Impact of protozoan grazing on nitrification and the ammonia- and nitrite-oxidizing bacterial communities in activated sludge

In activated sludge, protozoa feed on free-swimming bacteria and suspended particles, inducing flocculation and increasing the turnover rate of nutrients. In this study, the effect of protozoan grazing on nitrification rates under various conditions in municipal activated sludge batch reactors was examined, as was the spatial distribution of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) within the activated sludge. The reactors were monitored for ammonia, nitrite, nitrate, and total nitrogen concentrations, and bacterial numbers in the presence and absence of cycloheximide (a protozoan inhibitor), allylthiourea (an inhibitor of ammonia oxidation), and EDTA (a deflocculating agent). The accumulations of nitrate, nitrite, and ammonia were lower in batches without than with protozoa grazing. Inhibition of ammonia oxidation also decreased the amount of nitrite and nitrate accumulation. Inhibiting protozoan grazing along with ammonia oxidation further decreased the amounts of nitrite and nitrate accumulated. Induction of deflocculation led to high nitrate accumulation, indicating high levels of nitrification; this effect was lessened in the absence of protozoan grazing. Using fluorescent in situ hybridization and confocal laser scanning microscopy, AOB and NOB were found clustered within the floc, and inhibiting the protozoa, inhibiting ammonia oxidation, or inducing flocculation did not appear to lower the number of AOB and NOB present or affect their position within the floc. These results suggest that the AOB and NOB are present but less active in the absence of protozoa.     

Distinctive microbial ecology and biokinetics of autotrophic ammonia and nitrite oxidation in a partial nitrification bioreactor

Biological nitrogen removal (BNR) based on partial nitrification and denitrification via nitrite is a cost-effective alternate to conventional nitrification and denitrification (via nitrate). The goal of this study was to investigate the microbial ecology, biokinetics, and stability of partial nitrification. Stable long-term partial nitrification resulting in 82.1 +/- 17.2% ammonia oxidation, primarily to nitrite (77.3 +/- 19.5% of the ammonia oxidized) was achieved in a lab-scale bioreactor by operation at a pH, dissolved oxygen and solids retention time of 7.5 +/- 0.1, 1.54 +/- 0.87 mg O(2)/L, and 3.0 days, respectively. Bioreactor ammonia oxidizing bacteria (AOB) and nitrite oxidizing bacteria (NOB) populations were most closely related to Nitrosomonas europaea and Nitrobacter spp., respectively. The AOB population fraction varied in the range 61 +/- 45% and was much higher than the NOB fraction, 0.71 +/- 1.1%. Using direct measures of bacterial concentrations in conjunction with independent activity measures and mass balances, the maximum specific growth rate (micro(max)), specific decay (b) and observed biomass yield coefficients (Y(obs)) for AOB were 1.08 +/- 1.03 day(-1), 0.32 +/- 0.34 day(-1), and 0.15 +/- 0.06 mg biomass COD/mg N oxidized, respectively. Corresponding micro(max), b, and Y(obs) values for NOB were 2.6 +/- 2.05 day(-1), 1.7 +/- 1.9 day(-1), and 0.04 +/- 0.02 mg biomass COD/mg N oxidized, respectively. The results of this study demonstrate that the highly selective partial nitrification operating conditions enriched for a narrow diversity of rapidly growing AOB and NOB populations unlike conventional BNR reactors, which host a broader diversity of nitrifying bacteria. Further, direct measures of microbial abundance enabled not only elucidation of mixed community microbial ecology but also estimation of key engineering parameters describing bioreactor systems supporting these communities.     

