Effect of aeration on denitrification byOchrobactrum anthropi SY509

Aeration was found to affect the biological denitrification byOchrobactrum anthropi SY509. Although cell growth was vigorous under 1 vvm of aeration and an agitation speed of 400 rpm in a 3-L jar fermentor, almost no nitrate was removed. Yet under low agitation speeds (100, 200, and 300 rpm), denitrification occurred when the dissolved oxygen was exhausted shortly after the inoculation of the microorganism.Ochrobactrum anthropi SY509 was found to express highly active denitrifying enzymes under anaerobic conditions. The microorganism also synthesized denitrifying enzymes under aerobic conditions (1 vvm and 400 rpm), yet their activity was only 60% of the maximum level under anaerobic conditions and the nitrate removal efficiency was merely 15%. However, although the activities of the denitrifying enzymes were inhibited in the presence of oxygen, they were fully recovered when the conditions were switched to anaerobic conditions.     

Denitrification Potential in Lake Sediment Increases Across a Gradient of Catchment Agriculture

Intensification of catchment agriculture has increased nutrient loads and accelerated eutrophication in some lakes, often resulting in episodic harmful algal blooms or prolonged periods of anoxia. The influence of catchment agriculture on lake sediment denitrification capacity as a nitrogen (N) removal mechanism in lakes is largely unknown, particularly in contrast to research on denitrification in agricultural streams and rivers. We measured denitrification enzyme activity (DEA) to assess sediment denitrification potential in seven monomictic and three polymictic lakes that range in the proportion of agriculture in the catchment from 3 to 96% to determine if there is a link between agricultural land use in the lake catchment and sediment denitrification potential. We collected sediment cores for DEA measurements over 3 weeks in austral spring 2008 (October–November). Lake Okaro, with 96% catchment agriculture, had approximately 15 times higher DEA than Lake Tikitapu, with 3% catchment agriculture (232.2 ± 55.9 vs. 15.9 ± 4.5 μg N gAFDM⁻¹ h⁻¹, respectively). Additionally, sediment denitrification potential increased with the proportion of catchment in agriculture (R ² = 0.85, P < 0.001). Our data suggest that lakes retain a high capacity to remove excess N via denitrification under increasing N loads from higher proportions of catchment agriculture. However, evidence from the literature suggests that despite a high capacity for denitrification and longer water residence times, lakes with high N loads will still remove a smaller proportion of their N load. Lakes have a denitrification potential that reflects the condition of the lake catchment, but more measurements of in situ denitrification rates across lake catchments is necessary to determine if this capacity translates to high N removal rates.     

闽江河口短叶茳芏群落湿地沉积物反硝化强度对盐度的响应

为了探讨河口区湿地沉积物反硝化强度对盐度的响应,2015年7月选择闽江河口覆盖短叶茳芏的鳝鱼滩和道庆洲湿地沉积物为研究对象,采用密封厌氧培养,计算不同培养时期在不同盐度下的反硝化速率。结果表明,在反硝化培养的初期,鳝鱼滩的反硝化速率从低盐度到高盐度分别为(15.5±1.38)、(4.28±8.46)、(12.94±0.24)mg kg~(-1)d~(-1),道庆洲分别为(31.93±4.89)、(30.66±5.375)、(36.69±3.44)mg kg~(-1)d~(-1),鳝鱼滩的10天反硝化速率在0和10盐度下降幅度分别是5天的36.97%、53.01%,高于道庆洲22.64%、28.84%,后期随着时间的变化整体上呈现减小的趋势并趋于0。另外,鳝鱼滩和道庆洲反硝化活性随着时间呈现逐渐增加的趋势,其鳝鱼滩变化幅度在95.6%—99.8%,道庆洲为90.0%—96.7%,整个培养期间,鳝鱼滩的反硝化活性均高于道庆洲的反硝化活性。以上结果反映出盐度对闽江河口湿地沉积物的反硝化速率起到一定的抑制作用。     

