Activity Of Anammox Bacteria Research Articles

A combined upflow anaerobic sludge bed, aerobic, and anoxic fixed-bed reactor system for removing tetramethylammonium hydroxide and nitrogen from light-emitting diode wastewater

ABSTRACTA laboratory study using a combined upflow anaerobic sludge bed (UASB) and aerobic and anoxic fixed-bed reactor system was undertaken to explore its capability for removing tetramethylammonium hydroxide (TMAH) and nitrogen from light-emitting diode wastewater. When the organic loading rate was maintained at 0.26–0.65 kg TMAH m−3 d−1, the UASB reactor removed 70–100% of TMAH through methanogenesis. When the -N loading rate was maintained at 0.73–1.4 kg -N m−3 d−1, the aerobic reactor oxidized 31–59% of -N to -N through nitritation. When the nitrogen loading rate was maintained at 0.42–0.75 kg N m−3 d−1, the anoxic reactor removed 27–63% of nitrogen through anammox. The performance data of the combined reactor system agreed well with the stoichiometric relationships of methanogenesis, nitritation, and anammox. The batch studies showed that a higher initial TMAH concentration of up to 2520 mg L−1 gave a higher methanogenic activity of up to 16 mL CH4 g−1 VSS d−1. An increase in the initial TMAH concentration of up to 500 mg L−1 gradually decreased the activity of ammonia-oxidizing bacteria; whereas an increase in the initial TMAH concentration of up to 47 mg L−1 imposed a marked inhibiting effect on the activity of anammox bacteria.

Distribution and activity of anaerobic ammonium-oxidising bacteria in natural freshwater wetland soils.

Anaerobic ammonium oxidation (anammox) process plays a significant role in the marine nitrogen cycle. However, the quantitative importance of this process in nitrogen removal in wetland systems, particularly in natural freshwater wetlands, is still not determined. In the present study, we provided the evidence of the distribution and activity of anammox bacteria in a natural freshwater wetland, located in southeastern China, by using (15)N stable isotope measurements, quantitative PCR assays and 16S rRNA gene clone library analysis. The potential anammox rates measured in this wetland system ranged between 2.5 and 25.5 nmol N2 g(-1) soil day(-1), and up to 20% soil dinitrogen gas production could be attributed to the anammox process. Phylogenetic analysis of 16S rRNA genes showed that anammox bacteria related to Candidatus Brocadia, Candidatus Kuenenia, Candidatus Anammoxoglobus and two novel anammox clusters coexisted in the collected soil cores, with Candidatus Brocadia and Candidatus Kuenenia being the dominant anammox genera. Quantitative PCR of hydrazine synthase genes showed that the abundance of anammox bacteria varied from 2.3 × 10(5) to 2.2 × 10(6) copies g(-1) soil in the examined soil cores. Correlation analyses suggested that the soil ammonium concentration had significant influence on the activity of anammox bacteria. On the basis of (15)N tracing technology, it is estimated that a total loss of 31.1 g N m(-2) per year could be linked the anammox process in the examined wetland.

Role of Anaerobic Ammonium Oxidation (Anammox) in Nitrogen Removal from a Freshwater Aquifer.

Anaerobic ammonium oxidation (anammox) couples the oxidation of ammonium with the reduction of nitrite, producing N2. The presence and activity of anammox bacteria in groundwater were investigated at multiple locations in an aquifer variably affected by a large, wastewater-derived contaminant plume. Anammox bacteria were detected at all locations tested using 16S rRNA gene sequencing and quantification of hydrazine oxidoreductase (hzo) gene transcripts. Anammox and denitrification activities were quantified by in situ (15)NO2(-) tracer tests along anoxic flow paths in areas of varying ammonium, nitrate, and organic carbon abundances. Rates of denitrification and anammox were determined by quantifying changes in (28)N2, (29)N2, (30)N2, (15)NO3(-), (15)NO2(-), and (15)NH4(+) with groundwater travel time. Anammox was present and active in all areas tested, including where ammonium and dissolved organic carbon concentrations were low, but decreased in proportion to denitrification when acetate was added to increase available electron supply. Anammox contributed 39-90% of potential N2 production in this aquifer, with rates on the order of 10 nmol N2-N L(-1) day(-1). Although rates of both anammox and denitrification during the tracer tests were low, they were sufficient to reduce inorganic nitrogen concentrations substantially during the overall groundwater residence times in the aquifer. These results demonstrate that anammox activity in groundwater can rival that of denitrification and may need to be considered when assessing nitrogen mass transport and permanent loss of fixed nitrogen in aquifers.

