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1.
Summary A strain of Thiobacillus denitrificans was isolated after enrichment under anaerobic conditions by the continuous culture technique using thiosulfate as energy source and nitrate as electron acceptor and nitrogen source. The isolate was an active denitrifyer, the optimal conditions being 30°C and pH 7.5–8.0. Denitrification was inhibited by sulfate (the reaction product) above 5 g SO 4 = /l, whereas high concentrations of the substrates nitrate and thiosulfate were less harmful; nitrite affected denitrification above 0.2 g NO 2 /l. During the time course of denitrification in a batch culture growth and substrate consumption slowed down already after only half the substrate was utilized due to product inhibition. The following parameters were determined in continuous culture under nitrate limitation: max=0.11 h–1, K S=0.2 mg NO 3 /l, maximum denitrification rate=0.78 g NO 3 /g cells·h, g cells/g NO 3 , g cells/g S2O 3 = . Nitrite did not accumulate during steady state denitrification; the denitrification gas was almost pure N2. The concentrations of N2O and NO were below 1 ppm.  相似文献   

2.
Samples of sediment from Lake St. George, Ontario, Canada, were incubated in the laboratory under an initially aerobic gas phase and under anaerobic conditions. In the absence of added nitrate (NO3) there was O2-dependent production of nitrous oxide (N2O), which was inhibited by acetylene (C2H2) and by nitrapyrin, suggesting that coupled nitrification-denitrification was responsible. Denitrification of added NO3 was almost as rapid under an aerobic gas phase as under anaerobic conditions. The N2O that accumulated persisted in the presence of 0.4 atm of C2H2, but was gradually reduced by some sediment samples at lower C2H2 concentrations. Low rates of C2H2 reduction were observed in the dark, were maximal at 0.2 atm of C2H2, and were decreased in the presence of O2, NO3, or both. High rates of light-dependent C2H2 reduction occurred under anaerobic conditions. Predictably, methane (CH4) production, which occurred only under anaerobiosis, was delayed by added NO3 and inhibited by C2H2. Consumption of added CH4 occurred only under aerobic conditions and was inhibited by C2H2.  相似文献   

3.
Bradyrhizobium japonicum USDA 143 grew chemoorganotrophically under anoxic conditions with exogenous N2O as the sole terminal electron acceptor. Cell growth and dissimilatory N2O reduction were significantly inhibited by C2H2 when either N2O or N2O plus NO 3 served as terminal electron acceptor(s). Reduction of N2O accounted for 20% of the energy for cell growth in cultures supplied with NO 3 as the terminal electron acceptor. Nitrous oxide was produced stoichiometrically in cultures containing NO 3 and C2H2, but cell growth was proportionately reduced when compared with cultures supplied with an equal amount of NO 3 . Exogenous N2O delayed the reduction of NO 3 in cultures supplied with both electron acceptors. Direct amperometric monitoring of N2O respiration showed a specific activity of 0.082±0.004 moles N2O/min/mg cell protein, and azide inhibited cell respiration.  相似文献   

4.
Depth distributions of O2 respiration and denitrification activity were studied in 1- to 2-mm thick biofilms from nutrient-rich Danish streams. Acetylene was added to block the reduction of N2O, and micro-profiles of O2 and N2O in the biofilm were measured simultaneously with a polarographic microsensor. The specific activities of the two respiratory processes were calculated from the microprofiles using a one-dimensional diffusion-reaction model. Denitrification only occurred in layers where O2 was absent or present at low concentrations (of a fewM). Introduction of O2 into deeper layers inhibited denitrification, but the process started immediately after anoxic conditions were reestablished. Denitrification activity was present at greater depth in the biofilm when the NO3 concentration in the overlying water was elevated, and the deepest occurrence of denitrification was apparently determined by the depth penetration of NO3 . The denitrification rate within each specific layer was not affected by an increase in NO3 concentration, and the half-saturation concentration (Km) for NO3 therefore considered to be low (<25M). Addition of 0.2% yeast extract stimulated denitrification only in the uppermost 0.2 mm of the denitrification zone indicating a very efficient utilization of the dissolved organic matter within the upper layers of the biofilm.  相似文献   

