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Archaeal nitrification in the ocean.

Marine Crenarchaeota are the most abundant single group of prokaryotes in the ocean, but their physiology and role in marine biogeochemical cycles are unknown. Recently, a member of this clade was isolated from a sea aquarium and shown to be capable of nitrification, tentatively suggesting that Crenarchaeota may play a role in the oceanic nitrogen cycle. We enriched a crenarchaeote from North Sea water and showed that its abundance, and not that of bacteria, correlates with ammonium oxidation to nitrite. A time series study in the North Sea revealed that the abundance of the gene encoding for the archaeal ammonia monooxygenase alfa subunit (amoA) is correlated with a decline in ammonium concentrations and with the abundance of Crenarchaeota. Remarkably, the archaeal amoA abundance was 1-2 orders of magnitude higher than those of bacterial nitrifiers, which are commonly thought to mediate the oxidation of ammonium to nitrite in marine environments. Analysis of Atlantic waters of the upper 1,000 m, where most of the ammonium regeneration and oxidation takes place, showed that crenarchaeotal amoA copy numbers are also 1-3 orders of magnitude higher than those of bacterial amoA. Our data thus suggest a major role for Archaea in oceanic nitrification.

Archaeal Proteins↗

Use of natural zeolite to enhance nitrification in biofilter.

To enhance nitrification, natural zeolite and activated carbon were tested as a media in a biofilter to treat wastewater containing relatively high concentrations of TKN. Using those media, the adsorption isotherms for ammonium ion were compared; the K values for natural zeolite and activated carbon were found to be 0.5117 and 0.0006, respectively. In comparison of the performance of the two media, two identical, lab-scale biofilters were then operated for 4 months. The effect of NH3-N loading rates on the performance was investigated. The results showed that higher NH3-N removal efficiency and faster nitrification were achieved in the biofilter with natural zeolite throughout the experimental period. Nitrosomonas and Nitrobacter, two principal nitrifiers, in biofilm grown on two different media were counted and compared. Nitrobacter which is the more fragile of the two principal nitrifiers was outgrown in the biofilm on natural zeolite media. The reason for this may be due to the ammonium ion exchanging capacity of natural zeolite which provided the favorable environment for Nitrobacter.

Biodegradation, Environmental↗

Effect of temperature and dissolved oxygen on biological nitrification at high ammonia concentrations.

Effect of temperature and dissolved oxygen concentration on nitrification rate were investigated with enrichment cultures of nitrifying bacteria. Values of specific nitrite oxidation rate in the absence of ammonia were 2.9-12 times higher than maximum specific ammonia oxidation rates at the same temperatures. The presence of high ammonia levels reversed this relationship, causing maximum specific nitrite oxidation rates to fall to 19 to 45% as high as maximum specific ammonia oxidation rates. This result suggests that nitrification at high ammonia levels will invariably result in nitrite accumulation. The K(O2) for nitrite oxidation in the presence of high ammonia levels was higher than the K(O2) for ammonia oxidation when temperature exceeded 18 degrees C, whereas the opposite was true at lower temperatures. These results indicate that low oxygen tensions will exacerbate nitrite accumulation when water temperature is high.

Ammonia↗

Oxygen-limited nitrification and denitrification in a lab-scale rotating biological contactor.

Rotating Biological Contactors (RBC) treating highly nitrogenous wastewaters are known to give rise to nitrogen losses, generally assumed to be due to concomitant nitrification and denitrification. In this study, a lab-scale nitrifying RBC reactor was shown to bring about losses of nitrogen of the order of 10 to 20% at ammonium loading rates of up to 2129 mg N m(-2) d(-1), when no extra carbon source was added. These higher removal values could be due to Oxygen-Limited Autotrophic Nitrification and Denitrification (OLAND), because the reactor was operated at oxygen limitation. When methanol was added as a soluble organic carbon source to further diminish oxygen transfer into the deeper parts of the biofilm, the RBC achieved 84% nitrogen removal (loading rate 2300 mg N m(-2) d(-1)) at a COD/N ratio of 3.1. The latter also suggests that the process was occurring parallel to conventional denitrification.

Bioreactors↗

Nitrification activities in full-scale treatment plants with varying salt loads.