Evidence for anaerobic ammonium oxidation process in freshwater sediments of aquaculture ponds

The anaerobic ammonium oxidation (anammox) process, which can simultaneously remove ammonium and nitrite, both toxic to aquatic animals, can be very important to the aquaculture industry. Here, the presence and activity of anammox bacteria in the sediments of four different freshwater aquaculture ponds were investigated by using Illumina-based 16S rRNA gene sequencing, quantitative PCR assays and (15)N stable isotope measurements. Different genera of anammox bacteria were detected in the examined pond sediments, including Candidatus Brocadia, Candidatus Kuenenia and Candidatus Anammoxoglobus, with Candidatus Brocadia being the dominant anammox genus. Quantitative PCR of hydrazine synthase genes showed that the abundance of anammox bacteria ranged from 5.6 × 10(4) to 2.1 × 10(5) copies g(-1) sediment in the examined ponds. The potential anammox rates ranged between 3.7 and 19.4 nmol N2 g(-1) sediment day(-1), and the potential denitrification rates varied from 107.1 to 300.3 nmol N2 g(-1) sediment day(-1). The anammox process contributed 1.2-15.3% to sediment dinitrogen gas production, while the remainder would be due to denitrification. It is estimated that a total loss of 2.1-10.9 g N m(-2) per year could be attributed to the anammox process in the examined ponds, suggesting that this process could contribute to nitrogen removal in freshwater aquaculture ponds.

Fast start-up of the anammox process with addition of reduced graphene oxides

In this study, two upflow column reactors were built up to investigate the effects of reduced graphene oxide (RGO) on the startup of anammox process. Continuous experiments indicated that the start-up period of anammox process with RGO addition could be shortened from 67 to 49days. The further investigation proved that the anammox reactor with RGO addition possessed relatively higher anammox bacteria activity and stronger stability even against the high nitrogen loading rate (NLR) impacting. After 220days cultivation, the nitrogen removal rate of R2 (RGO addition) reached 1.08×103g-N/(m3d), which was 27.4% higher than 0.846×103g-N/(m3d) of the R1 (the control). The hydrazine dehydrogenase (HDH) activity increased dramatically with RGO addition, which could be the trigger of fast anammox start-up. Compared with the control reactor, the quantity of anammox bacteria in R2 increased during the whole operation period by quantitative PCR and fluorescence in situ hybridization analysis.

Chemical speciation of heavy metals in excess sludge treatment by thermal hydrolysis and anaerobic digestion process

Adequate treatment of excess sludge from municipal wastewater treatment plants (WWTP) is a critical issue nowadays in WWTP management. Thermal hydrolysis and anaerobic digestion are the methods often used to reduce the organic content of the excess sludge. However, these two methods have very limited effects on heavy metals’ removal. As the toxicity of heavy metals is closely related to not only their quantity but also in their chemical speciations, more in-depth studies on heavy metals speciation distribution are needed. In this paper, the speciation of eight kinds of heavy metals before and after thermal hydrolysis and anaerobic digestion were investigated using a modified BCR (Bureau Communautaire de Référence: a four-step sequential extraction process proposed by the European Community Bureau of Reference in 1992) sequential extraction process. The results showed that the stable content of eight kinds of heavy metals increased by various extents after thermal hydrolysis. The chemical speciation of heavy metals during thermal hydrolysis and anaerobic digestion exhibited significant changes from non-stable to relatively stable state. The existence of H2S can reduce the activity of anammox bacteria, and thus can lead to the deterioration of anaerobic biological treatment system. However, the toxicity of H2S and heavy metal ions can be significantly inhibited simultaneously by the interaction between heavy metals and H2S without secondary pollution, which is beneficial for excess sludge disposal.

Using electric field to enhance the activity of anammox bacteria.

