The influence of glucose and other sources of carbon on nitrate reduction rates in two temperate forest soils

The effects of different sources of organic carbon on the potential NO ₃ ⁻ reduction rates in an acid mull and an anmoor (anmoor is a type of soil which is rich in organic matter, remains water saturated most of the time and where organic and mineral fractions are closely associated) were studied under anaerobic incubation. The disappearance of NO ₃ ⁻ was higher in the anmoor than in the mull in all cases. This contradicts our previous findings where the apparent denitrification was higher in the mull than in the anmoor in the presence of added glucose. This seems to indicate that drying of soil samples might have caused the formation of stable soil aggregates in the case of anmoor (containing 16% of organic matter). Thus a favourable condition was created for securing improved diffusion of nutrients in the anmoor. Glucose, a readily available source of carbon, led to a very high degree of NO ₃ ⁻ disappearance. Among other sources, the cellulose caused higher reduction of NO ₃ ⁻ than litter and lignin. The accumulation although small, of NH ₄ ⁺ which was presumably formed (at least partially) during dissimilatory reduction of NO ₃ ⁻ , and its biological immobilization were more apparent in the anmoor than in the mull.     

Inorganic Nitrogen Metabolism in Barley Roots Under Poorly Aerated Conditions

Nitrate reduction, nitrite reduction, and ammonium assimilationwere measured over c. 24 h in excised sterile barley roots,in air or under low oxygen tensions. Partial anoxia had relativelylittle effect, but the pathway of nitrogen assimilation wasseverely inhibited during complete anoxia, when the uptake ofnitrate ceased. Much of the nitrate which was present in theroots at the time of excision was apparently unavailable forassimilation. None of the reactions of the pathway served inplace of oxygen as an electron acceptor under anaerobic conditions.The concentration of nitrate in the external solution duringgrowth and during the experimental treatments had no directeffect on anaerobic ethanol formation, although an indirecteffect was noted which was due to variations in the carbohydratecontent of the tissue.     

Aerobic and anaerobic nitrate and nitrite reduction in free-living cells of Bradyrhizobium sp. (Lupinus)

Induction, energy gain, effect on growth, and interaction of nitrate and nitrite reduction of Bradyrhizobium sp. (Lupinus) USDA 3045 were characterized. Both nitrate and nitrite were reduced in air, although nitrite reduction was insensitive to ammonium inhibition. Anaerobic reduction of both ions was shown to be linked with energy conservation. A dissimilatory ammonification process was detected, which has not been reported in rhizobia so far. Nevertheless, anaerobic conversion of nitrate to ammonium was lower than 40%, which suggests the presence of an additional, nitrite reductase of denitrifying type. Nitrite toxicity caused a non-linear relationship between biomass produced and >2 mM concentrations of each N oxyanion consumed. At > or =5 mM initial concentrations of nitrate, a stoichiometric nitrite accumulation occurred and nitrite remained in the medium. This suggests an inhibition of nitrite reductase activity by nitrate, presumably due to competition with nitrate reductase for electron donors. Lowering of growth temperature almost completely diminished nitrite accumulation and enabled consumption as high as 10 mM nitrate, which confirms such a conclusion.     

农田和森林土壤中氧化亚氮的产生与还原

采用土壤淤浆方法对丹麦农田和山毛榉森林土壤反硝化过程中Ｎ２Ｏ的产生与还原进行了研究。同时考察了硝酸根和铵离子对反硝化作用的影响。结果表明,森林土壤反硝化活性大于农田土壤,但农田土壤中Ｎ２Ｏ还原活性大于森林土壤,表现在农田和森林土壤中Ｎ２Ｏ／Ｎ２的产生比率分别为０．１１和３．６５。硝酸根和铵离子能促进两种土壤中的Ｎ２Ｏ产生,但可降低农田土壤中的Ｎ２Ｏ还原速率,与农田土壤相比,硝酸根可降低森林土壤Ｎ２     

Nitrogen removal from wastewaters at low C/N ratios with ammonium and acetate as electron donors

A denitrifying upflow anaerobic sludge blanket (UASB) reactor was operated at different nitrate loading rates at a C/N ratio of 1.2, with acetate as an electron donor. This resulted in an increase in the accumulation of nitrite. After this, the UASB reactor was supplemented with 100 mg NH4+-Nl(-1) d(-1), while acetate was gradually limited in the medium. This prevented nitrite accumulation at a C/N ratio of 0.6 due to an enhanced nitrite reduction rate achieved in the reactor. An increasing amount of ammonium was consumed when the C/N ratio was lowered in the medium. This suggested that ammonium was used as an alternative electron donor during denitrification, which is supported by nitrogen balances. Nitrite was shown to be toxic for the nitrogen removal process at 200-400 mg NO2--N(l(-1) when the C/N ratio was decreased to 0.4 leading to formation of ammonium. The present study showed that addition of ammonium as an alternative electron donor for denitrification achieved a nitrogen removal process with negligible accumulation of undesirable intermediates.     