The effect of salt on the nitrification activity in full-scale wastewater treatment plants (WWTP) was investigated. Not only the activity of ammonia and nitrite oxidisers was measured, but also the nitrifying population was assessed (by Fluorescent In Situ Hybridisation)--in full-scale domestic and industrial WWTPs, operated under various salt levels. The results demonstrate a decline in the activity of ammonia and nitrite oxidisers with an increase in salt content: the domestic WWTP with the lowest salt level (0.13g Cl- l(-1)) had the highest specific activity of ammonia and nitrite oxidisers (4.3 and 2.4 mg N (gVSS)(-1) h(-1), respectively), while the lowest specific activities of ammonia and nitrite oxidisers (1.1 and 0.5 mg N (gVSS)(-1) h(-1)) were measured at the highest NaCl concentration (16g Cl- l(-1)). However, comparing the nitrification activity of different types of sludge developed under different operational conditions with the reported values was not directly possible. So we have used the activated sludge model (ASM) to translate the routine operational data into parameters to enable the calculation of the actual fraction of nitrifiers and consequently the actual specific activity of ammonia and nitrite oxidisers. Expressing the activity of ammonia oxidisers in terms of actual specific activity makes the results from pure cultures, enriched cultures, pilot scale and full scale WWTPs comparable. Moreover, these results confirm the behaviour of nitrifiers under salt stress and validate the results obtained from pure and enriched cultures to be extrapolated to full scale.

Ammonia↗

Factors affecting nitrification in situ in a heated stream.

Nitrification, previously shown to occur in the stream under study, ensued only when the stream received heated (28 degree C or above) discharges and was negligible when water temperatures were 17 degree C or lower. The most probable number os ammonium oxidizer in stream bed sediments exceeded intrite oxidizers by ratios of 43:1 to 1,113:1, and the average rate of ammonium oxidation in this stream, 0.50 mg of NH4-N/liter per h exceeded the rate of nitrite oxidation, 0.29 mg of NO2-N/liter per h, resulting in an accumulation of nitrite. Nitrification rates were influenced by the dissolved oxygen concentration, increasing during daylight hours and exhibiting maximum rates during the summer months.

Ammonia↗

Nitrification in the intertidal zone: influence of effluent type and effect of tannin on nitrifiers.

Nitrification by intertidal sediments was measured by using a tide simulator that approximated the cycle of seawater on tidal flats. Sediments were chosen from sites affected by industrial and municipal effluents and pastoral seepage and runoff. The ability of sediments from different sites to nitrify endogenous nitrogen varied markedly. All sites exhibited an initial lag before activity commenced. The duration of this lag and the rate of nitrate production were different at each site. The sediments were also capable of oxidizing NH3-N supplied to them in seawater. This "nitrification potential" was highest at sites receiving nitrogenous effluents (slaughterhouse and sewage), but was also substantial in sediments affected by bark extract effluent and pasture runoff. The lowest potential and the longest lag were exhibited by sediments in an apple cannery effluent area. Enrichment cultures of nitrifying microorganisms were obtained from all sites using NH4+ as a source of energy, but enrichments for nitrite oxidizers were unsuccessful. Concentrated pine bark tannins, similar in origin to those in effluents at the well-nitrifying chipmill site, were tested for toxicity to pure cultures of nitrifying bacteria. Two Nitrobacter strains and one Nitrosomonas strain were unaffected by tannins even at 5 mg/ml. A Nitrosolobus and a Nitrosospira strain were inhibited partially at 5 mg/ml and only slightly or not at all at 1 mg/ml.

Ammonia↗

Heterotrophic nitrification in an Acid forest soil and by an Acid-tolerant fungus.

Nitrate was formed from ammonium at pH 3.2 to 6.1 in suspensions of a naturally acid forest soil; the maximum rates of formation occurred at ca. pH 4 to 5. Nitrate was also formed from soil nitrogen in suspensions incubated at 50 degrees C. Autotrophic nitrifying bacteria could not be isolated from this soil. Enrichment cultures produced nitrate in a medium with beta-alanine if much soil was added to the medium, and nitrite but not nitrate was formed in the presence of small amounts of soil. Nitrification by these enrichments was abolished by eucaryotic but not procaryotic inhibitors. A strain of Absidia cylindrospora isolated from this soil was found to produce nitrate and nitrite in a medium with beta-alanine at pH values ranging from 4.0 to 4.8. Nitrate production by A. cylindrospora required the presence of sterile soil. Free and bound hydroxylamine, hydroxamic acids, and primary aliphatic nitro compounds did not accumulate during the conversion of beta-alanine to nitrite by the fungus. The organism also formed nitrite from ammonium in a medium containing acetate. We suggest that nitrification in this soil is a heterotrophic process catalyzed by acid-tolerant fungi and not by autotrophs or heterotrophs in nonacid microsites.

Journal Article↗

Environmental Parameters Regulating Gaseous Nitrogen Losses from Two Forested Ecosystems via Nitrification and Denitrification.

