiments were made to better understand the mechanisms of N2O emissions from activated sludge in denitrifying conditions found in urban WWTPs, i.e. under anoxic and low oxygenation conditions. The results showed that in completely anoxic conditions, denitrification, related to a periplasmic nitrate reductase activity, is the major producer of N2O (100% of the N2O production), whereas the nitrate ammonifying activity is not significant. In a gradient of low oxygenation, the highest N2O emissions (49.7+/-3.8 microg N2O-N/g SS/h on average) occurred at a dissolved-oxygen concentration of around 0.3mg O2/L. Below 0.3mg O2/L, heterotrophic denitrification appeared to be the major process responsible for the N2O emission (100% at zero oxygenation). From 0.4 to 1.1mg O2/L, N2O emissions were due to two processes: (i) heterotrophic denitrification that represented about 40% of the N2O production, and (ii) autotrophic nitrifier denitrification that accounted for about 60%. The N2O emissions from activated sludge represented on average 0.4% of reduced NO3(-) in anoxic conditions. The N2O emissions associated with denitrification of entire nitrogen load would amount to 155 T N2O-N/year, if all the Paris wastewater was treated by a process using activated sludge.     

Reductive soil disinfestation (RSD) alters gross N transformation rates and reduces NO and N2O emissions in degraded vegetable soils

Background and aims

Continuous vegetable cultivation in greenhouses can easily induce soil degradation, which considerably affects the development of sustainable vegetable production. Recently, the reductive soil disinfestation (RSD) is widely used as an alternative to chemical soil disinfestations to improve degraded greenhouse vegetable soils. Considering the importance of nitrogen (N) for plant growth and environment effect, the internal N transformation processes and rates should be well investigated in degraded vegetable soils treated by RSD, but few works have been undertaken.

Methods

Three RSD-treated and three untreated degraded vegetable soils were chosen and a ¹⁵?N tracing incubation experiment differentially labeled with ¹⁵NH₄NO₃ or NH₄ ¹⁵NO₃ was conducted at 25 °C under 50 % water holding capacity (WHC) for 96 h. Soil gross N transformation rates were calculated using a ¹⁵?N tracing model combined with Markov Chain Monte Carlo Metropolis algorithm (Müller et al. 2007), while the emissions of N₂O and NO were also measured.

Results

RSD could significantly enhance the soil microbial NH₄ ⁺ immobilization rate, the heterotrophic and autotrophic nitrification rates, and the NO₃ ^? turnover time. The ratio of heterotrophic nitrification to total inorganic N supply rate (mineralization + heterotrophic nitrification) increased greatly from 5.4 % in untreated vegetable soil to 56.1 % in treated vegetable soil. In addition, low release potential of NO and N₂O was observed in RSD-treated vegetable soil, due to the decrease in the NO and N₂O product ratios from heterotrophic and autotrophic nitrifications. These significant differences in gross N transformation rates, the supply processes and capacity of inorganic N, and the NO and N₂O emissions between untreated and treated vegetable soils could be explained by the elimination of accumulated NO₃ ^?, increased pH, and decreased electrical conductivity (EC) caused by RSD. Noticeably, the NO₃ ^? consumption rates were still significantly lower than the NO₃ ^? production rates in RSD-treated vegetable soil.

Conclusions

Except for improving soil chemical properties, RSD could significantly alter the supply processes of inorganic N and reduce the release potential of N₂O and NO in RSD-treated degraded vegetable soil. In order to retard the re-occurrence of NO₃ ^? accumulation, acidification and salinization and to promote the long-term productivity of greenhouse vegetable fields, the rational use of N fertilizer should be paid great attention to farmers in vegetable cultivation.     

The effect of different N substrates on biological N2O production from forest and agricultural light textured soils

N₂O emissions from two slightly alkaline sandy soils, from arable land and a woodland, were determined in a laboratory experiment in which the soils were incubated with different sources of nitrogen, with or without glucose, and with 0, 1 and 100 mL C₂H₂ L^-1. Large differences in the rate of N₂O production were observed between the two soils and between the different N treatments. The arable soil showed very low N₂O emissions derived from reduced forms of N as compared with the N₂O which was produced when the soil was provided with NO ₂ ^- or NO ₃ ^- and a C source, suggesting a very active denitrifier population. In contrast, the woodland soil showed a very low denitrification activity and a much higher N₂O production derived from the oxidation of NH ₄ ⁺ and reduction of NO ₂ ^- by some processes probably mediated by autotrophic or heterotrophic nitrifiers or dissimilatory NO ₂ ^- reducers. In both soils, the highest N₂O emissions were induced by NO ₂ ^- addition. Those emissions were demonstrated to have a biological origin, as no significant N₂O emissions were measured when the soil was autoclaved.     

Nitrogen oxide gas emissions from temperate forest soils receiving long-term nitrogen inputs

From spring 2000 through fall 2001, we measured nitric oxide (NO) and nitrous oxide (N₂O) fluxes in two temperate forest sites in Massachusetts, USA that have been treated since 1988 with different levels of nitrogen (N) to simulate elevated rates of atmospheric N deposition. Plots within a pine stand that were treated with either 50 or 150 kg N ha^?1 yr^?1 above background displayed consistently elevated NO fluxes (100–200 µg N m^?2 h^?1) compared to control plots, while only the higher N treatment plot within a mixed hardwood stand displayed similarly elevated NO fluxes. Annual NO emissions estimated from monthly sampling accounted for 3.0–3.7% of N inputs to the high‐N plots and 8.3% of inputs to the Pine low‐N plot. Nitrous oxide fluxes in the N‐treated plots were generally < 10% of NO fluxes. Net nitrification rates (NRs) and NO production rates measured in the laboratory displayed patterns that were consistent with field NO fluxes. Total N oxide gas flux was positively correlated with contemporaneous measurements of NR and concentration. Acetylene inhibited both nitrification and NO production, indicating that autotrophic nitrification was responsible for the elevated NO production. Soil pH was negatively correlated with N deposition rate. Low levels (3–11 µg N kg^?1) of nitrite () were detected in mineral soils from both sites. Kinetic models describing NO production as a function of the protonated form of (nitrous acid [HNO₂]) adequately described the mineral soil data. The results indicate that atmospheric deposition may generate losses of gaseous NO from forest soils by promoting nitrification, and that the response may vary significantly between forest types under similar climatic regimes. The lowering of pH resulting from nitrification and/or directly from deposition may also play a role by promoting reactions involving HNO₂.