Anaerobic nitrate reduction to ammonium in two strains isolated from coastal marine sediment: A dissimilatory pathway

Abstract: A total of 28 nitrate-reducing bacteria were isolated from marine sediment (Mediterranean coast of France) in which dissimilatory reduction of nitrate to ammonium (DRNA) was estimated as 80% of the overall nitrate consumption. Thirteen isolates were considered as denitrifiers and ten as dissimilatory ammonium producers. ¹⁵N ammonium production from ¹⁵N nitrate by an Enterobacter sp. and a Vibrio sp., the predominant bacteria involved in nitrate ammonification in marine sediment, was characterized in pure culture studies. For both strains studied, nitrate-limited culture (1 mM) produced ammonium as the main product of nitrate reduction (> 90%) while in the presence of 10 mM nitrate, nitrite was accumulated in the spent media and ammonia production was less efficient. Concomitantly with the dissimilation of nitrate to nitrite and ammonium the molar yield of growth on glucose increased. Metabolic products of glucose were investigated under different growth conditions. Under anaerobic conditions without nitrate, ethanol was formed as the main product; in the presence of nitrate, ethanol disappeared and acetate increased concomitantly with an increased amount of ammonium. These results indicate that nitrite reduction to ammonium allows NAD regeneration and ATP synthesis through acetate formation, instead of ethanol formation which was favoured in the absence of nitrate.     

The effects of different electron donors on anaerobic nitrogen transformations and denitrification processes in Lake Taihu sediments

Nitrogen transformations in anaerobic sediments and leachate in Lake Taihu were simulated in the laboratory. Ammonium, nitrate and nitrite were analyzed after incubation under anaerobic conditions. Different reductive states and pH values were obtained by using different electron donors, such as glucose, sucrose, potato starch and sodium acetate. Chemical nitrogen transformation mechanisms were discussed relative to physico-chemical properties of lake sediment. Results demonstrated that nitrogen transformations in anaerobic conditions supplemented with different electron donors varied, and supplementation with certain electron donors may enhance nitrogen removal from anaerobic sediments. Among the four electron donors studied, higher nitrogen removal efficiencies were observed with acetate and starch. Saccharides, such as glucose, sucrose and starch, stimulate nitrate reduction to nitrite, while acetate stimulates nitrate reduction to ammonium.     

Dissimilatory nitrate reduction in fungi under conditions of hypoxia and anoxia: a review

Recent progress in studies of anaerobic nitrate reduction and nitrous oxide formation in fungi has been reviewed. Current understanding of the biochemistry of nitrate and nitrite reduction to nitrous oxide and ammonium under oxygen limitation is presented, with emphasis on patterns of fungal co-denitrification, properties of the enzymes involved, and prevalence of nitrate respiration among fungal species.     

Dissimilatory nitrate reduction in fungi under conditions of hypoxia and anoxia: A review

Recent progress in studies of anaerobic nitrate reduction and nitrous oxide formation in fungi has been reviewed. Current understanding of the biochemistry of nitrate and nitrite reduction to nitrous oxide and ammonium under oxygen limitation is presented, with emphasis on patterns of fungal co-denitrification, properties of the enzymes involved, and prevalence of nitrate respiration among fungal species.     

Nitrite utilization by excised Zea mays L. roots under anaerobic conditions

Under both aerobic and anaerobic conditions exogenously supplied nitrite was utilized by sterile excised Zea mays L. root. A slightly greater quantity of nitrite was used under aerobic conditions than under anaerobiosis. The uncoupler of oxidative phosphorylation, pentachlorophenol (PCP), diminished the utilization of nitrite under aerobic and anaerobic conditions resulting in a net accumulation of nitrite rather than a net disappearance of nitrite. Nitrite supplied together with nitrite resulted in a slight reduction in the level of nitrite utilized. Supply of exogenous nitrite had no effect on nitrate reduction under aerobic or anaerobic conditions. A net accumulation of nitrite occurs only when roots are supplied with nitrate in the absence of added nitrite. However, the level of nitrite accumulated under anaerobiosis, when roots were supplied with nitrate only, was found to be a fraction of the quantity of nitrite utilized when roots were supplied with nitrite under anaerobiosis. Nitrate utilization far exceeded the level of nitrite accumulated under anaerobiosis when roots were supplied with nitrate only.     