Gaseous N losses from disturbed and reference forested watersheds at the Coweeta Hydrologic Laboratory in western North Carolina were studied by in situ N(2)O diffusion measurements and laboratory incubations throughout a 10-month period. Soil temperature, percent base saturation, and water-filled pore space accounted for 43% of the variation in in situ N(2)O diffusion measurements. Laboratory incubations distinguished the gaseous N products of nitrification and denitrification. Nitrifying activity, ambient NO(3), and nitrification N(2)O were positively correlated with percent base saturation. However, differences between watersheds in soil N substrate caused by presence of leguminous black locust in the disturbed watershed were confounded with differences in soil acidity. Denitrification was most strongly affected by soil moisture, which in turn was determined by precipitation events and slope position. Gaseous N losses from well-drained midslope and toeslope landscape positions appeared to be minor relative to other N transformations. Favorable conditions for denitrification occurred at a poorly drained site near the stream of the disturbed watershed. Laboratory incubations revealed high rates of NO(3) reduction in these soils. We speculate that the riparian zone is a major site of depletion of NO(3) from the soil solution via denitrification.

Journal Article↗

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.

Aerobiosis↗

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.

Journal Article↗

Distinguishing nitrous oxide production from nitrification and denitrification on the basis of isotopomer abundances.

The intramolecular distribution of nitrogen isotopes in N2O is an emerging tool for defining the relative importance of microbial sources of this greenhouse gas. The application of intramolecular isotopic distributions to evaluate the origins of N2O, however, requires a foundation in laboratory experiments in which individual production pathways can be isolated. Here we evaluate the site preferences of N2O produced during hydroxylamine oxidation by ammonia oxidizers and by a methanotroph, ammonia oxidation by a nitrifier, nitrite reduction during nitrifier denitrification, and nitrate and nitrite reduction by denitrifiers. The site preferences produced during hydroxylamine oxidation were 33.5 +/- 1.2 per thousand, 32.5 +/- 0.6 per thousand, and 35.6 +/- 1.4 per thousand for Nitrosomonas europaea, Nitrosospira multiformis, and Methylosinus trichosporium, respectively, indicating similar site preferences for methane and ammonia oxidizers. The site preference of N2O from ammonia oxidation by N. europaea (31.4 +/- 4.2 per thousand) was similar to that produced during hydroxylamine oxidation (33.5 +/- 1.2 per thousand) and distinct from that produced during nitrifier denitrification by N. multiformis (0.1 +/- 1.7 per thousand), indicating that isotopomers differentiate between nitrification and nitrifier denitrification. The site preferences of N2O produced during nitrite reduction by the denitrifiers Pseudomonas chlororaphis and Pseudomonas aureofaciens (-0.6 +/- 1.9 per thousand and -0.5 +/- 1.9 per thousand, respectively) were similar to those during nitrate reduction (-0.5 +/- 1.9 per thousand and -0.5 +/- 0.6 per thousand, respectively), indicating no influence of either substrate on site preference. Site preferences of approximately 33 per thousand and approximately 0 per thousand are characteristic of nitrification and denitrification, respectively, and provide a basis to quantitatively apportion N2O.

Ammonia↗

Effect of chemicals used as nitrification inhibitors on the denitrification process.

Several chemicals used as nitrification inhibitors were tested to determine their effect on dentrification by a Pseudomonas sp. and in soil. Denitrification by the bacterium was suppressed by 2-chloro-6(-trichloromethyl)-pyridine (N-Serve) at a concentration of 50 ppm, while 2,5-dichloroaniline caused the accumulation of nitrite in the culture medium. The nitrification inhibitors had little effect on the denitrifying activity in soil under anaerobic conditions. 2-Sulfanilamidothiazole inhibited denitrification to some extent and samples supplied with potassium azide produced N2O rather than N2 as the predominant gas.

Anaerobiosis↗

Influence of nutrients, hexadecane, and temporal variations on nitrification and exopolysaccharide composition of river biofilms.

