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PubMed2005

Culture-independent techniques for rapid detection of bacteria associated with loss of chloramine residual in a drinking water system.

Chloramination is often the disinfection regimen of choice for extended drinking water systems. However, this process is prone to instability due to the growth of nitrifying bacteria. This is the first study to use alternative approaches for rapid investigation of chloraminated drinking water system instability in which flow cytometric cell sorting of bacteria with intact membranes (membrane-intact fraction) (BacLight kit) or with active esterases (esterase-active fraction) (carboxyfluorescein diacetate) was combined with 16S rRNA gene-directed PCR and denaturing gradient gel electrophoresis (DGGE). No active bacteria were detected when water left the water treatment plant (WTP), but 12 km downstream the chloramine residual had diminished and the level of active bacteria in the bulk water had increased to more than 1 x 10(5) bacteria ml(-1). The bacterial diversity in the system was represented by six major DGGE bands for the membrane-intact fraction and 10 major DGGE bands for the esterase-active fraction. PCR targeting of the 16S rRNA gene of chemolithotrophic ammonia-oxidizing bacteria (AOB) and subsequent DGGE and DNA sequence analysis revealed the presence of an active Nitrosospira-related species and Nitrosomonas cryotolerans in the system, but no AOB were detected in the associated WTP. The abundance of active AOB was then determined by quantitative real-time PCR (qPCR) targeting the amoA gene; 3.43 x 10(3) active AOB ml(-1) were detected in the membrane-intact fraction, and 1.40 x 10(4) active AOB ml(-1) were detected in the esterase-active fraction. These values were several orders of magnitude greater than the 2.5 AOB ml(-1) detected using a routine liquid most-probable-number assay. Culture-independent techniques described here, in combination with existing chemical indicators, should allow the water industry to obtain more comprehensive data with which to make informed decisions regarding remedial action that may be required either prior to or during an instability event.

Bacteria↗
PubMed1994

Oxidation of hydroxylamine by cytochrome P-460 of the obligate methylotroph Methylococcus capsulatus Bath.

An enzyme capable of the oxidation of hydroxylamine to nitrite was isolated from the obligate methylotroph Methylococcus capsulatus Bath. The absorption spectra in cell extracts, electron paramagnetic resonance spectra, molecular weight, covalent attachment of heme group to polypeptide, and enzymatic activities suggest that the enzyme is similar to cytochrome P-460, a novel iron-containing protein previously observed only in Nitrosomonas europaea. The native and subunit molecular masses of the M. capsulatus Bath protein were 38,900 and 16,390 Da, respectively; the isoelectric point was 6.98. The enzyme has approximately one iron and one copper atom per subunit. The electron paramagnetic resonance spectrum of the protein showed evidence for a high-spin ferric heme. In contrast to the enzyme from N. europaea, a 13-nm blue shift in the soret band of the ferrocytochrome (463 nm in cell extracts to 450 nm in the final sample) occurred during purification. The amino acid composition and N-terminal amino acid sequence of the enzyme from M. capsulatus Bath was similar but not identical to those of cytochrome P-460 of N. europaea. In cell extracts, the identity of the biological electron acceptor is as yet unestablished. Cytochrome c-555 is able to accept electrons from cytochrome P-460, although the purified enzyme required phenazine methosulfate for maximum hydroxylamine oxidation activity (specific activity, 366 mol of O2 per s per mol of enzyme). Hydroxylamine oxidation rates were stimulated approximately 2-fold by 1 mM cyanide and 1.5-fold by 0.1 mM 8-hydroxyquinoline.

Amino Acid Sequence↗
PubMed1997

Metal selectivity of in situ microcolonies in biofilms of the Elbe river.