Excessive production of ammonium from nitrate by some methanol-assimilating yeast strains

Summary Some strains ofCandida boidinii excrete ammonium when grown in nitrate methanol medium under conditions of oxygen limitation but not in wellaerated cultures or under anaerobic conditions. With other carbon sources ammonium excretion is observed only in the late exponential and in the stationary growth phase. Ammonium excretion by methanol-grown cultures approximately equals assimilatory nitrate reduction. The share of total nitrate reduction in the electron transport of methanol-grown cultures is about 18%. Increase in cell yield of oxygen-limited cultures due to nitrate addition was not observed. Nitrate reduction byC. boidinii appears to be not of the true dissimilatory type. Nor is it restricted to the assimilatory type: it is not inhibited or repressed by ammonium and the ammonium excretion occurs in quantities too large to be attributed to amino acid degradation.     

On the Physiological Role of Anaerobically Synthesized Lipids in <Emphasis Type="Italic">Oryza sativa</Emphasis> Seedlings

The objective of this work was to elucidate a possible adaptive role of lipid biosynthesis and unsaturated fatty acids (FAs), esterified to lipids, as terminal acceptors of electrons, alternative to molecular oxygen, in the shoots of rice seedlings (Oryza sativa L.) under conditions of strict anoxia. Biosynthesis of lipids and their accumulation, as well as the reduction of double bonds in unsaturated FAs, were studied by electron microscopic observation of the accumulation of lipid bodies in the cytoplasm and by the biochemical analysis of FAs in shoot lipids before and after anaerobic incubation of the shoots. The experiments were carried out with intact coleoptiles after 5 and 8 days of anaerobic germination of seeds (primary anoxia) and with detached shoots, preliminarily grown in air and then subjected to anoxia in the presence of 2% glucose for 48 h (secondary anoxia). In these experiments, lipid bodies did not accumulate in the cytoplasm under anoxic conditions. Lipid bodies appeared only during 48-h anaerobic incubation of detached coleoptiles in the absence of exogenous glucose, when mitochondria degraded. There was no change either in the double bond index of FAs, or in the qualitative and quantitative composition of FAs during shoot anaerobic incubation. We conclude that neither lipids synthesized under anaerobic conditions nor esterified unsaturated FAs are involved in plant adaptation to anaerobiosis as terminal acceptors of electrons, alternative to molecular oxygen. Lipid biosynthesis under anoxic conditions, which was demonstrated for anoxia-tolerant seedlings of Oryza sativa and Echinochloa phyllopogon in experiments with radioactive precursors, ¹⁴C-acetate and ³H-glycerol, is only the manifestation of a turnover of saturated FAs and various classes of lipids, which stabilizes cell membranes under adverse conditions of strict anoxia.__________Translated from Fiziologiya Rastenii, Vol. 52, No. 4, 2005, pp. 540–548.Original Russian Text Copyright © 2005 by Generosova, Vartapetian.     

Influence of NO3 - and NH4 + nutrition on fermentation,nitrate reductase activity and adenylate energy charge of roots of Carex pseudocyperus L. and Carex sylvatica Huds. exposed to anaerobic nutrient solutions

The adenylate energy charge, production of ethanol and lactate, and nitrate reductase activity were determined in order to study the influence of different nitrogen sources on the metabolic responses of roots of Carex pseudocyperus L. and Carex sylvatica HUDS. exposed to anaerobic nutrient solutions. Determination of adenylates was carried out by means of a modified HPLC technique. Total quantity of adenylates was higher in Carex pseudocyperus than in Carex sylvatica under all conditions. In contrast, the adenylate energy charge was only slightly different between the species and decreased more or less in relation to the applied nitrogen source under oxygen deficiency. The adenylate energy charge in roots of plants under nitrate nutrition showed a smaller decrease under anaerobic environmental conditions than plants grown with ammonium or nitrate/ammonium. Roots of nitrate-fed plants showed a lower ethanol and lactate production than ammonium/nitrate- and ammonium-fed plants. Ethanol production was higher in C. pseudocyperus, formation of lactate was lower compared to that in Carex sylvatica. The activity of enzymes involved in fermentation processes (ADH, LDH and PDC) was enhanced significantly after 24 hours of exposure to anaerobic nutrient solutions in roots of both species. The induction of these enzymes was only slightly influenced by different nitrogen supply. In vivo nitrate reductase activity increased almost 3-fold compared to the aerobic treatment in both species and overcompensated loss of NADH reoxidation capacity caused by decrease of ethanol and lactate development. Induction of in vitro nitrate reductase activity was enhanced 313% in C. pseudocyperus and 349% in C. sylvatica under anaerobic environmental conditions and nitrate supply. These results indicate that nitrate may serve as an alternative electron acceptor in anaerobic plant root metabolism and that the nitrate-supported energy charge may be due to an accelerated glycolytic flux resulting from a more effective NADH reoxidation capacity by nitrate reduction plus fermentation than by fermentation alone.Abbreviations ADH alcohol dehydrogenase - AEC adenylate energy charge - DMSO dimethyl sulfoxide - EDTA ethylen diamine tetraacetic acid - HPLC high performance liquid chromatography - LDH lactate dehydrogenase - NRA nitrate reductase activity - PCA perchloric acid - PDC pyruvate decarboxylase - PVP polyvinylpyrrolidone - PVPP polyvinylpolypyrrolidone - TCA trichloroacetic acid, Tris-tris(hydroxymethyl)aminomethane     