Denitrification in lucerne nodules is not involved in nitrite detoxification

Dissimilatory reduction of ionic nitrogen oxides to gaseous forms such as nitrous oxide or nitrogen can be carried out by free living or symbiotic forms of some strains of Rhizobium meliloti. In this paper we investigate whether bacteroid denitrification plays a role in the alleviation of the inhibitory effects of nitrate on nitrogen fixation both in bacteroid incubations as in whole nodules. The presence of a constitutive nitrate reductase (NR) activity in isolated bacteroids caused nitrite accumulation in the incubation medium, and acetylene reduction activity in these bacteroids was progressively inhibited, since nitrite reductase (NiR) activity was unable to reduce all the nitrite produced by NR and denitrification occurred slowly. Even nodules infiltrated with nitrate and nitrite failed to increase gaseous forms of nitrogen substantially, indicating that nitrite availability was not limiting denitrification by bacteroids. In spite of the low rates of bacteroidal denitrification, the effect of nodule denitrification on the inhibition of nitrogen fixation by nitrate in whole plants was tested. For that purpose, lucerne plants (Medicago sativa L. cv. Aragon) were inoculated with two Rhizobium meliloti strains: 102-F-65 (non denitrifying) and 102-F-51 (a highly denitrifying strain). After a seven days nitrate treatment, both strains showed the same pattern of inhibition, and it occurred before any nitrate or nitrite accumulation within the nodules could be detected. This observation, together with the lack of alleviation of the ARA inhibition in the denitrifying strain, and the limited activity of dissimilatory nitrogen reduction present in these bacteroids, indicate a role other than nitrite detoxification for denitrification in nodules under natural conditions.     

Dynamics of Soil Denitrifier Populations: Relationships between Enzyme Activity, Most-Probable-Number Counts, and Actual N Gas Loss

To better understand temporal variability in soil denitrification, denitrifying enzyme activity (DEA) and denitrifier populations (as determined by most-probable-number [MPN] counts) were measured in field and laboratory experiments. Measurements of DEA and MPN provided highly contradictory indications of denitrifier dynamics. In laboratory incubations, under conditions favoring active denitrification, the synthesis of new denitrifying enzymes and the actual amount of denitrification were closely related. In other experiments, however, both DEA and MPN counts were poor indicators of actual denitrification. In some cases, we found significant increases in DEA but no significant production of N gas. Except with unnaturally high substrate amendments, changes in DEA were small relative both to the persistently high DEA background and to changes in MPN. As estimated by MPN counts, denitrifier populations increased significantly during denitrification events. It was apparent that only a small fraction of the denitrifiers were included in the MPN counts, but it appeared that this isolatable fraction increased during periods of active denitrifier growth. Use of DEA as an index of biomass of cells which have synthesized denitrifying enzymes suggested that denitrifier populations were persistent, stable, and much larger than indicated by MPN procedures.     

Denitrification in deep subsurface sediments

Dissimilatory nitrate reduction (denitrification) in subsurface sediments by indigenous microflora was investigated in samples obtained over a range of depths from 0 to 289 m. Denitrifying activity in sediment samples retrieved from similar stratigraphic horizons at four different sites was determined by measuring the accumulation of N₂O using the acetylene blockage technique. Denitrification was detected in unamended samples which received only prereduced deionized water at almost all depths in all sediments sampled. The surface sediments showed the highest denitrification activity. In the deeper sediments, denitrifying activity was much higher in saturated sandy samples and lower or absent in drier clay samples. Addition of nitrate enhanced denitrification activity in all samples from below the water table down to the maximum depth sampled (289 m), while addition of a carbon (succinate) source in general had no stimulatory effect. These results show that denitrifying microorganisms were present in all of the deep subsurface sediments tested in this study. Furthermore, these results suggest that adequate supplies of metabolizable organic carbon were available to support denitrifying activity. However, denitrification may be limited by inadequate supplies of nitrate in the sediments.     

