The nitritation step is the fundament for the biological nitrogen removal regardless of the traditional nitrification and denitrification process, the nitrite shunt process or the anammox process. ...Thus, exploring the effective nitritation performance is an important aspect of biological nitrogen removal. This study explored the upper limit of nitritation rate by increasing hydraulic residence time with the well-mixed and continuous granular sludge-type reactor characterized with low complexity and easy operation. The results showed that with the nitrogen loading rate of 1.0 kg/m3/d, the nitrite production rate could reach up to 0.65 kg/m3/d with the nitrite production efficiency of 63.49%, which is remarkable compared to that in the previously similar research. The microbial analysis indicated that ammonia-oxidizing bacteria was successfully enriched (13.27%) and genus Nitrosomonas was the dominant bacteria type. Besides, the activity of ammonium oxidizing bacteria in the continuous flow reactor was higher than that of other reactor types. The growth of vorticella on the sludge was also found in the reactor. The test of specific sludge activity and the microbe analysis both indicated that the nitrite-oxidizing bacteria was well inhibited during the whole experiment, which indicated the strategy of simply adjusting the dissolved oxygen is effective for running of nitritation process. The phosphorus removal performance was also achieved with a removal efficiency of 23.53%. The functional composition of the microbial community in the samples was predicted and finally transformation mechanism of nitrogen in sludge was drawn. In sum, this study indicated the superior performance of the granule sludge-type nitritation process and give a reference for the application of biological nitrogen removal technology.
The control of nitrite-oxidizing bacteria (NOB) challenges the implementation of partial nitritation and anammox (PN/A) processes under mainstream conditions. The aim of the present study was to ...understand how operating conditions impact microbial competition and the control of NOB in hybrid PN/A systems, where biofilm and flocs coexist. A hybrid PN/A moving-bed biofilm reactor (MBBR; also referred to as integrated fixed film activated sludge or IFAS) was operated at 15 °C on aerobically pre-treated municipal wastewater (23 mgNH4-N L−1). Ammonium-oxidizing bacteria (AOB) and NOB were enriched primarily in the flocs, and anammox bacteria (AMX) in the biofilm. After decreasing the dissolved oxygen concentration (DO) from 1.2 to 0.17 mgO2 L−1 - with all other operating conditions unchanged - washout of NOB from the flocs was observed. The activity of the minor NOB fraction remaining in the biofilm was suppressed at low DO. As a result, low effluent NO3− concentrations (0.5 mgN L−1) were consistently achieved at aerobic nitrogen removal rates (80 mgN L−1 d−1) comparable to those of conventional treatment plants. A simple dynamic mathematical model, assuming perfect biomass segregation with AOB and NOB in the flocs and AMX in the biofilm, was able to qualitatively reproduce the selective washout of NOB from the flocs in response to the decrease in DO-setpoint. Similarly, numerical simulations indicated that flocs removal is an effective operational strategy to achieve the selective washout of NOB. The direct competition for NO2− between NOB and AMX - the latter retained in the biofilm and acting as a “NO2-sink” - was identified by the model as key mechanism leading to a difference in the actual growth rates of AOB and NOB (i.e., μNOB < μAOB in flocs) and allowing for the selective NOB washout over a broad range of simulated sludge retention times (SRT = 6.8–24.5 d). Experimental results and model predictions demonstrate the increased operational flexibility, in terms of variables that can be easily controlled by operators, offered by hybrid systems as compared to solely biofilm systems for the control of NOB in mainstream PN/A applications.
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•Hybrid PN/A systems provide increased operational flexibility for NOB control•AOB and NOB enrich primarily in the flocs, and AMX in the biofilm (“NO2-sink”)•AMX use NO2− allowing to differentiate AOB and NOB growth rates•A decrease in DO or an increase in floc removal leads to selective NOB washout from flocs•The activity of the minor NOB fraction in the biofilm is suppressed at limiting DO
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•A strategy for recovering partial nitritation in a mainstream PN/A system is proposed.•Selectively reviving AOB after thoroughly inhibiting AOB and NOB by FNA is applied.•NOB reduced ...largely whereas AOB declined merely after 1.35 mg/L FNA treatment.•Ca. Kuenenia and Ca. Brocadia formed important links with other N cycle processes.