Nitrite-oxidizing bacteria guild ecology associated with nitrification failure in a continuous-flow reactor

Nitrification is an important process for nitrogen removal in many wastewater treatment plants, which requires the mutualistic oxidation of ammonia to nitrate by ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB). However, this process can be quite unpredictable because both guilds are conditionally sensitive to small changes in operating conditions. Here, dynamics are examined within the NOB guild in two parallel chemostats operated at low and high dilution rates (0.10 and 0.83 day(-1), respectively) during periods of varying nitrification performance. NOB and AOB guild abundances and nitrogen-oxidation efficiency were relatively constant over time in the 0.10 day(-1) reactor; however, the 0.83 day(-1) reactor had two major disturbance episodes that caused destabilization of the NOB guild, which ultimately led to nitrification failure. The first episode caused the extinction of Nitrospira spp. from the system, resulting in chronic incomplete ammonia oxidation and nitrite accumulation. The second episode caused complete loss of nitrification activity, likely resulting from metal toxicity and the previous extinction of Nitrospira spp. from the system. These results exemplify the types of changes that can occur within the NOB guild that result in process impairment or failure, and provide one possible explanation for why nitrification is often unstable at higher dilution rates.     

Effects of aeration cycles on nitrifying bacterial populations and nitrogen removal in intermittently aerated reactors

The effects of the lengths of aeration and nonaeration periods on nitrogen removal and the nitrifying bacterial community structure were assessed in intermittently aerated (IA) reactors treating digested swine wastewater. Five IA reactors were operated in parallel with different aeration-to-nonaeration time ratios (ANA). Populations of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) were monitored using 16S rRNA slot blot hybridizations. AOB species diversity was assessed using amoA gene denaturant gradient gel electrophoresis. Nitrosomonas and Nitrosococcus mobilis were the dominant AOB and Nitrospira spp. were the dominant NOB in all reactors, although Nitrosospira and Nitrobacter were also detected at lower levels. Reactors operated with the shortest aeration time (30 min) showed the highest Nitrosospira rRNA levels, and reactors operated with the longest anoxic periods (3 and 4 h) showed the lowest levels of Nitrobacter, compared to the other reactors. Nitrosomonas sp. strain Nm107 was detected in all reactors, regardless of the reactor's performance. Close relatives of Nitrosomonas europaea, Nitrosomonas sp. strain ENI-11, and Nitrosospira multiformis were occasionally detected in all reactors. Biomass fractions of AOB and effluent ammonia concentrations were not significantly different among the reactors. NOB were more sensitive than AOB to long nonaeration periods, as nitrite accumulation and lower total NOB rRNA levels were observed for an ANA of 1 h:4 h. The reactor with the longest nonaeration time of 4 h performed partial nitrification, followed by denitrification via nitrite, whereas the other reactors removed nitrogen through traditional nitrification and denitrification via nitrate. Superior ammonia removal efficiencies were not associated with levels of specific AOB species or with higher AOB species diversity.     

泉州西湖沉积物中硝化细菌的分布及其作用

比较研究泉州西湖沉积物中氨氧化细菌(AOB)和亚硝酸盐氧化细菌(NOB)的分布及氨氧化潜力和亚硝酸盐(NO2?)氧化潜力。结果表明: 西湖沉积物中存在高浓度的有机质(OM)、总氮(TN)和氨氮。AOB生物量为1.1×106?6.4×106 个/g干土, 显著高于NOB生物量4.2×105?7.4×105 个/g 干土(配对t-检验, P<0.05)。对于NOB, 硝化杆菌属(Nitrobacter)和硝化螺菌属(Nitrospira)同时存在于西湖沉积物中, 以Nitrobacter为优势种群。AOB和NOB生物量的差异一定程度上导致西湖沉积物中氨氧化潜力显著高于NO2?氧化潜力(配对t-检验, P<0.05), NO2?氧化过程成为硝化作用的限制步骤。另外, 西湖沉积物中存在的较高浓度氨氮, 一方面促进了AOB的生长和活性, 导致较高速率的氨氧化过程, 另一方面却对亚硝酸盐氧化过程产生选择性抑制, 这也是导致NO2?氧化潜力较低的主要原因之一。     

Effect of temperature and free ammonia on nitrification and nitrite accumulation in landfill leachate and analysis of its nitrifying bacterial community by FISH