Electric field was applied to enhance the activity of anaerobic ammonium oxidation (anammox) bacteria for nitrogen removal in this study. The effect of different distributive modes with the total electric field time of 6 h/24 h (R2) and 18 h/24 h (R3) at 2 V/cm was determined through long-term tests. With the continuous application time increasing from 3 to 9 h in R3 (1 to 3 h in R2), the total nitrogen removal rate (NRR) showed an acceleration trend. When the electric field was applied continuously for 9 h (R3, mode 3, application-rest time: 9 h-3 h), the NRR dramatically increased 38.7 % over that of control on day 160. Besides, it was demonstrated that the increase of crude enzyme activities and the cell quantities were the main reasons for the enhancement of nitrogen removal performance of the anammox process. Additionally, transmission electron microscope observation proved the morphological change of anammox biomass under electric field application.

Statistical analysis to evaluate the effects of temperature and pH on anammox activity

Anaerobic ammonium oxidation (anammox) is the most efficient process of nitrogen removal from wastewater. A central composite design was employed to study the individual and interactive effects of temperature and pH on the activity of anammox bacteria. A total of 13 experiments including 5 center point replicates with specific anammox activity as a response were carried out. The maximum specific anammox activity (0.0286 ± 0.0003 g N2–N g VSS−1 d−1) was observed at temperature and pH of 43 °C and 7.5, respectively. The results of estimated regression coefficients for specific anammox activity suggested that both temperature and pH positively affects the specific anammox activity of bacteria. The effect of pH was found to be more significant than temperature as evident from the P value (P < 0.05). Further, contour plot revealed that the interaction between pH and temperature is also significant. Regression model equation was developed to predict the specific anammox activity at particular pH and temperature. The regression model was validated by carrying out experiment at lower temperatures (25, 20 and 15 °C) with varying pH, which revealed that an appropriate anammox activity could be maintained at lower temperature by changing pH. The study suggested that higher pH should be maintained in wastewater treatment plant to achieve the high anammox activity at lower temperatures.

Identification of the release and effects of AHLs in anammox culture for bacteria communication

Quorum sensing (QS) is a microorganism communication system. This study verified the potential existence of the QS system among ANaerobic AMMonium OXidation (anammox) bacteria. The release of three Acyl Homoserine Lactones (AHLs; C6-HSL, C8-HSL, C12-HSL) by anammox culture was confirmed. By conducting series of batch test, the introduction of exogenous C6-HSL and C8-HSL could significantly increase activity of anammox bacteria, and meanwhile, C6-HSL significant increased the growth rate, finally resulting in the increased ammonium nitrogen removal performance of 35% and 20%, respectively, whereas the exogenous C12-HSL apparently promoted the growth of heterotrophic bacteria in the community. To further identify how exogenous signal molecular affects anammox culture, analyses of anammox culture supernatant and biomass extraction revealed that these additional signal molecules activated bacteria by inducing the production of endogenous signal molecular by anammox culture. This study might open a new window to overcome the disadvantage of the long-start-up period of anammox reactor restricting the development of current anammox process.

Occurrence and importance of anaerobic ammonium-oxidising bacteria in vegetable soils.

The quantitative importance of anaerobic ammonium oxidation (anammox) has been described in paddy fields, while the presence and importance of anammox in subsurface soil from vegetable fields have not been determined yet. Here, we investigated the occurrence and activity of anammox bacteria in five different types of vegetable fields located in Jiangsu Province, China. Stable isotope experiments confirmed the anammox activity in the examined soils, with the potential rates of 2.1 and 23.2 nmol N2 g(-1) dry soil day(-1), and the anammox accounted for 5.9-20.5% of total soil dinitrogen gas production. It is estimated that a total loss of 7.1-78.2 g N m(-2) year(-1) could be linked to the anammox process in the examined vegetable fields. Phylogenetic analyses showed that multiple co-occurring anammox genera were present in the examined soils, including Candidatus Brocadia, Candidatus Kuenenia, Candidatus Anammoxoglobus and Candidatus Jettenia, and Candidatus Brocadia appeared to be the most common anammox genus. Quantitative PCR further confirmed the presence of anammox bacteria in the examined soils, with the abundance varying from 2.8 × 10(5) to 3.0 × 10(6) copies g(-1) dry soil. Correlation analyses suggested that the soil ammonium concentration had significant influence on the activity and abundance of anammox bacteria in the examined soils. The results of our study showed the presence of diverse anammox bacteria and indicated that the anammox process could serve as an important nitrogen loss pathway in vegetable fields.