The ultrastructure of natural complex biofilm communities of the Elbe river grown in situ on microscopic glass coverslips was studied by using transmission electron microscopy and energy-dispersive x-ray (EDX) analysis. Characteristic microcolonies which measured between 3.3 and 9.3 microm in diameter were frequently observed. They had an outer envelope and harbored 6 to 30 cells. The cells formed short rods measuring 1.09 +/- 0.28 microm (n = 10) in length and 0.55 + 0.07 microm (n = 21) in width. They were surrounded by a thick layer of electron-transparent, nonosmicated matter, 120 to 300 nm thick. Individual cells exhibited a unique ultrastructural trait, namely, a concentric membrane stack which completely surrounded the cytoplasm. It consisted of three membrane doublets, which showed an overall thickness of 57 to 66 nm. The center-to-center spacing between two membrane doublets was 22.2 +/- 1.0 nm (n = 12). The bacterial cell wall seemed to be of the gram-negative type. The fact that upon shrinkage hexagonal clefts appeared proved the cells to be tightly packed, and septum formation by binary fissions was observed. All of these morphological details indicate that the cells within these microcolonies were actively growing and did not represent spore-like states. EDX analysis showed that only the electron-dense surface deposit of the microcolonies contained Mn and Fe in significant amounts, while these two elements were absent from the intercellular space and the cytoplasm of the microorganisms. In contrast, aluminum ions were able to penetrate the outer envelope of the microcolonies and were detected in the intercellular space. They were, however, completely absent from the microbial cytoplasm, indicating a filter cascade with respect to aluminum. From the ultrastructural data together with the deposition of iron and manganese on the microcolony surface, it appears that these organisms may belong to the genus Siderocapsa or Nitrosomonas. They do not precisely match any of the described species and may therefore represent a new species.

Biofilms↗
PubMed1998

Cytochrome P460 genes from the methanotroph Methylococcus capsulatus bath.

P460 cytochromes catalyze the oxidation of hydroxylamine to nitrite. They have been isolated from the ammonia-oxidizing bacterium Nitrosomonas europaea (R. H. Erickson and A. B. Hooper, Biochim. Biophys. Acta 275:231-244, 1972) and the methane-oxidizing bacterium Methylococcus capsulatus Bath (J. A. Zahn et al., J. Bacteriol. 176:5879-5887, 1994). A degenerate oligonucleotide probe was synthesized based on the N-terminal amino acid sequence of cytochrome P460 and used to identify a DNA fragment from M. capsulatus Bath that contains cyp, the gene encoding cytochrome P460. cyp is part of a gene cluster that contains three open reading frames (ORFs), the first predicted to encode a 59,000-Da membrane-bound polypeptide, the second predicted to encode a 12, 000-Da periplasmic protein, and the third (cyp) encoding cytochrome P460. The products of the first two ORFs have no apparent similarity to any proteins in the GenBank database. The overall sequence similarity of the P460 cytochromes from M. capsulatus Bath and N. europaea was low (24.3% of residues identical), although short regions of conserved residues are present in the two proteins. Both cytochromes have a C-terminal, c-heme binding motif (CXXCH) and a conserved lysine residue (K61) that may provide an additional covalent cross-link to the heme (D. M. Arciero and A. B. Hooper, FEBS Lett. 410:457-460, 1997). Gene probing using cyp indicated that a cytochrome P460 similar to that from M. capsulatus Bath may be present in the type II methanotrophs Methylosinus trichosporium OB3b and Methylocystis parvus OBBP but not in the type I methanotrophs Methylobacter marinus A45, Methylomicrobium albus BG8, and Methylomonas sp. strains MN and MM2. Immunoblot analysis with antibodies against cytochrome P460 from M. capsulatus Bath indicated that the expression level of cytochrome P460 was not affected either by expression of the two different methane monooxygenases or by addition of ammonia to the culture medium.

Amino Acid Sequence↗
PubMed2002

Phylogeny and functional expression of ribulose 1,5-bisphosphate carboxylase/oxygenase from the autotrophic ammonia-oxidizing bacterium Nitrosospira sp. isolate 40KI.