The cause of seasonal failure of a nitrifying municipal landfill leachate treatment plant utilizing a fixed biofilm was investigated by wastewater analyses and batch respirometric tests at every treatment stage. Nitrification of the leachate treatment plant was severely affected by the seasonal temperature variation. High free ammonia (NH3-N) inhibited not only nitrite oxidizing bacteria (NOB) but also ammonia oxidizing bacteria (AOB). In addition, high pH also increased free ammonia concentration to inhibit nitrifying activity especially when the NH4-N level was high. The effects of temperature and free ammonia of landfill leachate on nitrification and nitrite accumulation were investigated with a semi-pilot scale biofilm airlift reactor. Nitrification rate of landfill leachate increased with temperature when free ammonia in the reactor was below the inhibition level for nitrifiers. Leachate was completely nitrified up to a load of 1.5 kg NH4-N m(-3)d(-1) at 28 degrees C. The activity of NOB was inhibited by NH3-N resulting in accumulation of nitrite. NOB activity decreased more than 50% at 0.7 mg NH3-N L(-1). Fluorescence in situ hybridization (FISH) was carried out to analyze the population of AOB and NOB in the nitrite accumulating nitrifying biofilm. NOB were located close to AOB by forming small clusters. A significant fraction of AOB identified by probe Nso1225 specifically also hybridized with the Nitrosomonas specific probe Nsm156. The main NOB were Nitrobacter and Nitrospira which were present in almost equal amounts in the biofilm as identified by simultaneous hybridization with Nitrobacter specific probe Nit3 and Nitrospira specific probe Ntspa662.     

Effective and robust partial nitrification to nitrite by real-time aeration duration control in an SBR treating domestic wastewater

Achieving sustainable partial nitrification to nitrite has been proven difficult in treating low strength nitrogenous wastewater. Real-time aeration duration control was used to achieve efficient partial nitrification to nitrite in a sequencing batch reactor (SBR) to treat low strength domestic wastewater. Above 90% nitrite accumulation ratio was maintained for long-term operation at normal condition, or even lower water temperature in winter. Partial nitrification established by controlling aeration duration showed good performance and robustness even though encountering long-term extended aeration and starvation period. Process control enhanced the successful accumulation of ammonia oxidizing bacteria (AOB) and washout of nitrite oxidizing bacteria (NOB). Scanning electron microscope observations indicated that the microbial morphology showed a shift towards small rod-shaped clusters. Fluorescence in situ hybridization (FISH) results demonstrated AOB were the dominant nitrifying bacteria, up to 8.3 ± 1.1% of the total bacteria; on the contrary, the density of NOB decreased to be negligible after 135 days operation since adopting process control.     

Partial nitrification in a sequencing batch reactor treating acrylic fiber wastewater

A sequencing batch reactor was employed to treat the acrylic fiber wastewater. The dissolved oxygen and mixed liquor suspended solids were 2–3 and 3,500–4,000 mg/L, respectively. The results showed ammonium oxidizing bacteria (AOB) had superior growth rate at high temperature than nitrite oxidizing bacteria (NOB). Partial nitrification could be obtained with the temperature of 28 °C. When the pH value was 8.5, the nitrite-N accumulation efficiency was 82 %. The combined inhibitions of high pH and free ammonium to NOB devoted to the nitrite-N buildup. Hydraulic retention time (HRT) was a key factor in partial nitrification control, and the optimal HRT was 20 h for nitrite-N buildup in acrylic fiber wastewater treatment. The ammonium oxidation was almost complete and the transformation from nitrite to nitrate could be avoided. AOB and NOB accounted for 2.9 and 4.7 %, respectively, corresponding to the pH of 7.0. When the pH was 8.5, they were 6.7 and 0.9 %, respectively. AOB dominated nitrifying bacteria, and NOB was actually washed out from the system.     