Starting up or recovering partial nitritation is a major challenge for achieving or maintaining stable partial nitritation/anammox (PN/A) during mainstream wastewater treatment. This study presents a novel strategy for recovering the nitrite pathway by selectively reviving ammonium oxidizing bacteria (AOB) after thoroughly inhibiting AOB and nitrite oxidizing bacteria (NOB) using free nitrous acid (FNA). A sequencing batch reactor was operated for PN/A to treat real domestic wastewater for 423 days, during which twice FNA treatment was temporarily implemented. Results showed that with a single 0.45 mg/L FNA treatment on flocculent sludge, the NO3−-N concentration during the aerobic period showed an uptrend again and the partial nitritation performance was deteriorated. In contrast, 1.35 mg/L FNA treatment induced the inhibition of both AOB and NOB leading to regressive ammonium oxidation, but a subsequently higher DO (1.5 mg/L) and longer aeration duration recovered partial nitritation. For the relative abundances of the acquired biomass related to nitrogen conversion, Nitrosomonas, Nitrospira and Nitrolancea increased to 9.65%, 10.27% and 4.35%, respectively, at the beginning of the 1.35 mg/L FNA treatment, and Nitrospira and Nitrolancea decreased to 2.80% and 0.03% whereas Nitrosomonas declined to 8.71% after 76 days. Ca. Brocadia showed less resilience after the 1.35 mg/L FNA treatment, with the relative abundance decreasing from 13.38% to 0.62% due to insufficient nitrite. Molecular ecological network analysis indicates that among anammox taxa, Ca. Kuenenia and Ca. Brocadia formed important links with other N cycle processes. Moreover, the proposed strategy shows operational flexibility because it can be easily used to control NOB in mainstream PN/A applications offered by flocculent sludge systems.
•Higher NH4+-N load led to higher oxygen transfer rate and lower NOB activity.•NOB was inhibited by low DO concentration in the limited space of aerobic biofilm.•Closed-end aeration mode favored ...Nitrosomonas and inhibited Nitrospira.•A strategy was proposed for quick start-up and stable mainstream PN in MABR.
The proliferation of nitrite oxidizing bacteria (NOB) still remains as a major challenge for nitrogen removal in mainstream wastewater treatment process based on partial nitrification (PN). This study investigated different operational conditions to establish mainstream PN for the fast start-up of membrane aerated biofilm reactor (MABR) systems. Different oxygen controlling strategies were adopted by employing different influent NH4+-N loads and oxygen supply strategies to inhibit NOB. We indicated the essential for NOB suppression was to reduce the oxygen concentration of the inner biofilm and the thickness of aerobic biofilm. A higher NH4+-N load (7.4 g-N/(m2·d)) induced higher oxygen utilization rate (14.4 g-O2/(m2·d)) and steeper gradient of oxygen concentration, which reduced the thickness of aerobic biofilm. Employing closed-end oxygen supply mode exhibited the minimum concentration of oxygen to realize PN, which was over 46% reduction of the normal open-end oxygen mode. Under the conditions of high NH4+-N load and closed-end oxygen supply mode, the microbial community exhibited a comparative advantage of ammonium oxidizing bacteria over NOB in the aerobic biofilm, with a relative abundance of Nitrosomonas of 30–40% and no detection of Nitrospira. The optimal fast start-up strategy was proposed with open-end aeration mode in the first 10 days and closed-end mode subsequently under high NH4+-N load. The results revealed the mechanism of NOB inhibition on the biofilm and provided strategies for a quick start-up and stable mainstream PN simultaneously, which poses great significance for the future application of MABR.
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•Continuous aeration in membrane-aerated biofilms resulted in full nitrification.•Intermittent aeration suppressed nitrite oxidation and supported anammox process.•Biofilm pH ...presented periodic upshifts with aeration switches.•Free ammonia speciation likely caused the inhibition of nitrite-oxidizing bacteria.•Heterotrophs established nitrous oxide-reduction zones under intermittent aeration.
Membrane-aerated biofilm reactors (MABR) are being applied for autotrophic nitrogen removal, yet control of nitrogen turnover remains challenging in MABR counter-diffusion biofilms. In this study, we regulated microbial activities in two lab-scale MABRs by providing continuous versus intermittent aeration. Nitrogen consumption by different functional microbial groups was estimated from bulk measurements via a mass balance approach. Nitrite-oxidizing bacteria (NOB) proliferated under continuous aeration while they were significantly suppressed under intermittent aeration, and NOB suppression activated anaerobic ammonium oxidation. Nitritation performance in the MABR was studied through long-term bulk measurements and in situ biofilm microprofiles of dissolved oxygen (DO) and pH. During intermittent aeration pH effects rather than DO effects determined nitritation success, especially ammonia speciation, which serves as substrate and inhibitor in nitrification processes. Biofilm transition phases were monitored upon aeration switches. Canonical correspondence analysis suggested that the relative transition after anoxia and aeration intermittency were less decisive for biofilm performance than the relative aeration duration. Heterotrophic bacteria displayed minor denitrification rates with aeration control, but contributed to mitigation of nitrous oxide (N2O) emissions. N2O production hotspots were identified at the top of the anoxic biofilm zone under continuous aeration. Instead, under intermittent aeration an anoxic N2O reduction zone was established. Our observations support intermittent aeration control of MABRs as a simple strategy for energy-efficient nitrogen removal with low N2O emission. .