The autotrophic ammonia-oxidizing bacteria (AOB), which play an important role in the global nitrogen cycle, assimilate CO(2) by using ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO). Here we describe the first detailed study of RubisCO (cbb) genes and proteins from the AOB. The cbbLS genes from Nitrosospira sp. isolate 40KI were cloned and sequenced. Partial sequences of the RubisCO large subunit (CbbL) from 13 other AOB belonging to the beta and gamma subgroups of the class Proteobacteria are also presented. All except one of the beta-subgroup AOB possessed a red-like type I RubisCO with high sequence similarity to the Ralstonia eutropha enzyme. All of these new red-like RubisCOs had a unique six-amino-acid insert in CbbL. Two of the AOB, Nitrosococcus halophilus Nc4 and Nitrosomonas europaea Nm50, had a green-like RubisCO. With one exception, the phylogeny of the AOB CbbL was very similar to that of the 16S rRNA gene. The presence of a green-like RubisCO in N. europaea was surprising, as all of the other beta-subgroup AOB had red-like RubisCOs. The green-like enzyme of N. europaea Nm50 was probably acquired by horizontal gene transfer. Functional expression of Nitrosospira sp. isolate 40KI RubisCO in the chemoautotrophic host R. eutropha was demonstrated. Use of an expression vector harboring the R. eutropha cbb control region allowed regulated expression of Nitrosospira sp. isolate 40KI RubisCO in an R. eutropha cbb deletion strain. The Nitrosospira RubisCO supported autotrophic growth of R. eutropha with a doubling time of 4.6 h. This expression system may allow further functional analysis of AOB cbb genes.

Amino Acid Sequence↗
PubMed2006

The yjeB (nsrR) gene of Escherichia coli encodes a nitric oxide-sensitive transcriptional regulator.

Microarray studies of the Escherichia coli response to nitric oxide and nitrosative stress have suggested that additional transcriptional regulators of this response remain to be characterized. We identify here the product of the yjeB gene as a negative regulator of the transcription of the ytfE, hmpA and ygbA genes, all of which are known to be upregulated by nitrosative stress. Transcriptional fusions to the promoters of these genes were expressed constitutively in a yjeB mutant, indicating that all three are targets for repression by YjeB. An inverted repeat sequence that overlaps the -10 element of all three promoters is proposed to be a binding site for the YjeB protein. A similar inverted repeat sequence was identified in the tehA promoter, which is also known to be sensitive to nitrosative stress. The ytfE, hmpA, ygbA, and tehA promoters all caused derepression of a ytfE-lacZ transcriptional fusion when present in the cell in multiple copies, presumably by a repressor titration effect, suggesting the presence of functional YjeB binding sites in these promoters. However, YjeB regulation of tehA was weak, as judged by the activity of a tehA-lacZ fusion, perhaps because YjeB repression of tehA is masked by other regulatory mechanisms. Promoters regulated by YjeB could be derepressed by iron limitation, which is consistent with an iron requirement for YjeB activity. The YjeB protein is a member of the Rrf2 family of transcriptional repressors and shares three conserved cysteine residues with its closest relatives. We propose a regulatory model in which the YjeB repressor is directly sensitive to nitrosative stress. On the basis of similarity to the nitrite-responsive repressor NsrR from Nitrosomonas europaea, we propose that the yjeB gene of E. coli be renamed nsrR.

Escherichia coli↗
PubMed2005

Bacterial diversity in water samples from uranium wastes as demonstrated by 16S rDNA and ribosomal intergenic spacer amplification retrievals.

Bacterial diversity was assessed in water samples collected from several uranium mining wastes in Ger many and in the United States by using 16S rDNA and ribosomal intergenic spacer amplification retrievals. The results obtained using the 16S rDNA retrieval showed that the samples collected from the uranium mill tailings of Schlema/Alberoda, Germany, were predominated by Nitrospina-like bacteria, whereas those from the mill tailings of Shiprock, New Mexico, USA, were predominated by gamma-Pseudomonas and Frauteria spp. Additional smaller populations of the Cytophaga-Flavobacterium-Bacteroides group and alpha- and delta-Proteobacteria were identified in the Shiprock samples as well. Proteobacteria and Cytophaga-Flavobacterium-Bacteroides were also found in the third uranium mill tailings studied, Gittersee/Coschütz, Germany, but the groups of the predominant clones were rather small. Most of the clones of the Gittersee/Coschütz samples represented individual sequences, which indicates a high level of bacterial diversity. The samples from the fourth uranium waste studied, Steinsee Deponie B1, Germany, were predominantly occupied by Acinetobacter spp. The ribosomal intergenic spacer amplification retrieval provided results complementary to those obtained by the 16S rDNA analyses. For instance, in the Shiprock samples, an additional predominant bacterial group was identified and affiliated with Nitrosomonas sp., whereas in the Gittersee/Coschütz samples, anammox populations were identified that were not retrieved by the applied 16S rDNA approach.