从典型硝化细菌到全程氨氧化微生物：发现及研究进展

生物硝化过程在全球氮循环中起关键性作用,被认为由氨氮氧化成亚硝酸盐和亚硝酸盐氧化成硝酸盐两个步骤组成,分别由氨氧化微生物(Ammonia oxidizing microorganisms,AOM)和硝化细菌(Nitrite oxidizing bacteria,NOB)催化完成。AOM包括氨氧化细菌(Ammonia oxidizing bacteria,AOB)和氨氧化古菌(Ammonia oxidizing archaea,AOA),AOB与AOA分布广泛,两者的相对丰度和氨氮浓度密切相关。2015年底,3个硝化螺菌属(Nitrospira)谱系Ⅱ的NOB被证实含有AOM的特征功能酶,包括氨单加氧酶(AMO)和羟胺脱氢酶(HAO),并证明NOB同时具有氨氧化和亚硝酸盐氧化的能力,命名为全程氨氧化微生物(Complete ammonia oxidizer,Comammox)。根据AMO的α亚基基因amoA的相似性将Comammox分为两大分支clade A和clade B。它们广泛分布于自然环境和人工系统,包括土壤(稻田、森林)、淡水(湿地、河流、湖泊沉积物、蓄水层)、污水处理厂和自来水厂等。本文综述了Comammox的发现及其最新的研究进展,并展望了Comammox作为氮循环关键功能菌群的研究方向和应用前景。     

Activated sludge under free ammonia treatment using gel immobilization technology for long-term partial nitrification with different initial biomass

To achieve stable partial nitrification, activated sludge from a wastewater treatment plant using free ammonia (FA) inhibition was immobilized in a polyvinyl alcohol carrier. After FA treatment at 16.44 mg L⁻¹ for 1 day, due to the increased growth rate gap between ammonium-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB), AOB enrichment and NOB inhibition were achieved within 12 days, with AOB and NOB accounting for 65.61 and 0.05%, respectively. Subsequently, with dissolved oxygen concentrations of 4−5 mg L⁻¹, pH of 7.6–7.8 and temperature of 25 ± 1 °C, the immobilized carrier made of activated sludge achieved more than 90% and more than 86% of nitrite accumulation rate at the influent ammonia concentration of 90−110 mg L⁻¹ and 35−50 mg L⁻¹, respectively. After 50 days operation, the NOB content was 0.10%, indicating the immobilized carrier provided favorable conditions for maintaining the low NOB content. Furthermore, due to the low NOB content in the inoculum and the oxygen-limited environment formed by the increase in the AOB numbers in the carrier, immobilized carrier with different initial biomass (1, 2.5 and 5%) can achieve stable partial nitrification.     

短程硝化启动运行中功能菌群变化研究

【目的】短程硝化-厌氧氨氧化是可实现的最短生物脱氮工艺,短程硝化是实现该工艺的重要环节和必要条件。【方法】采用序批式反应器(SBR)来实现短程硝化过程的启动和稳定运行,并对该过程中的相关功能菌群变化进行检测分析。【结果】通过控制低DO浓度(<1 mg/L)和逐步提高氨氮进水负荷,可抑制氨氧化细菌(NOB)菌群增殖并促进亚硝酸氧化菌(AOB)菌群规模显著扩大,实现短程硝化过程的启动和稳定运行。在氨氮进水负荷为0.055 kg/(m3.d)时,平均氨氮去除容积负荷和污泥负荷可达到0.043kg/(m3.d)和0.16 kg/(kg.d),平均亚硝酸盐积累率可达到83.4%。在短程硝化启动和稳定运行过程中,NOB菌群密度从2.0×105CFU/mL降至1.5×104CFU/mL,相对丰度从5.51%降至2.14%;AOB菌群密度从4.5×104CFU/mL增加至1.5×107CFU/mL,相对丰度从0.18%增加至7.25%。【结论】AOB菌群规模的扩大是实现短程硝化和氨氮去除能力提高的主要原因,同时较高的进水氨氮浓度和负荷也会造成亚硝化活性的抑制。     