5.
Two of nine sulfate reducing bacteria tested,Desulfobulbus propionicus andDesulfovibrio desulfuricans (strain Essex 6), were able to grow with nitrate as terminal electron acceptor, which was reduced to ammonia. Desulfovibrio desulfuricans was grown in chemostat culture with hydrogen plus limiting concentrations of nitrate, nitrite or sulfate as sole energy source. Growth yields up to 13.1, 8.8 or 9.7 g cell dry mass were obtained per mol nitrate, nitrite or sulfate reduced, respectively. The apparent half saturation constants (K s) were below the detection limits of 200, 3 or 100 mol/l for nitrate, nitrite of sulfate, respectively. The maximum growth rates {ie63-1} raised from 0.124 h-1 with sulfate and 0.150 h-1 with nitrate to 0.193 h-1 with nitrite as electron acceptor. Regardless of the electron acceptor in the culture medium, cell extracts exhibited absorption maxima corresponding to cytochromec and desulfoviridin. Nitrate reductase was found to be inducible by nitrate or nitrite, whereas nitrite reductase was synthesized constitutively. The activities of nitrate and nitrite reductases with hydrogen as electron donor were 0.2 and 0.3 mol/min·mg protein, respectively. If limiting amounts of hydrogen were added to culture bottles with nitrate as electron acceptor, part of the nitrate was only reduced to the level of nitrite. In media containing nitrate plus sulfate or nitrite plus sulfate, sulfate reduction was suppressed.The results demonstrate that the ammonification of nitrate or nitrite can function as sole energy conserving process in some sulfate-reducing bacteria.  相似文献   

6.
Production of nitric oxide in Nitrosomonas europaea by reduction of nitrite   总被引:1,自引:0,他引:1  
Nitrosomonas europaea and Nitrosovibrio sp. produced NO and N2O during nitrification of ammonium. Less then 15% of the produced NO was due to chemical decomposition of nitrite. Production of NO and especially of N2O increased when the bacteria were incubated under anaerobic conditions at decreasing flow rates of air, or at increasing cell densities. Low concentrations of chlorite (10 M) inhibited the production of NO and N2, but not of nitrite indicating that NO and N2O were not produced during the oxidative conversion of ammonium to nitrite. NO and N2O were produced during reduction of nitrite with hydrazine as electron donor in almost stoichiometric quantities indicating that reduction of nitrite was the main source of NO and N2O.  相似文献   

7.
Nitrate and nitrite concentrations in the water and nitrous oxide and nitrite fluxes across the sediment-water interface were measured monthly in the River Colne estuary, England, from December 1996 to March 1998. Water column concentrations of N2O in the Colne were supersaturated with respect to air, indicating that the estuary was a source of N2O for the atmosphere. At the freshwater end of the estuary, nitrous oxide effluxes from the sediment were closely correlated with the nitrite concentrations in the overlying water and with the nitrite influx into the sediment. Increases in N2O production from sediments were about 10 times greater with the addition of nitrite than with the addition of nitrate. Rates of denitrification were stimulated to a larger extent by enhanced nitrite than by nitrate concentrations. At 550 μM nitrite or nitrate (the highest concentration used), the rates of denitrification were 600 μmol N · m−2 · h−1 with nitrite but only 180 μmol N · m−2 · h−1 with nitrate. The ratios of rates of nitrous oxide production and denitrification (N2O/N2 × 100) were significantly higher with the addition of nitrite (7 to 13% of denitrification) than with nitrate (2 to 4% of denitrification). The results suggested that in addition to anaerobic bacteria, which possess the complete denitrification pathway for N2 formation in the estuarine sediments, there may be two other groups of bacteria: nitrite denitrifiers, which reduce nitrite to N2 via N2O, and obligate nitrite-denitrifying bacteria, which reduce nitrite to N2O as the end product. Consideration of free-energy changes during N2O formation led to the conclusion that N2O formation using nitrite as the electron acceptor is favored in the Colne estuary and may be a critical factor regulating the formation of N2O in high-nutrient-load estuaries.  相似文献   