Anaerobic ammonium oxidation discovered in a denitrifying fluidized bed reactor

Abstract Until now, oxidation of ammonium has only been known to proceed under aerobic conditions. Recently, we observed that NH₄⁺ was disappearing from a denitrifying fluidized bed reactor treating effluent from a methanogenic reactor. Both nitrate and ammonium consumption increased with concomitant gas production. A maximum ammonium removal rate of 0.4 kg N · m⁻³ · d⁻¹ (1.2 mM/h) was observed. The evidence for this anaerobic ammonium oxidation was based on nitrogen and redox balances in continuous-flow experiments. It was shown that for the oxidation of 5 mol ammonium, 3 mol nitrate were required, resulting in the formation of 4 mol dinitrogen gas. Subsequent batch experiments confirmed that the NH₄⁺ conversion was nitrate dependent. It was concluded that anaerobic ammonium oxidation is a new process in which ammonium is oxidized with nitrate serving as the electron acceptor under anaerobic conditions, producing dinitrogen gas. This biological process has been given the name ‘Anammox” (anaerobic ammonium oxidation), and has been patented.     

Nitrogen transformation under different dissolved oxygen levels by the anoxygenic phototrophic bacterium <Emphasis Type="Italic">Marichromatium gracile</Emphasis>

Marichromatium gracile: YL28 (M. gracile YL28) is an anoxygenic phototrophic bacterial strain that utilizes ammonia, nitrate, or nitrite as its sole nitrogen source during growth. In this study, we investigated the removal and transformation of ammonium, nitrate, and nitrite by M. gracile YL28 grown in a combinatorial culture system of sodium acetate-ammonium, sodium acetate-nitrate and sodium acetate-nitrite in response to different initial dissolved oxygen (DO) levels. In the sodium acetate-ammonium system under aerobic conditions (initial DO?=?7.20–7.25 mg/L), we detected a continuous accumulation of nitrate and nitrite. However, under semi-anaerobic conditions (initial DO?=?4.08–4.26 mg/L), we observed a temporary accumulation of nitrate and nitrite. Interestingly, under anaerobic conditions (initial DO?=?0.36–0.67 mg/L), there was little accumulation of nitrate and nitrite, but an increase in nitrous oxide production. In the sodium acetate-nitrite system, nitrite levels declined slightly under aerobic conditions, and nitrite was completely removed under semi-anaerobic and anaerobic conditions. In addition, M. gracile YL28 was able to grow using nitrite as the sole nitrogen source in situations when nitrogen gas produced by denitrification was eliminated. Taken together, the data indicate that M. gracile YL28 performs simultaneous heterotrophic nitrification and denitrification at low-DO levels and uses nitrite as the sole nitrogen source for growth. Our study is the first to demonstrate that anoxygenic phototrophic bacteria perform heterotrophic ammonia-oxidization and denitrification under anaerobic conditions.     

Role of phospholipids in the aerobic endogenous metabolism of freshwater mussel spermatozoa

Aerobic incubation of the spermatozoa of freshwater mussel (Hyriopsis schlegelii) caused a reduction in acyl-ester content and a corresponding diminution in lipid-phosphorus content. Anaerobic incubation also caused a reduction in acyl ester but a smaller reduction in lipid phosphorus, while it brought on an accumulation of free fatty acids. It was mainly palmitic and stearic acids which were accumulated during the anaerobic incubation, and they were preferentially metabolized during the aerobic incubation. Complex lipids from the spermatozoa consisted mainly of diacyl glycerophospholipids, in which phosphatidylethanolamine was predominant, followed by lecithin, while plasmalogen was a minor component. After aerobic incubation of the spermatozoa, there was a marked decrease in ethanolamine-containing phospholipid fraction. However, no diminution was observed in the plasmalogen content. These results lead to the conclusion that mussel spermatozoa utilize as a primary energy source fatty acids which are derived from the breakdown of intracellular diacyl glycerophospholipids.     