硝化作用研究的新发现:单步硝化作用与全程氨氧化微生物

硝化作用是氨被微生物氧化为硝酸盐的过程,分别由氨氧化微生物(AOB和AOA)和亚硝酸盐氧化细菌(NOB)主导完成.一个世纪以来,我们把这个分步硝化过程当成唯一的硝化途径来学习和研究.虽然根据动力学理论推测,环境中应该存在单步硝化作用,即由一种微生物单独完成整个硝化过程,将NH₃氧化为NO₃^-,但一直没有研究能直接证明该种微生物的存在.直到2015年底,3个科研团队分别在不同环境中发现了3种不同的经过纯培养的细菌(Candidatus Nitrospira nitrosa、Candidatus Nitrospira nitrificans和Candidatus Nitrospira inopinata)和一种未经过纯培养的细菌(类Nitrospira),它们都具备单独将氨氧化为硝酸盐的能力,这些微生物被定义为全程氨氧化微生物(Comammox).单步硝化作用和全程氨氧化微生物的发现终结了传承百年的理论,并引发了众多关于全球氮素循环的重要科学问题,如这些微生物在环境中的生态位点及其在硝化作用中的相对贡献等.本文就单步硝化作用及全程氨氧化微生物的发现作了简要概述.     

Nitrification in hybrid bioreactors treating simulated domestic wastewater

Aim

To provide deeper insights into nitrification process within aerobic bioreactors containing supplemental physical support media (hybrid bioreactors).

Methods and Results

Three bench‐scale hybrid bioreactors with different media size and one control bioreactor were operated to assess how biofilm integrity influences microbial community conditions and bioreactor performance. The systems were operated initially at a 5‐day hydraulic retention time (HRT), and all reactors displayed efficient nitrification and chemical oxygen demand (COD) removal (>95%). However, when HRT was reduced to 2·5 days, COD removal rates remained high, but nitrification efficiencies declined in all reactors after 19 days. To explain reduced performance, nitrifying bacterial communities (ammonia‐oxidizing bacteria, AOB; nitrite‐oxidizing bacteria, NOB) were examined in the liquid phase and also on the beads using qPCR, FISH and DGGE. Overall, the presence of the beads in a reactor promoted bacterial abundances and diversity, but as bead size was increased, biofilms with active coupled AOB–NOB activity were less apparent, resulting in incomplete nitrification.

Conclusions

Hybrid bioreactors have potential to sustain effective nitrification at low HRTs, but support media size and configuration type must be optimized to ensure coupled AOB and NOB activity in nitrification.

Significance and Impact of the Study

This study shows that AOB and NOB coupling must be accomplished to minimize nitrification failure.     

Stimulatory Effect of Xenobiotics on Oxidative Electron Transport of Chemolithotrophic Nitrifying Bacteria Used as Biosensing Element

Electron transport chain (ETCh) of ammonium (AOB) and nitrite oxidizing bacteria (NOB) participates in oxidation of ammonium to nitrate (nitrification). Operation of ETCh may be perturbed by a range of water-soluble xenobiotics. Therefore, consortia of nitrifying bacteria may be used as a biosensor to detect water contamination. A surprising feature of this system is an increase of oxygen consumption, detected in the presence of certain inhibitors of ETCh. Thus, to shed light on the mechanism of this effect (and other differences between inhibitors) we monitored separately respiration of the bacteria of the first (AOB - Nitrosomonas) and second (NOB -Nitrobacter) stages of nitrification. Furthermore, we measured plasma membrane potential and the level of reduction of NAD(P)H. We propose a novel model of ETCh in NOB to explain the role of reverse electron transport in the stimulation of oxygen consumption (previously attributed to hormesis).     