Molecular and geochemical constraints on anaerobic ammonium oxidation (anammox) in a riparian zone of the Seine Estuary (France)

To expand the limited knowledge about the ecological significance of anaerobic ammonium oxidation (anammox) in continental aquatic and terrestrial ecosystems, we studied community structure, abundance, and activity of anammox bacteria in soils and sediments in the wetland of Trou Deshayes, a riparian zone in the Seine Estuary, France. Combining (i) molecular analyses of the genes coding for anammox bacterial 16S rRNA and the enzyme hydrazine oxidoreductase (hzo), (ii) quantification of unique anammox bacterial membrane lipids (i.e. ladderanes), and, (iii) 15N-isotope label incubation experiments with intertidal sediments and irregularly flooded soils nearby, we demonstrated that anammox bacteria were ubiquitous in the studied wetland ecosystem. In both soils and sediments, detected anammox bacteria were related to Candidatus ‘Brocadia’. 16S rRNA genes were generally lower in the more oxygenated soils, but on the same order of magnitude (107–108 copies g−1 d.w.) as found for other river estuaries, riparian zones and agricultural soils. While the C20-ladderane fatty acid with five cyclobutane moieties (C20-[5]-FA) was found in both sediments and soils, other ladderane species were detected only in the wetland sediments. The observed differential ladderane distribution suggests intra-genus differences in the community composition of anammox bacteria between the sediments and the floodplain soils. While the abundance of anammox bacteria was significantly lower in the soils versus the sediments, the potential anammox rates were similar (≤15 and ≤22 nmol N2 d−1 g−1 w.w. sediment and soil, respectively), suggesting lower cell-specific anammox rates in the sediments. The observed potential rates of anammox were rather low, leaving canonical denitrification as the main fixed N removal pathway in this riparian zone. The relative contribution of anammox to the total N2 production (between 3 and 8 %) was similar at all sites, highlighting the dependence of the anammox process on nitrite supply from denitrification across environmental boundaries. Due to this coupling, the dependence of organotrophic denitrification on the quality and stoichiometry of OM also seems to affect the anammox bacterial community. Our results suggest that N removal and mitigation of N supply from agriculture in wetlands by anammox is limited, and much less important than denitrification.

Observability of anammox activity in single-stage nitritation/anammox reactors using mass balances

Theoretically, mass balances based on microbial kinetics allow the determination of the activity of anammox bacteria (AMX) and heterotrophic bacteria (HET). In practise, the variance of the resulting activities is too high.

Intermittent aeration in one-stage partial nitritation/anammox process

This study investigated the influence of different aeration strategies, characterized by the ratio (R) between non-aerated and aerated phase duration and dissolved oxygen concentration (DO), on one stage partial nitritation/anammox process performance and efficiency treating reject water. The activity of anammox bacteria, ammonium oxidizers, nitrite oxidizers and denitrifiers was determined by measuring specific anammox activity (SAA), oxygen uptake rate (OUR) and nitrate utilization rate (NUR). The results showed that with the same DO level during continuous aeration and intermittent aeration R=1/3 (in one hour), nitrogen removal efficiencies were similar. This means that introduction of intermittent aeration with R=1/3 can shorten the duration of aeration without any loss of process efficiency. Around 17–20% of the total decrease in ammonium and nitrate occurred in the non-aerated phase when intermittent aeration with R=1/3 was applied. Moreover, introduction of non-aerated phases enhanced the activity of anammox bacteria and limited that of nitrite oxidizers.

Effect of zero-valent iron on the start-up performance of anaerobic ammonium oxidation (anammox) process.

The long start-up time of anaerobic ammonium oxidation (anammox) process hinders the widespread application of anammox technology in practical wastewater treatment when anammox seed sludge is not available. Meanwhile, the production of nitrate cannot meet the increasingly more strict discharge standards. To combine the chemical nitrate reduction to ammonium with biological nitrogen removal, two anammox upflow anaerobic sludge blanket reactors packed with different types of zero-valent iron (ZVI), microscale ZVI (mZVI) and nanoscale ZVI (nZVI), were developed to accelerate the start-up of anammox process. The results revealed that anammox start-up time shortened from 126 to 105 and 84 days with the addition of mZVI and nZVI. The nitrogen removal performance was also improved remarkably by adding ZVI, especially in the start-up stage. The value of dissolved oxygen showed that ZVI could be regarded as a useful deoxidant to create anaerobic condition for the proliferation of anammox bacteria. ZVI was favorable for the secretion of EPS, which would represent the activity of anammox bacteria. The result of real-time quantitative PCR (qPCR) further confirmed that the proliferation of anammox bacteria was enhanced by ZVI.