Bacteria↗
PubMed2005

Dissimilatory metabolism of nitrogen oxides in bacteria: comparative reconstruction of transcriptional networks.

Bacterial response to nitric oxide (NO) is of major importance since NO is an obligatory intermediate of the nitrogen cycle. Transcriptional regulation of the dissimilatory nitric oxides metabolism in bacteria is diverse and involves FNR-like transcription factors HcpR, DNR, and NnrR; two-component systems NarXL and NarQP; NO-responsive activator NorR; and nitrite-sensitive repressor NsrR. Using comparative genomics approaches, we predict DNA-binding motifs for these transcriptional factors and describe corresponding regulons in available bacterial genomes. Within the FNR family of regulators, we observed a correlation of two specificity-determining amino acids and contacting bases in corresponding DNA recognition motif. Highly conserved regulon HcpR for the hybrid cluster protein and some other redox enzymes is present in diverse anaerobic bacteria, including Clostridia, Thermotogales, and delta-proteobacteria. NnrR and DNR control denitrification in alpha- and beta-proteobacteria, respectively. Sigma-54-dependent NorR regulon found in some gamma- and beta-proteobacteria contains various enzymes involved in the NO detoxification. Repressor NsrR, which was previously known to control only nitrite reductase operon in Nitrosomonas spp., appears to be the master regulator of the nitric oxides' metabolism, not only in most gamma- and beta-proteobacteria (including well-studied species such as Escherichia coli), but also in gram-positive Bacillus and Streptomyces species. Positional analysis and comparison of regulatory regions of NO detoxification genes allows us to propose the candidate NsrR-binding motif. The most conserved member of the predicted NsrR regulon is the NO-detoxifying flavohemoglobin Hmp. In enterobacteria, the regulon also includes two nitrite-responsive loci, nipAB (hcp-hcr) and nipC (dnrN), thus confirming the identity of the effector, i.e. nitrite. The proposed NsrR regulons in Neisseria and some other species are extended to include denitrification genes. As the result, we demonstrate considerable interconnection between various nitrogen-oxides-responsive regulatory systems for the denitrification and NO detoxification genes and evolutionary plasticity of this transcriptional network.

Amino Acid Motifs↗
PubMedDate not supplied

DNA-probing for genes coding for denitrification, N2-fixation and nitrification in bacteria isolated from different soils.

Bacteria isolated from different layers of four soils of the Cologne area were analyzed for denitrifying, nitrifying and N2-fixing isolates by colony hybridization using gene probes. In the soils tested, the percentage of denitrifying bacteria among the total population isolated was 3-8% (in one case exceptionally 15%) and thus small. Denitrifying bacteria were particularly enriched in the upper layer (depth approximately 5 cm) and were present only in low amounts at 25 cm depth in two gleysol soils. Nitrate apparently did not determine the distribution of denitrifying bacteria in these soils. The potential denitrification activity of different soil layers coincided with the distribution pattern of isolates assessed by DNA-probing. The total number of bacteria and of denitrifying isolates was considerably higher in or at the roots of plants than in the bulk, root-free soil adjacent to the plants. The percentage of the isolated aerobic N2-fixing bacteria varied between 0-3%, and these bacteria could be isolated mainly from the upper 5 cm layer. A small portion of the isolates hybridized with the probe coding for part of one subunit of ammonia monooxygenase from Nitrosomonas europaea. The investigation showed that DNA-probing can provide useful information about the relative distribution of denitrifying and N2-fixing bacteria in different soils and their layers.