Effects of roxithromycin on ammonia-oxidizing bacteria and nitrite-oxidizing bacteria in the rhizosphere of wheat

In a pot-cultural experiment, the impact of the antibiotic roxithromycin (ROX) addition was assessed on the diversities of microbial structure and function communities, especially involved in ammonia and nitrite oxidation in wheat rhizosphere soil with and without the addition of earthworms. The abundances of ammonia-oxidizing bacteria (AOB), nitrite-oxidizing bacteria (NOB), and total bacteria were surveyed by the quantitative PCR. The quantities of total bacteria, AOB, and NOB with earthworms were higher than those without earthworms because of the synergistic effect. ROX inhibited the growth of AOB in all treatments, although the quantities of AOB were in a light increase in medium and heavy polluted treatments compared with that in the light polluted treatments. Different from AOB, the quantities of NOB were lowest in light polluted treatments, but the quantities of NOB were rapidly increased in medium and heavy polluted treatments compared with that in the control. These results indicated that the application of ROX principally had a negative effect on nitrification performance by affecting the abundances and relative ratios of both AOB and NOB in soil communities, which affected the N cycle in an agricultural ecosystem. According to the metabolic diversities evaluated by the biologic assay, the tendency of metabolic diversities was quite contrary to the quantities of NOB in all treatments and showed the contrast growing relation of autotrophic and heterotrophic bacteria under ROX pollution pressure in agricultural ecosystems.     

Geospatial variation in co‐occurrence networks of nitrifying microbial guilds

Microbial communities transform nitrogen (N) compounds, thereby regulating the availability of N in soil. The N cycle is defined by interacting microbial functional groups, as inorganic N‐products formed in one process are the substrate in one or several other processes. The nitrification pathway is often a two‐step process in which bacterial or archaeal communities oxidize ammonia to nitrite, and bacterial communities further oxidize nitrite to nitrate. Little is known about the significance of interactions between ammonia‐oxidizing bacteria (AOB) and archaea (AOA) and nitrite‐oxidizing bacterial communities (NOB) in determining the spatial variation of overall nitrifier community structure. We hypothesize that nonrandom associations exist between different AO and NOB lineages that, along with edaphic factors, shape field‐scale spatial patterns of nitrifying communities. To address this, we sequenced and quantified the abundance of AOA, AOB, and Nitrospira and Nitrobacter NOB communities across a 44‐hectare site with agricultural fields. The abundance of Nitrobacter communities was significantly associated only with AOB abundance, while that of Nitrospira was correlated to AOA. Network analysis and geostatistical modelling revealed distinct modules of co‐occurring AO and NOB groups occupying disparate areas, with each module dominated by different lineages and associated with different edaphic factors. Local communities were characterized by a high proportion of module‐connecting versus module‐hub nodes, indicating that nitrifier assemblages in these soils are shaped by fluctuating conditions. Overall, our results demonstrate the utility of network analysis in accounting for potential biotic interactions that define the niche space of nitrifying communities at scales compatible to soil management.     

Activity of Nitrifying Biofilms Constructed on Low-Density Polyester Enhances Bioremediation of a Coastal Wastewater Effluent

Summary Nitrifying biofilms were constructed on low density polyester Dacron for the bioremediation of nitrogen from wastewater effluent of a municipal treatment plant. Dacron disks were inoculated with wastewater sludge enriched for 15 days for either ammonia- or nitrite-oxidizing bacteria (AOB or NOB, respectively) and packed into glass bioreactors. Wastewater effluent containing high levels of ammonia, nitrite, and phosphate was collected and fed to inoculated and uninoculated bioreactors. Both inoculated bioreactors showed stable nitrification efficiencies, removing 96 and 76% of the ammonia and 12 and 35% of the nitrite for AOB- and NOB-inoculated bioreactors, respectively. Efficiencies of phosphate removal were similar in both inoculated and uninoculated bioreactors, indicating that nitrifiers were not required for this process. AOB-inoculated bioreactors accumulated nitrite mid-way through the experiment and had low rates of conversion to nitrate, suggesting slow nitrite oxidizer growth. DGGE and sequence analysis of AOB 16S rRNA genes showed enrichment of Nitrosomonas spp. in both inoculated bioreactors, and a dominance of Nitrosospira spp. in non-inoculated bioreactors. This study describes an inexpensive and efficient technology for removing ammonia and nitrite from wastewater effluents of municipal treatment plants before its release to the environment.     