Microbial composition of the activated sludge of Moscow wastewater treatment plants

The contribution of the major technologically important microbial groups (ammonium- and nitrite-oxidizing, phosphate-accumulating, foam-inducing, and anammox bacteria, as well as planctomycetes and methanogenic archaea) was characterized for the aeration tanks of the Moscow wastewater treatment facilities. FISH investigation revealed that aerobic sludges were eubacterial communities; the metabolically active archaea contributed insignificantly. Stage II nitrifying microorganisms and planctomycetes were significant constituents of the bacterial component of activated sludge, with Nitrobacter spp. being the dominant nitrifier. No metabolically active anammox bacteria were revealed in the sludge from aeration tanks. The sludge from the aeration tanks using different wastewater treatment technologies were found to differ in characteristics. Abundance of the nitrifying and phosphate-accumulating bacteria in the sludges generally correlated with microbial activity, in microcosms and with efficiency of nitrogen and phosphorus removal from wastewater. The highest microbial numbers and activity were found in the sludges of the tanks operating according to the technologies developed in the universities of Hanover and Cape Town. The activated sludge from the Novokur yanovo facilities, where abundant growth of filamentous bacteria resulted in foam formation, exhibited the lowest activity The group of foaming bacteria included Gordonia spp. and Acinetobacter spp., utilizing petroleum and motor oils, Sphaerotilus spp. utilizing unsaturated fatty acids, and Candidatus 'Microthrix parvicella'. Thus, the data on abundance and composition of metabolically active microorganisms obtained by FISH may be used for the technological control of wastewater treatment.

Start-up of low-temperature anammox in UASB from mesophilic yeast factory anaerobic tank inoculum

Robust start-up of the anaerobic ammonium oxidation (anammox) process from non-anammox-specific seeding material was achieved by using an inoculation with sludge-treating industrial -, organics- and N-rich yeast factory wastewater. N-rich reject water was treated at 20°C, which is significantly lower than optimum treatment temperature. Increasing the frequency of biomass fluidization (from 1–2 times per day to 4–5 times per day) through feeding the reactor with higher flow rate resulted in an improved total nitrogen removal rate (from 100 to 500 g m−3d−1) and increased anammox bacteria activity. As a result of polymerase chain reaction (PCR) tests, uncultured planctomycetes clone 07260064(4)-2-M13-_A01 (GenBank: JX852965) was identified from the biomass taken from the reactor. The presence of anammox bacteria after cultivation in the reactor was confirmed by quantitative PCR (qPCR); an increase in quantity up to ∼2×106 copies g VSS−1 during operation could be seen in qPCR. Statistical modelling of chemical parameters revealed the roles of several optimized parameters needed for a stable process.

The effects of zero-valent iron (ZVI) and ferroferric oxide (Fe3O4) on anammox activity and granulation in anaerobic continuously stirred tank reactors (CSTR)

The anammox granules with Fe and Fe3O4 as cores were successfully obtained in anaerobic continuously stirred tank reactors (CSTR) in this study. The sizes [d(0.5)] of Fe-anammox and Fe3O4-anammox granules successively arrived to 417.3μm and 988.5μm on the 39th and 72nd day, corresponding to the specific anammox activity (SAA) of 0.24 and 0.239kgNkgVSS−1d−1, respectively. The SAA of blank-anammox granule was stable (about 0.20kgNkgVSS−1d−1) in the whole study. The anammox granulation mechanisms of common anammox granule (blank-anammox granule) and Fe-anammox/Fe3O4-anammox granule were extracellular polymers (ECP) bonding model and ion-bonding model, respectively. Results showed that the optimal oxidation–reduction potential (ORP) of anammox bacteria was 209mV. The reasonable concentration of Fe2+ and Fe3+ was beneficial to the granulation and activity of anammox bacteria by maintaining high ORP and gathering acylhomoserine lactones (AHLs). The quorum sensing (QS) material of anammox bacteria was identified as N-dodecanoyl homoserine lactone by LC–MS.