Bacteria↗
PubMed2006

Quantification of ammonia-oxidizing bacteria populations in full-scale sewage activated sludge systems and assessment of system variables affecting their performance.

This study carried out quantification of ammonia-oxidizing bacteria (AOB) populations in 12 full-scale sewage activated sludge systems that were different in ammonia removals and treatment processes during three different seasons. Experiment was divided into 3 parts: 1) analysis of AOB communities by PCR-DGGE-cloning-sequencing of 16S rRNA genes; 2) development of four real-time PCR primer sets for quantification of the particular AOB of interest; and 3) quantification of AOB populations by using the newly developed real-time PCR primer sets. The results suggested that all the primer sets gave good reproducibility and specificity for PCR amplification with the detection limits of 10(2) copies/PCR reaction. Although the 12 systems were different in several aspects, one of the identified sequence types of Nitrosomonas oligotropha cluster was the dominant AOB in every system and every season studied. However, the other sequence type of this cluster was not significantly involved in ammonia removals in the systems. The occurrence of N. communis cluster in the systems seemed to depend on the remaining oxygen concentrations in the sludge floc and thus the activity of aerobic heterotrophs in the aeration tanks. N. europaea-Nitrosococcus. mobilis solely existed in one A20 system of which the influent contained twice the chloride concentrations than those of other systems.

Ammonia↗
PubMed2006

Post-treatment of a slaughterhouse wastewater: stability of the microbial community of a sequencing batch reactor operated under oxygen limited conditions.

Slaughterhouse wastewater is a complex effluent with an important content of organic nitrogen. After an anaerobic treatment where most of the organic matter is removed, the nitrogen, remains as ammonium and post-treatment of the effluent is necessary. Sequencing batch reactor (SBR) technology has been developed to completely remove nitrogen in one single reactor combining aerobic and anoxic stages. Under oxygen limited conditions only nitrite is produced with concomitant energy saving. The stability and diversity of the microbial community from a nitrifying denitrifying SBR operated under oxygen limited conditions were studied using molecular and respirometric methods. The AOB (ammonia oxidizing bacteria) community was relatively stable Nitrosomonas being the dominant genera although Nitrosospira and Nitrosococcus were detected in low proportions. Nitrite oxidizing bacteria were out competed during the operation under oxygen-limited conditions. After an increase of the DO in the reactor Nitrobacter spp were detected suggesting that they remained in the system. Changes in the AOB and denitrifying communities were observed after the DO increase. Sedimentation problems were detected during operation, this could be related to the predominance of Thauera spp detected by FISH and T-RFLP.

Abattoirs↗
PubMedDate not supplied

Quantification of nitrifying bacterial populations in a full-scale nitrifying trickling filter using fluorescent in situ hybridization.

Fluorescent in situ hybridization (FISH) was used to quantify the ammonia-oxidizing populations within intact biofilm samples collected from a full-scale nitrifying trickling filter (NTF). Ammonia, nitrite, and nitrate concentrations were measured for aqueous samples taken in conjunction with biofilm samples at multiple filter depths. Correlation coefficients for individual sampling events, calculated by simple linear regression of FISH signal area and ammonia removal rates, ranged from 0.558 when using probe NEU23a to 0.982 when using probe Nso190. The improved correlations with Nso190 suggest that genera other than Nitrosomonas are present in this system. Percent biofilm coverage, as determined by 4',6-diamidino-2-phenylindole counterstaining and dry weight biomass measurements, did not change throughout the NTF. This indicates that biofilm growth is fairly uniform throughout the filter even if nitrifier growth is not.

Ammonia↗
PubMedDate not supplied

Linking nitrifying biofilm structure and function through fluorescent in situ hybridization and evaluation of nitrification capacity.