•Current molecular technologies to identify and detect AOB/NOB are summarized.•Strategies for partial nitrification via nitrite are critically reviewed.•Combination of partial nitrification and ...Anammox benefits wastewater treatment.•Existing problems and directions for future research in partial nitrification are suggested.
Partial nitrification has gained broad interests in the biological nitrogen removal (BNR) from wastewater, since it alleviates carbon limitation issues and acts as a shortcut nitrogen removal system combined with anaerobic ammonium oxidation (Anammox) process. The occurrence and maintenance of partial nitrification relies on various conditions, which favor ammonium oxidizing bacteria (AOB) but inhibit or limit nitrite oxidizing bacteria (NOB). The studies of the AOB and NOB activities have been conducted by state-of-the-art molecular techniques, such as Polymerase Chain Reaction (PCR), Quantitative PCR, denaturing gradient gel electrophoresis (DGGE), Fluorescence in situ hybridization (FISH) technique, Terminal Restriction Fragment Length Polymorphism (T-RFLP), Live/Dead BacLight, and quinone profile. Furthermore, control strategies for obtaining partial nitrification are mainly focused on the pH, temperature, dissolved oxygen concentration, real-time aeration control, sludge retention time, substrate concentration, alternating anoxic and aerobic operation, inhibitor and ultrasonic treatment. Existing problems and further perspectives for the scale-up of partial nitrification are also proposed and suggested.
Nobiletin (Nob) is a major component among the most reported polymethoxyflavones (PMFs), which possesses multiple efficacious healthcare activities. Owing to its high melting point and poor water ...solubility, the oral bioavailability of Nob needs to be improved via loading Nob on carriers. To take full advantage of Nob, the interaction mechanism between Nob and vehicles should be clarified. Herein, β-lactoglobulin (β-LG) was selected as the vehicle and further investigated the binding mechanism between Nob and β-LG. The binding stoichiometry of complex was found to be 1:1 by analysis of intrinsic fluorescence experiment. The results also confirmed by isothermal titration calorimetry (ITC) measurement that the binding behavior between β-LG and Nob was a spontaneously endothermic process driving mainly by hydrophobic interaction. Moreover, competitive binding and molecular docking method indicated the Nob was primary bound to internal calyx of β-LG at neutral pH. UV spectrophotometry revealed that the solubility of Nob was enhanced to 3 times by forming complex. Furthermore, Nob could alter secondary structure of β-LG by a transition from α-helix to β-sheet and lead to small increase on surface hydrophobicity of β-LG. This work will provide some valuable information on clarifying the interaction between protein and PMFs, which contributing to improve the poor bioavailability of PMFs.
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•Nobiletin had influence on the secondary and tertiary structure of β-lactoglobulin.•The hydrophobic interaction played a vital role in the interaction between Nobiletin and β-lactoglobulin.•The formation of β-lactoglobulin/Nobiletin complex was a spontaneously endothermic process.•At neutral pH, Nobiletin was primary bound into internal calyx of β-lactoglobulin.•Nobiletin solubility could be improved up to 3 times by forming β-lactoglobulin/Nobiletin complex.
•A concept of AnMBR-PN/A process was proposed for mainstream treatment.•In-situ FA/FNA exposure eliminated Nitrospira while retained Nitosonomas community.••DO limitation prevented switch of the NOB ...community in the mainstream conditions.•The effluent TN of the mainstream PN/A system was lower than 10 mg-N/L.•Enhanced hydrodynamic shear force resulted in sludge settleability deterioration.