Partial nitrification to nitrite using low dissolved oxygen concentration as the main selection factor

Partial nitrification to nitrite (nitritation) can be achieved in a continuous process without sludge retention by wash out of nitrite oxidising bacteria (NOB) while retaining ammonia oxidising bacteria (AOB), at elevated temperatures (the SHARON process) and, as demonstrated in this paper, also at low dissolved oxygen (DO) concentrations. Enriched AOB was attained at a low DO concentration (0.4 mg l⁻¹) and a dilution rate of 0.42 day⁻¹ in a continuous process. A higher oxygen affinity of AOB compared to NOB seemed critical to achieving this. This was verified by determining the oxygen half saturation constant, K _o, with similar oxygen mass transfer resistances for enriched AOB and NOB as 0.033 ± 0.003 mg l⁻¹ and 0.43 ± 0.08 mg l⁻¹, respectively. However, the extent of nitritation attained was found to be highly sensitive to process upsets.     

Unravelling the reasons for disproportion in the ratio of AOB and NOB in aerobic granular sludge

In this study, we analysed the nitrifying microbial community (ammonium-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB)) within three different aerobic granular sludge treatment systems as well as within one flocculent sludge system. Granular samples were taken from one pilot plant run on municipal wastewater as well as from two lab-scale reactors. Fluorescent in situ hybridization (FISH) and quantitative PCR (qPCR) showed that Nitrobacter was the dominant NOB in acetate-fed aerobic granules. In the conventional system, both Nitrospira and Nitrobacter were present in similar amounts. Remarkably, the NOB/AOB ratio in aerobic granular sludge was elevated but not in the conventional treatment plant suggesting that the growth of Nitrobacter within aerobic granular sludge, in particular, was partly uncoupled from the lithotrophic nitrite supply from AOB. This was supported by activity measurements which showed an approximately threefold higher nitrite oxidizing capacity than ammonium oxidizing capacity. Based on these findings, two hypotheses were considered: either Nitrobacter grew mixotrophically by acetate-dependent dissimilatory nitrate reduction (ping-pong effect) or a nitrite oxidation/nitrate reduction loop (nitrite loop) occurred in which denitrifiers reduced nitrate to nitrite supplying additional nitrite for the NOB apart from the AOB.     

Nitrite Removal Performance and Community Structure of Nitrite-Oxidizing and Heterotrophic Bacteria Suffered with Organic Matter

Altlhough ammonia oxidation and ammonia-oxidizing bacteria (AOB) have been extensively studied, nitrite oxidation and nitrite-oxidizing bacteria (NOB) are still not well understood. In this article, the effect of organic matter on NOB and heterotrophic bacteria was investigated with functional performance analysis and bacterial community shift analysis. The results showed that at low concentrations of initial sodium acetate [initial sodium acetate (ISA) = 0.5 or 1 g/L], the nitrite removal rate was higher than that obtained under autotrophic conditions and the bacteria had a single growth phase, whereas at high ISA concentrations (5 or 10 g/L), continuous aerobic nitrification and denitrification occurred in addition to higher nitrite removal rates, and the bacteria had double growth phases. The community structure of total bacteria strikingly varied with the different concentrations of ISA; the dominant populations shifted from autotrophic and oligotrophic bacteria (NOB, and some strains of Bacteroidetes, Alphaproteobacteria, Actinobacteria, and green nonsulfur bacteria) to heterotrophic and denitrifying bacteria (strains of Gammaproteobacteria, especially Pseudomonas stutzeri and P. nitroreducens). The reasons that nitrite removal rate increased with supplement of organic matters were discussed.