Biofilms were cultivated on polycarbonate strips in rotating annular reactors using South Saskatchewan River water during the fall of 1999 and the fall of 2001. The reactors were supplemented with carbon (glucose), nitrogen (NH(4)Cl), phosphorus (KH(2)PO(4)), or combined nutrients (CNP), with or without hexadecane. The impact of these treatments on nitrification and on the exopolysaccharide composition of river biofilms was determined. The results showed that the biofilms had higher NH4(+) oxidation, NO3(-) production, and N2O production activities in fall 1999 than fall 2001 when grown with CNP but had higher activities in fall 2001 than fall 1999 when grown with individual nutrients. The exopolysaccharide amounts and proportions were generally higher in fall 1999 than fall 2001, as a consequence of the higher nutrient levels in the river water in the first year of this study. The addition of P and especially CNP stimulated NH4(+) oxidation by the biofilms, showing a P limitation in this river ecosystem. The presence of hexadecane negatively affected these activities and lowered the amounts of exopolysaccharides in CNP and P biofilms in fall 1999 but increased the biofilm activities and exopolysaccharide amounts in CNP biofilm in fall 2001. Antagonistic, synergistic, and independent effects between nutrients and hexadecane were also observed. This study demonstrated that the biofilm autotrophic nitrification activity in the South Saskatchewan River was limited by P, that this activity and the exopolysaccharide amounts and proportions were dependent on the nutrient concentrations in the river water, and suggested that exopolysaccharides may play a protective role for biofilm microorganisms against toxic pollutants.

Alkanes↗

Nitrification and denitrification processes for biologic treatment of industrial effluents.

Nitrification process performance was evaluated using a three-phase fluidized-bed bioreactor. A synthetic effluent was used for this experiment containing 180-230 mg/L of chemical oxygen demand (COD), 25-30 mg/L of N-NH4+, 12 to 13 mg/L of total phosphorous, and micronutrients. The bioreactor used for denitrification behaved as completely mixed. The results indicate that the nitrification process was efficient, reaching efficiencies of about 98%. The best results related to the efficiency of the denitrification process were obtained when the processes were supplemented with the carbon source. The results indicated an efficiency of 86-98% COD removal.

Bacteria↗

Influence of closed loop control on microbial diversity in a nitrification process.

This paper compares two control strategies for a nitrification process. The objective is to achieve partial nitrification and thus to accumulate nitrite instead of nitrate. To this end, change in temperature setpoint and active control of oxygen and ammonia concentrations are evaluated in the long term. Evaluation is made on the control performances that are obtained, but also--and more importantly--on the microbial diversity. In particular, it is shown that the combined oxygen and ammonia control strategy is more appropriate since shift in the temperature setpoint strongly affects the composition of the microbial ecosystem present in the reactor whereas active control of oxygen and ammonia does not.

Ammonia↗

Comparison of membrane biofouling in nitrification and denitrification for the membrane bioreactor (MBR).

Batch filtration tests were conducted to compare the characteristics of membrane biofouling with regard to nitrification and denitrification. A Modified Fouling Index (MFI) was obtained using a stirred cell tester. The denitrification assays showed higher membrane fouling rates than the nitrification assays. The fouling became worse, not only due to pore blocking resistance, but also from cake layer resistance after denitrification. The Extracellular Polymeric Substances (EPS) concentration and relative hydrophobicity were decreased after denitrification, resulting in floc deterioration. The floc deterioration was assumed to have increased the cake layer resistance in the filtration test. The protein Soluble Microbial Products (SMP) concentration, portion of high molecular weight in carbohydrate SMP and relative hydrophobicity were increased after denitrification, which was assumed to cause membrane pore blocking. The changes in the EPS and SMP characteristics were the main fouling parameters in denitrification.

Biomass↗

The expanded bed biofilter: combined nitrification, solids destruction, and removal of bacteria.

Developed for tertiary nitrification, this biofilter also removed carbonaceous BOD (cBOD) and (SS). Because the biofilter is expanded, it cannot clog, and therefore does not require backflushing; yet, it removed a significant proportion of the influent SS. This unanticipated capability was due to the activities of heterotrophic bacteria, protozoa, and metazoa (nematode and oligochaete worms). The expanded bed is an intensified process, which is based on natural immobilization of microbes to small support particles. Using glassy coke as the support material, an attached layer of microbes develops, forming particulate biofilms having a superficial surface area of 1 800 m2 m(-3)(expandedbed). Autotrophic nitritifiers (Nitrosomonas spp.) were detected in the biofilm using rRNA-based molecular methods and were likely responsible, at least in part, for reducing the ammonia concentration by up to 99% (to 0.1 mg L(-1)), while the other organisms reduced cBOD and SS by up to 56% and 62%, respectively. Furthermore, the influent concentrations of Escherichia coli, coliform and heterotrophic bacteria were reduced by over 80%. It thereby provides a single process solution for combined tertiary nitrification and solids removal. Operating the process to consistently achieve < 0.5mg NH3N L(-1) and at the same time removing a significant fraction of cBOD and SS, it can replace processes such as SAFs or NTFs followed by a sandfilter.

Ammonia↗