8.
The potential for denitrification in marl and peat sediments in the Shark River Slough in the Everglades National Park was determined by the acetylene blockage assay. The influence of nitrate concentration on denitrification rate and N2O yield from added nitrate was examined. The effects of added glucose and phosphate and of temperature on the denitrification potential were determined. The sediments readily denitrified added nitrate. N2O was released from the sediments both with and without added acetylene. The marl sediments had higher rates than the peat on every date sampled. Denitrification was nitrate limited; however, the yields of N2O amounted to only 10 to 34% of the added nitrate when 100 μM nitrate was added. On the basis of measured increases in ammonium concentration, it appears that the balance of added nitrate may be converted to ammonium in the marl sediment. The sediment temperature at the time of sampling greatly influenced the denitrification potential (15-fold rate change) at the marl site, indicating that either the number or the specific activity of the denitrifiers changed in response to temperature fluctuations (9 to 25°C) in the sediment. It is apparent from this study that denitrification in Everglades sediments is not an effective means of removing excess nitrogen which may be introduced as nitrate into the ecosystem with supply water from the South Florida watershed and that sporadic addition of nitrate-rich water may lead to nitrous oxide release from these wetlands.  相似文献   

9.
Denitrification in aquatic sediments was measured by an N2O reductase assay. Sediments consumed small added quantities of N2O over short periods (a few hours). In experiments with sediment slurries, N2O reductase activity was inhibited by O2, C2H2, heat treatment, and by high levels of nitrate (1 mM) or sulfide (10 mM). However, ambient levels of nitrate (<100 μM) did not influence activity, and moderate levels (about 150 μM) induced only a short lag before reductase activity began. Moderate levels of sulfide (<1 mM) had no effect on N2O reductase activity. Nitrous oxide reductase displayed Michaelis-Menten kinetics in sediments from freshwater (Km = 2.17 μM), estuarine (Km = 14.5 μM), and alkaline-saline (Km = 501 μM) environments. An in situ assay was devised in which a solution of N2O was injected into sealed glass cores containing intact sediment. Two estimates of net rates of denitrification in San Francisco Bay under approximated in situ conditions were 0.009 and 0.041 mmol of N2O per m2 per h. Addition of chlorate to inhibit denitrification in these intact-core experiments (to estimate gross rates of N2O consumption) resulted in approximately a 14% upward revision of estimates of net rates. These results were comparable to an in situ estimate of 0.022 mmol of N2O per m2 per h made with the acetylene block assay.  相似文献   

10.
Denitrification and N2O emission from urine-affected grassland soil   总被引:1,自引:0,他引:1  
Denitrification and N2O emission rates were measured following two applications of artificial urine (40 g urine-N m–2) to a perennial rye-grass sward on sandy soil. To distinguish between N2O emission from denitrification or nitrification, urine was also applied with a nitrification inhibitor (dicyandiamide, DCD). During a 14 day period following each application, the soil was frequently sampled, and incubated with and without acetylene to measure denitrification and N2O emission rates, respectively.Urine application significantly increased denitrification and N2O emission rates up to 14 days after application, with rates amounting to 0.9 and 0.6 g N m–2 day–1 (9 and 6 kg N ha–1 day–1), respectively. When DCD was added to the urine, N2O emission rates were significantly lower from 3 to 7 days after urine application onwards. Denitrification was the main source of N2O immediately following each urine application. 14 days after the first application, when soil water contents dropped to 15% (v/v) N2O mainly derived from nitrification.Total denitrification losses during the 14 day periods were 7 g N m–2, or 18% of the urine-N applied. Total N2O emission losses were 6.5 and 3 g N m–2, or 16% and 8% of the urine-N applied for the two periods. The minimum estimations of denitrification and N2O emission losses from urine-affected soil were 45 to 55 kg N ha–1 year–1, and 20 to 50 kg N ha–1 year–1, respectively.  相似文献   