Fluorescent in situ hybridization (FISH) was used to quantify the ammonia-oxidizing populations within intact biofilm samples collected from a full-scale nitrifying trickling filter (NTF). Ammonia, nitrite, and nitrate concentrations were measured for aqueous samples taken in conjunction with biofilm samples at multiple filter depths. The ammonia removal capability of collected biofilms was evaluated by transferring the biofilms to laboratory batch reactors. Nitrate plus nitrite generation rates were calculated for the full-scale NTF and for the bench-scale reactors. Correlation coefficients for individual sampling events, calculated by simple linear regression of FISH signal area and nitrate plus nitrite generation rates for the full-scale NTF, ranged from -0.882 to 0.984. Correlation coefficients obtained for FISH signal area and nitrate plus nitrite generation rates for the bench-scale reactors ranged from 0.853 to 0.994 when using probe Nso190 and from 0.919 to 0.986 when using probe Neu23a. Occasional differences between the number of ammonia-oxidizing bacteria quantified by the probe Nso190 versus the NEU23a probe with depth suggest that genera other than Nitrosomonas are present in the NTF.

Ammonia↗
PubMedDate not supplied

Calcium carbonate phosphate binding ion exchange filtration and accelerated denitrification improve public health standards and combat eutrophication in aquatic ecosystems.

Cultural eutrophication, the process by which a lake becomes rich in dissolved nutrients as a result of point and nonpoint pollutant sources, is a major cause of the loss of natural lake ecosystems throughout the world. The process occurs naturally in all lakes, but phosphate-rich nutrient runoff from sources such as storm drains and agricultural runoff is a major cause of excess phosphate-induced eutrophication. Especially in Madrona Marsh, one of the last remaining vernal marshes in the greater Los Angeles area, California, cultural eutrophication has become a major problem. In this study, calcium carbonate was found to be an excellent phosphate binder, reducing up to 70% of the phosphates in a given sample of water, and it posed relatively negligent ecological repercussions. This study involved the testing of this principle in both the laboratory and the real ecosystem. A calcium carbonate lacing procedure was first carried out to determine its efficacy in Madrona Marsh. Through this, ammonia was found to interfere with the solubility of calcium carbonate and therefore to be a hindrance to the reduction of phosphate. Therefore, various approaches for reduction of ammonia were tested, including aeration, use of fiber growth media, and plants, mainly Caulerpa verticellata, chosen for it hardiness, primarily in an attempt to increase population of Nitrobacter and Nitrosomonas. All were successful in moderately reducing ammonia levels. In addition, soil sampling, sediment analysis, microscopic plant analysis, microorganism and macroinvertebrate identification, and rate law formulations were conducted. The effect of phosphate and ammonia reduction on the populations of enterobacteria was also an important focus of this experiment. Varying concentrations of phosphate, ammonia, and calcium carbonate in conjunction with phosphate were tested in Madrona Marsh to determine their effects on the populations of enteropathogens on nonspecific blood agar, MacConkey agar, and Hektoen agar. Initial analyses suggest a strong correlation between phosphate concentrations and bacterial populations; a 66% decrease in phosphate resulted in a 35% reduction in bacterial populations and a 45% reduction in enteropathogenic populations. Likewise, a strong correlation was shown between calcium carbonate concentrations and bacterial reduction greater than that which can be attributed to the phosphate reduction alone. This was followed by the construction of various phosphate binding calcium carbonate filters, which used the ion exchange principle, including a spring loading filter, PVC pipe filter, and a galvanized filter. All were tested with the aid of Stoke's law formulation. The experiment was extremely successful in designing a working phosphate-binding and ammonia-reducing filter, and a large-scale agitator-clarifier filter system is currently being planned for construction in Madrona Marsh; this filter will reduce phosphate and ammonia levels substantially in the following years, bringing ecological, economical, and health-related improvements to the overall ecosystem and habitat.

Calcium Carbonate↗
PubMed1999

High Mortality in Zebrafish (Danio rerio).