Research of iron reduction and the iron reductase localization of anammox bacteria.

The iron-reducing capability of anammox bacteria was examined in this study using Percoll purified anammox bacteria. Anammox bacteria could reduce Fe(III) to Fe(II) with organic matters as the electron donor. The activity of anammox iron-reducing process was dependent on different electron donor, acceptor and pH. The highest iron-reducing activity of anammox bacteria was achieved with Fe(III)-NTA (nitrilotriacetic acid) as electron acceptor and formate as the electron donor at pH7. Similar to other iron reducers, 80 % of the iron reductase in anammox bacteria was located in the membrane fraction. Due to the chemical oxidant of NO2 (-) and the NO3 (-) dependent ferrous iron oxidation by anammox bacteria, the iron-reducing activity of anammox bacteria could be severely inhibited when iron-reducing pathway and the anammox process were coupled. However, the total nitrogen removal efficiency was not significantly affected in the presence of Fe(III). The iron-reducing capability of anammox bacteria could influence both N and Fe cycle on earth, and it is a potential way for wastewater treatment.

Reason Analysis of ANAMMOX Occurred in a Landfill Leachate Treatment System

The biological treatment process of landfill leachate in Beijing Liulitun landfill is a multistage A/O technology, in which a large amount of ANAMMOX (Anaerobic ammonia oxidation, ANAMMOX) bacteria were found in the sludge. There are several factors impacting the activity of ANAMMOX bacteria, including pH value, temperature and HRT which in this process are suitable for the survival of ANAMMOX bacteria. Especially, low dissolved oxygen is an essential factor as the provider of electron donor for nitrite formation. Although the high concentrations of organic matter, ammonia nitrogen and nitrite will inhibit the occurrence of ANAMMOX, ANAMMOX bacteria can self-detoxification by forming a low-poison habitat by consuming ammonia and nitrite as well as organic matter by heterotrophic ANAMMOX bacteria.

The role of pH on the resistance of resting- and active anammox bacteria to NO2- inhibition.

The anaerobic oxidation of ammonium (anammox) uses nitrite as terminal electron acceptor. The nitrite can cause inhibition to the bacteria that catalyze the anammox reaction. The literature shows a great divergence on the levels of NO2 (-) causing inhibition. Moreover, the conditions influencing the resistance of anammox bacteria to NO2 (-) inhibitory effect are not well understood. This work investigated the effect of the pH and the concentration of nitrite on the activity and metabolism of anammox granular sludge under different physiological conditions. Batch activity tests in a range of pH values were carried out in which either actively metabolizing cells or resting cells were exposed to nitrite in the presence or absence of the electron donating substrate ammonium, respectively. The response of the bacteria was evaluated by analyzing the specific anammox activity, the accumulation of nitric oxide, and the evolution of the ATP content in the biomass. Additionally, the effect of the pH on the tolerance of the biomass to single substrate feeding interruptions was evaluated in continuous anammox bioreactors. The results show that the influence of the pH on the NO2 (-) inhibition of anammox bacteria is greater under non-metabolizing conditions than during active metabolism. The exposure of resting cells to NO2 (-) (100 mg N L(-1) ) at pH values below 7.2 caused complete inhibition of the anammox activity. The inhibition was accompanied by accumulation of the intermediate, nitric oxide, in the gas phase. In contrast, just mild inhibition was observed for resting cells exposed to the same NO2 (-) concentration at pH values higher than 7.5 or any of the pH values tested in assays with actively metabolizing cells. ATP initially increased and subsequently decreased in time after resting cells were exposed to NO2 (-) suggesting an active response of the cells to nitrite stress. Furthermore, bioreactors operated at pH lower than 6.8 had greater sensitivity to NO2 (-) during an ammonium feed interruption than a bioreactor operated at pH 7.1. The results suggest that the ability of resting cells to tolerate NO2 (-) inhibition is seriously impeded at mildly acidic pH values; whereas actively metabolizing biomass is resistant to NO2 (-) toxicity over a wide range of pH values.