11.
Production and sources of N2O were determined in soil columns amended with autoclaved yeast cells either mixed into or added as 0.5 cm3 lumps to the soil in combination with no or 200 g NO3 --N g-1. At four occasions over a two-week study period, subsets of cores were measured for N2O production during 4-hour incubations under atmospheres of ambient air, 10 Pa of C2H2, and N2, respectively. Denitrification enzyme activity (DEA) was assessed in subsamples of cores that had been incubated continuously under air.Autoclaved yeast provided a C-source readily available for denitrifying bacteria in the soil. Nitrous oxide production was negligible in unamended columns whereas accumulated N2O losses in the presence of yeast material were substantial, varying between 15 to 49 ng N2O-N g-1 h-1. Mixing yeast into the soil caused the highest production of N2O followed by the yeast lump and no yeast treatments. Incubation in the presence of 10 Pa C2H2 indicated that denitrification was the sole source of N2O, in accordance with an increase in DEA. Nitrous oxide production and DEA peaked after 4–7 days of incubation, and both were unaffected by additional NO3 -. Two-to four-fold responses to anaerobiosis and accumulation of NO3 - and NH4 + in proximity of the lumps indicated that N2O production here was limited by relatively low C-availability. In contrast, 10- to 12-fold responses to anaerobiosis and no accumulation of inorganic N suggested a higher C-availability where yeast was mixed into the soil.  相似文献   

12.
The denitrification activity of five bacterial isolates was studied in samples of soil and freshwater sediments and from a wastewater treatment plant. The bacteria were exposed in a mineral medium supplied with Na2S2O3·5H2O (electron donor) and KNO3 (electron acceptor). The effect of different concentrations of donor and/or acceptor of electrons was evaluated by the method of acetylene inhibition of N2O-reductase. No positive influence on denitrification activity was observed with increased levels of individually added substrates. Three bacterial isolates exhibited a significant increase of denitrification activity when measured in the presence of both tested substances.  相似文献   

13.
Rhizobium hedysari strain HCNT 1 rapidly reduced nitrite to N2O, only slowly reduced nitrate to nitrite and did not exhibit nitrous oxide reductase activity. Nitrite reduction in this rhizobium strain may be a detoxification mechanism for conversion of nitrite, which inhibits O2 uptake, to non-toxic N2O. Concentrations of nitrite as small as 3 M diminished O2 uptake in whole cells. The bacterium did not couple energy conservation with nitrate or nitrite reduction. Cells neither grew anaerobically at the expense of these nitrogen oxides nor translocated protons during reduction of nitrite. Induction of nitrite reductase activity was not a response to the presence of nitrate or nitrite, but occurred instead when the O2 concentration in culture atmospheres fell to <16.5% of air saturation. Sensitivity of cytochrome o, which is synthesized only in cells grown under O2-limited conditions, may account for the toxicity of nitrite in strain HCNT 1.  相似文献   

14.
15.
N2O when used in most probable number tubes as the only electron acceptor was not consumed at dilutions, down to 10–3 per gram. Additions of fresh carbon source and NO3 after growth did not stimulate N2O reduction. Since pure cultures of denitrifiers grew well under the same conditions this result was unexpected, the explanation has not been found. This approach, if successful, has the advantage that only denitrifiers can reduce N2O to N2. Further work in designing a method to use N2O as an electron acceptor in MPN tubes is suggested.  相似文献   

16.
The capacity for dissimilatory reduction of NO3 to N2 (N2O) and NH4+ was measured in 15NO3-amended marine sediment. Incubation with acetylene (7 × 10−3 atmospheres [normal]) caused accumulation of N2O in the sediment. The rate of N2O production equaled the rate of N2 production in samples without acetylene. Complete inhibition of the reduction of N2O to N2 suggests that the “acetylene blockage technique” is applicable to assays for denitrification in marine sediments. The capacity for reduction of NO3 by denitrification decreased rapidly with depth in the sediment, whereas the capacity for reduction of NO3 to NH4+ was significant also in deeper layers. The data suggested that the latter process may be equally as significant as denitrification in the turnover of NO3 in marine sediments.  相似文献   