Effect of crude oil on denitrification and sulfate reduction in marine sediments

The denitrifying activity was measured in different types of sediment from the Mediterranean coast of France before, and after, a massive contamination (30–100 g kg^-1 sediment) of hydrocarbons. A closed system was used in order to maintain anoxic conditions and to control substrates and gaseous products concentrations. We have demonstrated that the respiratory metabolism was inhibited in all cases following an incubation time of 20 to 50 days. At this time, the addition of lactate restore the denitrifying activity. The inhibitory effect of crude oil was not related to an alteration of bacterial cells, but to changes in environmental conditions allowing denitrification. The presence of hydrocarbons in the sediments causes a decrease in the redox potential and a concomitant stimulation of the sulfate reduction.     

Differentiated free-living and sediment-attached bacterial community structure inside and outside denitrification hotspots in the river–groundwater interface

This study assessed the functional significance of attached and free-living bacterial communities involved in the process of denitrification in a shallow aquifer of a riparian zone (Garonne River, SW France). Denitrification enzyme activity (DEA), bacterial density (BD) and bacterial community composition (BCC) were measured in two aquifer compartments: the groundwater and the sandy fraction of the sediment deposit. Samples were collected in wells located inside (IHD) and outside (OHD) identified hotspots of denitrification. Despite high BD values (up to 1.14 × 10¹² cells m⁻³), DEA was not detected in the water compartment (< 0.32 mg N–N₂O m⁻³ d⁻¹). The sandy fraction showed detectable DEA (up to 1,389 mg N–N₂O m⁻³ d⁻¹) and, consistent with BD pattern, higher DEA values were measured in IHD zones than in OHD zones. The BCC assessed by 16S rDNA polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) partly supported this result: attached and free-living communities were significantly different (< 30% similarity) but patterns of BCC did not cluster according to IHD and OHD zones. Targeting the denitrifying communities by means of a culture enrichment step prior to 16S rDNA PCR-DGGE showed that the free-living and sediment attached communities differed. Most sequences obtained from DGGE profiles of denitrifying communities were affiliated to Proteobacteria and showed low genetic distance with taxa that have already been detected in aquifers (e.g., Azoarcus sp., Acidovorax sp. and Pseudomonas spp.). This study confirms that in the aquifer the sediment-attached fraction exhibits different functions (DEA) from free-living communities and suggests that this functional difference is related to the communities’ structure.     

Short-term influence of recolonisation by the polycheate worm [2pt] Nereis succinea on oxygen and nitrogen fluxes and [2pt] denitrification: a microcosm simulation

The short-term effects of sediment recolonisation by Nereis succinea on sediment-water column fluxes of oxygen and dissolved inorganic nitrogen, and rates of denitrification, were studied in microcosms of homogenised, sieved sediments. The added worms enhanced oxygen uptake by the sediments, due to the increased surface area provided by the burrow walls and the degree of stimulation was stable with time. Similarly, ammonium fluxes to the water column were stimulated by N. succinea, but declined over the 3 day incubation in all microcosms including the controls. Nitrate fluxes were generally greater in the faunated microcosms, but highly variable with time. Denitrification rates were positively stimulated by N. succinea populations, denitrification of water column nitrate was stimulated 10-fold in comparison to denitrification coupled to nitrification in the sediments. Rates of denitrification of water column nitrate were not significantly different from rates in undisturbed sediment cores with similar densities of N. succinea, whereas rates of coupled nitrification–denitrification were 3-fold lower in the experimental set-up. These results may reflect the relative growth rates of nitrifying and denitrifying bacteria, which allow more rapid colonisation of new burrow surfaces by denitrifier compared to nitrifier populations. The data indicate that recolonisation by burrowing macrofauna of the highly reduced sediments of the Sacca di Goro, Lagoon, Italy, following the annual dystrophic crisis, may play a significant role in the reoxidation and detoxification of the sediments. The increased rates of denitrification associated with the worm burrows, may promote nitrogen losses, but due to the low capacity of nitrifying bacteria to colonise the new burrow structures, these losses would be highly dependent upon water column nitrate concentrations.     