Patterns and drivers of anaerobic nitrogen transformations in sediments of thermokarst lakes

Significant attention has been given to the way in which the soil nitrogen (N) cycle responds to permafrost thaw in recent years, yet little is known about anaerobic N transformations in thermokarst lakes, which account for more than one-third of thermokarst landforms across permafrost regions. Based on the N isotope dilution and tracing technique, combined with qPCR and high-throughput sequencing, we presented large-scale measurements of anaerobic N transformations of sediments across 30 thermokarst lakes over the Tibetan alpine permafrost region. Our results showed that gross N mineralization, ammonium immobilization, and dissimilatory nitrate reduction rates in thermokarst lakes were higher in the eastern part of our study area than in the west. Denitrification dominated in the dissimilatory nitrate reduction processes, being two and one orders of magnitude higher than anaerobic ammonium oxidation (anammox) and dissimilatory nitrate reduction to ammonium (DNRA), respectively. The abundances of the dissimilatory nitrate reduction genes (nirK, nirS, hzsB, and nrfA) exhibited patterns consistent with sediment N transformation rates, while α diversity did not. The inter-lake variability in gross N mineralization and ammonium immobilization was dominantly driven by microbial biomass, while the variability in anammox and DNRA was driven by substrate supply and organic carbon content, respectively. Denitrification was jointly affected by nirS abundance and organic carbon content. Overall, the patterns and drivers of anaerobic N transformation rates detected in this study provide a new perspective on potential N release, retention, and removal upon the formation and development of thermokarst lakes.     

The influence of nitrate concentration upon the end-products of nitrate dissimilation by bacteria in anaerobic salt marsh sediment

Abstract Experiments were carried out with slurries of saltmarsh sediment to which varying concentrations of nitrate were added. The acetylene blocking technique was used to measure denitrification by accumulation of nitrous oxide, while reduction of nitrate to nitrite and ammonium was also measured. There was good recovery of reduced nitrate and at the smallest concentration of nitrate used (250 μM) there was approximately equal reduction to either ammonium or nitrous oxide (denitrification). Nitrite was only a minor end-product of nitrate reduction. As the nitrate concentration was increased the proportion of the nitrate which was denitrified to nitrous oxide increased, to 83% at the greatest nitrate concentration used (2 mM), while reduction to ammonium correspondingly decreased. This change was attributed either to a greater competitiveness by the denitrifiers for nitrate as the ratio of electron donor to electron acceptor decreased; or to the increased production of nitrite rather than ammonium by fermentative bacteria under high nitrate, the nitrite then being reduced to nitrous oxide by denitrifying bacteria.     

Nitrification and occurrence of Nitrobacter by MPN-PCR in low and high nitrifying coniferous forest soils

The nitrification of three coniferous forest soils was investigated: a podzol (Fontainebleau, France) with low N deposition and no nitrate accumulation, an acid mull (Haldon, UK) with low N deposition and moderate nitrate accumulation, and a podzol (Wekerom, The Netherlands) with high N deposition and high nitrate accumulation. Twenty-one months in situ lysimeter experiments and short-term and long-term in vitro incubations were performed to respectively establish the status of NO ₃ ^- accumulation and potential nitrification of these soils. These complementary approaches allowed to conclude that the absence of NO ₃ ^- accumulation in Fontainebleau soil was effectively due to a lack of nitrifing activity in the whole profile while the high NO ₃ ^- accumulation observed in the Wekerom soil essentially resulted from an active nitrifying activity in the Oh horizon of this soil. For the Haldon soil, the inadequacy between the lysimeter and the short-term in vitro experiment was discussed.Surprisingly, relatively high densities of the nitrite-oxidiser Nitrobacter (enumerated by PCR-MPN technique) were found in all the studied horizons of the three soils. Moreover, a long-term incubation experiment showed that inhibition of nitrification in the Oh Fontainebleau soil could be removed after submitting the soil samples to constant conditions during 82 days. This demonstrates that in these soils, ammonium-oxidisers (as well as nitrite-oxidisers) are present and that extrinsic factors were involved in the inhibition of nitrification.