A group of 100 adult zebrafish were housed in a new system at a stocking density of 20 fish per tank. Four weeks after arrival, 15 fish presented with petechial hemorrhages and ulceration on the surfaces of the skin. Samples of the fish were collected for histopathology, fungal culture, and bacterial culture and sensitivity. Water samples were analyzed for pH, ammonia, nitrite, and submitted for bacterial and fungal culture. Histologically, the epidermis had multiple areas of ulceration and mononuclear cell infiltrate. Gram-positive bacteria were observed beneath the surface of the skin and surrounding the outer aspect of the spinal cord. Both Aeromonas hydrophila and A. sobria were isolated from the affected fish, and a diagnosis of motile aeromonad septicemia (MAS) was made. Water from the tanks had a nitrite level of 1-5 ppm, a toxic concentration that indicated poor water quality. Because the housing system had been seeded with Nitrobacter spp. and Nitrosomonas spp. only 2 weeks prior to the arrival of the fish, a lack of colonizing nitrifying bacteria was deemed to be the cause of the high nitrite level, which, along with over-crowding, stressed the fish and increased their susceptibility to MAS. No further cases of septicemia were observed once the nitrite level and stocking density were reduced.

Journal Article↗
PubMed2002

Oxygen-limited nitrogen removal in a lab-scale rotating biological contactor treating an ammonium-rich wastewater.

A lab-scale Rotating Biological Contactor (RBC) was operated with the purpose of oxygen-limited (autotrophic) nitrification-denitrification of an ammonium-rich synthetic wastewater without Chemical Oxygen Demand (COD). Based on the field observations that RBCs receiving anaerobic effluents come to anoxic ammonium removal, the RBC was inoculated with methanogenic sludge. Some 100 days after the addition of the anaerobic sludge to the reactor as a possible means of a rapid initiation of the nitrogen (N) removal process, a maximum ammonium removal of 1,550 mg N m(-2) d(-1) was achieved. Batch tests with 15N labeled ammonium and nitrite indicated that a large part of that N was removed via oxygen-limited oxidation of ammonium with nitrite as the electron acceptor. The other part was removed via conventional denitrification, presumably with COD released from lysis of cells. Species identification of the most abundant microorganisms revealed that Nitrosomonas spp. were the dominant ammonium-oxidizers in the sludge. Thus far, the molecular characterization of the sludge could not show the presence of Planctomycetes among the most dominant species. Overall this experiment confirms the property of the RBC system to remove ammonium to nitrogen gas without the use of heterotrophic carbon source.

Bacteria, Anaerobic↗
PubMed2002

Production of gaseous nitrogen compounds in a novel process for ammonium removal.

The production of gaseous nitrogen compounds, particularly the greenhouse gas nitrous oxide, was investigated in a novel process for ammonium removal from wastewater. The process is based on the adsorption of ammonium on zeolite followed by bioregeneration. The zeolite serves the dual purpose of an ion exchanger and a physical carrier for nitrifying bacteria which bio-regenerate the ammonium saturated mineral. An analysis of the nitrifying population composition in the reactor fed with simulated secondary effluent (NH4+ = 50 mg/l) revealed that about half of the bacteria in the biofilm were common ammonium oxidizers Nitrosococcus mobilis and Nitrosomonas, while the other half were nitrite oxidizers. The amount of nitrogen losses, under different conditions, and the identification of the emitted gases (N2 or N2O) were investigated in two sets of experiments: (I) batch experiments using biomass originating from the ion exchange reactor with and without the addition of nitrite, and (II) continuous experiments using the ion exchange reactor with zeolite as the biomass carrier. In the batch experiments, nitrite and oxygen concentrations were determined as the major parameters responsible for the formation of gaseous nitrogen gas during ammonia oxidation by autotrophic bacteria. Continuous experiments showed that the major parameter significantly affecting nitrogen losses was the amount of ammonium adsorbed by the zeolite during the ion exchange phase. The amount of ammonium adsorbed determines the ammonium concentration during the initial period of bioregeneration, which in turn directly influences oxygen demand and the resulting concentrations of oxygen and nitrite. It was concluded that the formation of nitrogen gas compounds in the ion exchange/bioregeneration process can be eliminated by adjusting the operational regime to have a shorter adsorption phase resulting in smaller amounts of ammonium adsorbed per cycle.

Adsorption↗