The effect of crude oil on the colonization of meiofauna into salt marsh sediments

The effect of an oil spill on estuarine meiofauna was examined in a controlled colonization experiment. Forty-five replicate azoic sediment chambers treated with 0,133 or 381 mg hydrocarbons: 100 g dry sediment of South Louisiana crude oil were each quantitatively sampled with three replicate cores for colonizing meiofauna. Chambers were sampled on days 2, 5, 10, 30 and 60 postplacement in a Louisiana Spartina alterniflora (Loisel) salt marsh. Polychaetes showed a delayed colonization and reduced densities in oiled relative to non-oiled sediments. Nematode numbers were significantly depressed in the high oil treatment but no delay in colonization was identified. Only one species of meiobenthic copepods, Enhydrosoma woodini, displayed a reaction to the presence of the oil but only in the top centimeter of sediment. This species showed significantly decreased densities due to the heavily oiled treatment throughout the study until day 60 when numbers in the heavily oiled chambers were significantly higher than those in the non-oiled chambers. Species diversity (H') was calculated on the meiobenthic copepod assemblage and showed that diversity in the high oil treatment was generally lower than that in other treatments through day 30. Fewer species colonized the heavily oiled chambers before day 30. Principle Components Ordination conducted on the copepod assemblage could not identify an oil effect separate from a chamber effect for copepod community structure.     

Azospirillum affects Eh and potential denitrification in a soil

The effect of inoculation withAzospirillum brasilense (strain 7001) on soil Eh under anaerobic conditions (N₂ flux) was examined during 144 h at 26°C and 230 h at 18°C. The Eh values of the control (not inoculated) soil decreased to approximately 80 or 140 mV in both cases, whereas after 24 h of anaerobic incubation, the Eh of theAzospirillum-inoculated soil remained at higher values. After 144 and 230 h of anaerobic incubation, the denitrifying activity (measured in anaerobiosis with excess of e^- acceptor and donor) in the inoculated soil was seven and three times lower respectively, than in the non-inoculated soil. This indicates thatAzospirillum may affect the soil Eh and consequently any highly Eh-dependent microbial activity, such as denitrification.     

Denitrifying Bacterial Community Composition Changes Associated with Stages of Denitrification in Oxygen Minimum Zones

Denitrification in the ocean is a major sink for fixed nitrogen in the global N budget, but the process is geographically restricted to a few oceanic regions, including three oceanic oxygen minimum zones (OMZ) and hemipelagic sediments worldwide. Here, we describe the diversity and community composition of microbes responsible for denitrification in the OMZ using polymerase chain reaction, sequence and fragment analysis of clone libraries of the signature genes (nirK and nirS) that encode the enzyme nitrite reductase, responsible for key denitrification transformation steps. We show that denitrifying assemblages vary in space and time and exhibit striking changes in diversity associated with the progression of denitrification from initial anoxia through nitrate depletion. The initial denitrifying assemblage is highly diverse, but succession on the scale of 3–12 days leads to a much less diverse assemblage and dominance by one or a few phylotypes. This progression occurs in the natural environment as well as in enclosed incubations. The emergence of dominants from a vast reservoir of rare types has implications for the maintenance of diversity of the microbial population and suggests that a small number of microbial dominants may be responsible for the greatest rates of transformations involving nitrous oxide and global fixed nitrogen loss. Denitrifying blooms, driven by a few types responding to episodic environmental changes and distributed unevenly in time and space, are consistent with the sampling effect model of diversity–function relationships. Canonical denitrification thus appears to have important parallels with both primary production and nitrogen fixation, which are typically dominated by regionally and temporally restricted blooms that account for a disproportionate share of these processes worldwide.     

Use of chitin for controlling plant-parasitic nematodes

Cabbage plants were grown in soil amended with Clandosan (CLA) prepared from crustacean chitin (0.3% w/w). The plants were maintained in constant temperature tanks set to 15° or 30°C, in soils naturally infested with cyst nematodeHeterodera schachtii, or inoculated with the root-knot nematode,Meloidogyne javanica, respectively. At 30°C, after the first month following inoculation, CLA caused an increase in top fresh weight of plants but no reduction in nematode—induced root galling was recorded. However, when fresh plants were planted, CLA induced a large reduction in gall formation and caused an increase in top fresh weight of nematode-inoculated plants. At 15°C, CLA significantly affected the plants only after 60 days: an increase in top fresh weight and a reduction in the number of eggs per cyst were recorded. Ammonium was not detected in soil after 30 days, at 30°C, whereas at 15°C, CLA-treated soil contained twice as much ammonium as non-treated soil. After 60 days, ammonium was not detected at all. After 30 days nitrate concentrations in soil attained higher values at 30°C than at 15°C, whereas after 60 days high levels were detected only at 15°C. At 30°C, CLA induced an increase in the number of fungi, chitinolytic bacteria, and total amount of bacteria; at 15°C, such an increase was detected only with the chitinolytic microorganisms.Contribution from the Agricultural Research Organization (ARO), Bet Dagan, Israel No. 2196-E, 1987 series.     