In this pilot-scale study, an innovative mainstream treatment process that couples the anaerobic membrane reactor (AnMBR) with a one-stage PN/A system was proposed for advancing the concept of carbon neutrality in the municipal wastewater treatment plant. This work demonstrates the start-up procedure of a pilot-scale one-stage PN/A system for mainstream treatment. The 255-day start-up of the one-stage PN/A system involved the cultivation of ammonium-oxidizing bacteria (AOB) from the activated sludge, suppression of nitrite-oxidizing bacteria (NOB), investigation of in-situ growth kinetics of anammox bacteria (AnAOB), and the 50-day operation of the pilot-scale AnMBR-PN/A process for natural mainstream treatment. It is verified in the pilot-scale system for the first time that the in-situ free ammonia (FA) and free nitrous acid (FNA) exposure could effectively eliminate the Nitrospira (the NOB genus) while retaining the Nitosonomas (the AOB genus) community in the suspended sludge. NOB community rebounding was not detected even at the mainstream conditions with low nitrogen concentrations (Influent ammonium concentration=38±6 mg-NH4+-N/L) by intermittent aeration to control the system dissolved oxygen (DO) below 0.5 mg/L. The results of the mainstream treatment showed that the average effluent total nitrogen (TN) in the coupled process was generally lower than 10 mg-N/L, which meets the discharge limits of most prefectures in Japan. The investigated results of the in-situ anammox bacteria (AnAOB) growth kinetics suggested that the promoted start-up strategy of taking advantage of the warm months with higher mainstream temperature to achieve the rapid in-situ growth of the AnAOB is applicable in the investigated regions. From the perspective of the removal performance of the TN and organic substance, the AnMBR-PN/A process has great potential as the layouts of the carbon-neutral mainstream wastewater treatment plants.
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The partial nitritation-anammox (PNA) process is the most promising technique to treat municipal sewage; however, nitrite oxidizing bacteria (NOB) are a hindrance to achieve PNA. This study ...investigated the effects of selectively discharging flocs (<200 μm) to washout NOB in a sequencing batch reactor (SBR) over 200 d. The experiment was divided into three phases with different floc sludge retention times (SRTs; 30, 20 and 30 d). When the SRT of the flocs was reduced from 30 to 20 d to washout NOB, a significant reduction of ammonia oxidizing bacteria (AOB) and anaerobic ammonium-oxidizing (anammox) bacteria in the flocs was found. This indicates that a low floc SRT (20 d) leads to the loss of AOB and anammox bacteria in the flocs (<200 μm) and destroys PNA. Activity tests and qPCR analysis revealed the variations of functional bacteria in the granules and flocs, indicating that the enrichment of AOB, NOB, anammox bacteria in the granules is caused by the long-term discharging of flocs. High-throughput sequencing analysis revealed that the microbial shift of Tetrasphaera was significant in the flocs and may be connected to the enrichment of anammox bacteria and the stability of the PNA requires further research. All the obtained NOB sequences were affiliated with the genera Nitrospira and could further influence the PNA system. Overall, this study provides an in-depth understanding of the impact of discharging flocs to washout NOB and promotes the application of combing granules/floc PNA in sewage treatment.
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•Controlling floc SRTs of 30 d was beneficial to stable operation of PNA.•Floc SRTs of 20 d would result in the reduction of AOB and Anammox in flocs.•The distribution of NOB in granules and flocs was impacted by long-term discharging flocs.•The microbial community structure was seriously affected by discharging flocs.
Sludge treatment using free ammonia (FA) is an innovative approach that was recently reported effective achieving stable mainstream nitrogen removal via the nitrite pathway. This study aims to ...investigate the adaptation of nitrifying community and the response of nitrification performance to high-level of FA exposure under real wastewater conditions. Two parallel lab-scale sequencing batch reactors were operated and fed with real municipal wastewater, with one receiving sludge treatment by FA and another as a control. While the FA approach rapidly achieved partial nitrification with a nitrite accumulation ratio (NAR) of approximately 60%, the partial nitrification eventually failed due to nitrite-oxidizing bacteria (NOB) adaptation to FA inactivation. NOB activity in the inoculum was suppressed by 82% after exposure to FA at ~220 mg NH3-N/L. However, towards the end of the experiments, significantly higher NOB activities were observed after exposure to the same level of FA. Distinct behaviours of NOB observed in batch tests during the study supported the reactor operational data and strongly suggested the adaptation of NOB under the FA stress. Furthermore, microbial community analysis revealed the underlying mechanism of the observed adaptation: the dominant NOB changed from Nitrospira to Candidatus Nitrotoga. It is for the first time shown that Ca. Nitrotoga are highly resistant to FA inhibition and inactivation in comparison to Nitrospira and Nitrobacter. In addition, while the Nitrosomonas genus was always the dominant ammonia-oxidizing bacteria (AOB) throughout the study, different shift in a species level was observed.
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•Inactivation and adaptation of nitrifiers exposed to FA were investigated.•Dominant NOB genus shifted from Nitrospira to Ca. Nitrotoga under FA stress.•Ca. Nitrotoga are resistant to FA inhibition/inactivation compared with other NOB.•Nitrosomonas were always dominant AOB genus under FA stress.
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