17.
Four new isolates were obtained from denitrifying enrichments with various electron donors using sediment samples from hypersaline soda lakes. Based on 16S rRNA gene analysis and DNA-DNA hybridization results, they were all identified as members of the Gammaproteobacteria closely associated with the AlkalispirillumAlkalilimnicola group. Two isolates were obtained from samples enriched with nitrate as electron acceptor and H2 or polysulfide as electron donors, and another two strains were obtained with N2O as the electron acceptor and sulfide or acetate as electron donors. All four new isolates, together with the type strains of the genera Alkalispirillum and Alkalilimnicola originally described as obligate aerobes, were capable of anaerobic growth with acetate using either nitrate or N2O as electron acceptors. Their denitrification pathway, however, was disrupted at the level of nitrite. RuBisCO form I gene was detected and sequenced in the new isolates and in Alkalilimnicola halodurans but not in Alkalispirillum mobile. These data, together with the evidence of Oremland et al. (Appl Environ Microbiol 68:4795–4802, 2002) on the potential of Alkalilimnicola sp. MLHE-1 for autotrophic growth with arsenite as electron donor and nitrate as electron acceptor, demonstrate much higher metabolic diversity of this specific group of haloalkaliphilic Gammaproteobacteria than was originally anticipated.  相似文献   

18.
Wetlands are sources of denitrification-derived nitrous oxide (N2O). Thus, the denitrifier community of an N2O-emitting fen (pH 4.7 to 5.2) was investigated. N2O was produced and consumed to subatmospheric concentrations in unsupplemented anoxic soil microcosms. Total cell counts and most probable numbers of denitrifiers approximated 1011 cells·gDW−1 (where DW is dry weight) and 108 cells·gDW−1, respectively, in both 0- to 10-cm and 30- to 40-cm depths. Despite this uniformity, depth-related maximum reaction rate (vmax) values for denitrification in anoxic microcosms ranged from 1 to 24 and −19 to −105 nmol N2O h−1· gDW−1, with maximal values occurring in the upper soil layers. Denitrification was enhanced by substrates that might be formed via fermentation in anoxic microzones of soil. N2O approximated 40% of total nitrogenous gases produced at in situ pH, which was likewise the optimal pH for denitrification. Gene libraries of narG and nosZ (encoding nitrate reductase and nitrous oxide reductase, respectively) from fen soil DNA yielded 15 and 18 species-level operational taxonomic units, respectively, many of which displayed phylogenetic novelty and were not closely related to cultured organisms. Although statistical analyses of narG and nosZ sequences indicated that the upper 20 cm of soil contained the highest denitrifier diversity and species richness, terminal restriction fragment length polymorphism analyses of narG and nosZ revealed only minor differences in denitrifier community composition from a soil depth of 0 to 40 cm. The collective data indicate that the regional fen harbors novel, highly diverse, acid-tolerant denitrifier communities capable of complete denitrification and consumption of atmospheric N2O at in situ pH.Nitrous oxide (N2O) is a potent greenhouse gas with a global warming potential that is 300-fold higher than that of CO2, and its concentration increased from 270 ppb in 1750 to 319 ppb in 2005 (17). N2O can be produced in soils during denitrification, nitrification, the dissimilatory reduction of nitrate to nitrite and/or ammonium (hereafter referred to as dissimilatory nitrate reduction), or the chemical transformation of nitrite or hydroxylamine (5, 7, 49). The percentage of N2O produced in any of these processes is variable, depending mainly on the redox potential, pH, and C/N ratio (49). In anoxic ecosystems such as waterlogged soils, most of the N2O is considered to be denitrification derived (7, 9). Complete denitrification is the sequential reduction of nitrate to dinitrogen (N2) via nitrite, nitric oxide (NO), and N2O (75). The main product of denitrification varies with the organism and in situ conditions and is usually either N2O or N2 (68). N2O can occur as a by-product during dissimilatory nitrate reduction when accumulated nitrite interacts with nitrate reductase to form N2O (59). The production of N2O by dissimilatory nitrate reducers is favored in environments with large amounts of readily available organic carbon (65). Thus, their contribution to nitrate-dependent production of N2O in soils is likely insignificant compared to that of denitrifiers.The oxidoreductases involved in denitrification are termed dissimilatory nitrate reductase (Nar, encoded by narGHJI, or Nap, encoded by napEDABC), nitrite reductase (Nir, encoded by nirK and nirS), NO reductase (cNor and qNor, encoded by norBC and norB, respectively), and N2O reductase (Nos, encoded by nosZ) (75). Nitrate reductase is also found in dissimilatory nitrate reducers (60). narG can therefore be used as a molecular marker to assess both denitrifiers and dissimilatory nitrate reducers, whereas nosZ is specific for the assessment of denitrifiers (25, 43, 48).Denitrification in soils is regulated by temperature, pH, substrate (i.e., carbon) availability, and water content (10, 24, 66). Although denitrification increases with increasing temperature, it can still occur at temperatures below 0°C (10, 24). Low temperatures appear to limit the activity of N2O reductase more severely than other enzymes involved in denitrification and thus yield higher relative amounts of denitrification-derived N2O (24). Although denitrification activity usually decreases under acidic conditions, the relative percentage of N2O to total denitrification-derived nitrogenous gases increases with increasing acidity, a result attributed to the sensitivity of N2O reductase to low pH (27, 70). However, denitrifier communities can be adapted to the in situ pH of the system (40, 58, 73).Wetlands are ecosystems in which denitrification is likely a dominant source of emitted N2O (7, 44, 45). The identification and analysis of main drivers for N2O production (i.e., the microbiota catalyzing N2O production and consumption) is thus of major concern in such environments. Fens are specialized wetlands characterized by soil acidity (67). However, information on acid-tolerant denitrifier communities of such wetlands is scarce. It is hypothesized that fens harbor a diverse, hitherto unknown, denitrifier community that is adapted to in situ conditions and associated with N2O fluxes (i.e., fen denitrifiers are acid tolerant and have a high affinity for nitrate and N2O). Thus, the main objectives of the present study were to evaluate the capacities of denitrifier communities of an N2O-emitting fen (20) to produce or consume N2O and to determine if a novel and diverse denitrifier community was associated with these capacities.  相似文献   