Denitrification activity of Bradyrhizobium sp. isolated from Argentine soybean cultivated soils

Two hundred and fifty strains, all of them representatives of native Bradyrhizobium sp., isolated from soils cultivated with soybean have been characterized by their denitrification activity. In addition, the denitrification potential of those soils was also measured by evaluating the most-probable-number (MPN) of denitrifying bacteria and the denitrification enzyme assay (DEA). Of the 250 isolates tested, 73 were scored as probable denitrifiers by a preliminary screening method. Only 41 were considered denitrifiers because they produced gas bubbles in Durham tubes, cultures reached an absorbance of more than 0.1 and NO³⁻ and NO²⁻ were not present. Ten of these 41 were selected to confirm denitrification and to study denitrification genes. According to N₂O production and cell protein concentration with NO³⁻, the isolates could be differentiated in three categories of denitrifiers. The presence of the napA, nirK, norC and nosZ genes was detected by production of a diagnostic PCR product using specific primers. RFLP from the 16S-23S rDNA spacer region (IGS) revealed that denitrifiers strains could be characterized as Bradyrhizobium japonicum and strains which were non-respiratory denitrifiers as B. elkanii.     

Nitrogen Turnover in Drying Sediments of an Amazon Floodplain Lake

In the Amazon floodplain large areas are subject to annual cycles of drying and rewetting. The turnover of nitrogen in the periodically drying sediments is an important regulator of floodplain fertility. In the present study the transition of a lake sediment from flooded to dry conditions was studied with respect to microbial nitrogen turnover. Soil nitrogen pools, as well as the activity and abundance of denitrifying and nitrifying bacteria, were investigated during one dry season. During the first weeks after drying, most of the inorganic nitrogen vanished from the sediment. The process was inhibited by a nitrification inhibitor, showing that coupled nitrification–denitrification was responsible for the nitrogen loss. Assimilation by plants or microbes, as well as leaching, were not important mechanisms of nitrogen loss. During a period of only 10 days, 59% of the total denitrification and 94% of the total N₂O emission during the dry period occurred. Cell numbers of denitrifiers were not correlated with activities. Denitrification was not correlated with other sediment variables but was regulated by the patchy distribution of reduced and oxidized zones in the uppermost centimeters of the sediment. This heterogeneity was probably introduced by the bioturbation of small insects, which was restricted to a rather short time period shortly after drying.     

Spatial Heterogeneity in the Distribution of Denitrifying Bacteria Associated with Denitrification Activity Zones

Denitrifying enzyme activity (DEA) in artificially established denitrification activity zones was, on average, 3.5-fold greater than in bulk soil. Denitrifier biomass was between 1 and 4 orders of magnitude greater in activity zones. Denitrifier specific activity was lower in activity zones and more similar to specific activity values reported for pure cultures of denitrifying bacteria added to soil. Measurements of DEA and biomass carried out on a homogeneous subsample derived from a soil core containing a denitrification activity zone will overestimate the DEA and biomass of the bulk soil and underestimate the DEA and biomass of the activity zone.     

Effects of photoperiod on growth of and denitrification by Pseudomonas chlororaphis in the root zone of Glyceria maxima,studied in a gnotobiotic microcosm

The emergent macrophyte Glyceria maxima was subjected to different photoperiods and grown with ammonium or nitrate as nitrogen source in presterilized microcosms with spatially separated root and non-root compartments. The microcosms were inoculated with the denitrifying bacterium Pseudomonas chlororaphis. The effect of the plant and the photoperiod on growth and denitrification by P. chlororaphis was assessed. The plant had a strong positive effect on the growth of the bacteria. The bacterial numbers in the root compartment of the planted microcosms were 19-32 times higher than found in the non-root sediment of the unplanted systems. Lengthening the photoperiod resulted in elevated bacterial numbers due to the higher carbon exudation of the plant. This effect was greater still with the nitrate-fed plants, where additional P. chlororaphis growth could proceed via denitrification, indicating oxygen-limiting conditions in the microcosms. Higher porewater N₂O concentrations in the root compartments as compared to the non-root compartments, which were highest for the long photoperiod, were also indicative of a plant-induced stimulation of denitrification. An effect of a diurnal oxygen release pattern of G. maxima on denitrification could not be detected. The gnotobiotic microcosm used in this study represents a potential system for the study of the behaviour and interactions of important bacterial groups, such as nitrifying and denitrifying bacteria where plant roots drive bacterial activity.     

Riparian zone denitrification affects nitrogen flux through a tidal freshwater river

Tidal freshwater zones (TFZ) of coastal rivers link terrestrial watersheds to the ocean and are characterized by large, regularly inundated riparian zones. We investigated the effect of riparian denitrification on nitrogen flux in the TFZ Newport River, North Carolina (U.S.A.) by developing an empirical model of denitrification and parameterizing it using measured denitrification rates, sediment oxidation-reduction potential dynamics, and riparian topography. Denitrification rates were measured monthly in laboratory-incubated sediment cores by using a membrane inlet mass spectrometer to assess net water-borne N₂ flux from the cores. Annual average rates of denitrification in three intertidal riparian habitats, emergent marsh, mudflat, and hardwood forest, were 1864, 1956, and 2018 μg m^?2 h^?1, respectively. Laboratory experiments and in-situ monitoring revealed that the temporal lag between tidal inundation and reduced, denitrifying conditions was 4–5 h. Field measurements and remotely sensed data showed that the inundated surface area during high tide was three times greater than that at low tide. By combining data on denitrification, oxidation-reduction potential, and topography, the model predicted that the daily denitrification flux constituted 2–15% of the daily riverine nitrate flux during most of the year and >100% during low discharge periods. Current regional and global nitrogen budgets thus may overestimate nitrogen delivery to the ocean by not accounting for the TFZ denitrification.     

Urban Stormwater Runoff Drives Denitrifying Community Composition Through Changes in Sediment Texture and Carbon Content

The export of nitrogen from urban catchments is a global problem, and denitrifying bacteria in stream ecosystems are critical for reducing in-stream N. However, the environmental factors that control the composition of denitrifying communities in streams are not well understood. We determined whether denitrifying community composition in sediments of nine streams on the eastern fringe of Melbourne, Australia was correlated with two measures of catchment urban impact: effective imperviousness (EI, the proportion of a catchment covered by impervious surfaces with direct connection to streams) or septic tank density (which affects stream water chemistry, particularly stream N concentrations). Denitrifying community structure was examined by comparing terminal restriction fragment length polymorphisms of nosZ genes in the sediments, as the nosZ gene codes for nitrous oxide reductase, the last step in the denitrification pathway. We also determined the chemical and physical characteristics of the streams that were best correlated with denitrifying community composition. EI was strongly correlated with community composition and sediment physical and chemical properties, while septic tank density was not. Sites with high EI were sandier, with less fine sediment and lower organic carbon content, higher sediment cations (calcium, sodium and magnesium) and water filterable reactive phosphorus concentrations. These were also the best small-scale environmental variables that explained denitrifying community composition. Among our study streams, which differed in the degree of urban stormwater impact, sediment grain size and carbon content are the most likely drivers of change in community composition. Denitrifying community composition is another in a long list of ecological indicators that suggest the profound degradation of streams is caused by urban stormwater runoff. While the relationships between denitrifying community composition and denitrification rates are yet to be unequivocally established, landscape-scale indices of environmental impact such as EI may prove to be useful indicators of change in microbial communities.