19.
Chloroplasts were prepared from peas (Pisum sativum) in glucose-phosphate medium. In the presence of dl-glyceraldehyde, they catalyzed nitrite-dependent O2 evolution (mean of 13 preparations, 17.5 μmole per mg chlorophyll per hour, sd 3.64). The optimum concentration of nitrite was 0.5 mm; 0.12 mm nitrite supported Vmax/2. The reaction was accompanied by the consumption of nitrite; 55 to 80% of the nitrite-N consumed was recovered as ammonia. In short experiments (less than 10 minutes) the O2 to nitrite ratio approached 1.5, but thereafter decreased. There was no nitrite-dependent O2 evolution with chloroplasts from plants grown without added nitrate but such chloroplasts could assimilate ammonia at about the usual rate. The results are consistent with the reduction of nitrite to ammonia involving nitrate-induced nitrite reductase and a reductant generated by the chloroplast electron transport chain.  相似文献   

20.
The nitrous oxide reductase activity of Paracoccusdenitrificans can be conveniently measured using an electrochemical method for determining N2O. Introduction of this procedure has shown that (i) N2O reductase activity is reversibly inhibited by oxygen; (ii) antimycin strongly inhibits electron flow to N2O and that the inhibition is bypassed by tetramethyl-p-phenylenediamine; (iii) ascorbate plus tetramethyl-p-phenylenediamine, presumably by donating electrons to cytochrome c, is an effective reductant for nitrous oxide reductase; (iv) in the presence of the nitrous oxide reductase inhibitor, acetylene, N2O is promptly produced from nitrite, consistent with the product of nitrite reductase being N2O.  相似文献   

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