Inadvertently enriched cyanobacteria prompted bacterial phosphorus uptake without aeration in a conventional anaerobic/oxic reactor.

The enhanced biological phosphorus removal (EBPR) process requires alternate anaerobic and aerobic conditions, which are regulated respectively by aeration off and on. Recently, in an ordinary EBPR reactor, an abnormal orthophosphate concentration (PO43--P) decline in the anaerobic stage (namely non-aerated phosphorus uptake) aroused attention. It was not occasionally but occurred in each cycle and lasted for 101 d and shared about 16.63 % in the total P uptake amount. After excluding bio-mineralization and surface re-aeration, indoor light conditions (180 to 260 lx) inducing non-aerated P uptake were confirmed. High-throughput sequencing analysis revealed that cyanobacteria could produce oxygen via photosynthesis and were inhabited inside wall biofilm. The cyanobacteria (Pantalinema and Leptolyngbya ANT.L52.2) were incubated in a feeding transparent silicone hose, entered the reactor along with influent, and outcompeted Chlorophyta, which existed in the inoculum. Eventually, this work deciphered the reason for non-aerated phosphorus uptake and indicated its potential application in reducing CO2 emissions and energy consumption via the cooperation of microalgal-bacterial and biofilm-sludge.

Elucidating performance failure in the use of an Anaerobic-Oxic-Anoxic (AOA) plug-flow system for biological nutrient removal.

The Anaerobic-oxic-anoxic (AOA) process is a carbon-saving and high-efficiency way to treat municipal wastewater and gets more attention. Recent reports suggest that in the AOA process, well-performed endogenous denitrification (ED), conducted by glycogen accumulating organisms (GAOs), is crucial to advanced nutrient removal. However, the consensuses about starting up and optimizing AOA, and in-situ enriching GAOs, are still lacking. Hence, this study tried to verify whether AOA could be established in an ongoing anaerobic-oxic (AO) system. For this aim, a lab-scale plug-flow reactor (working volume of 40 L) previously operated under AO mode for 150 days, during that 97.87 % of ammonium was oxidized to nitrate and 44.4 % of orthophosphate was absorbed. Contrary to expectations, under AOA mode, little nitrate reduction (only 6.3 mg/L within 5.33 h) indicated the failure of ED. According to high-throughput sequencing analysis, GAOs (Candidatus_Competibacter and Defluviicoccus) were enriched within the AO period (14.27 % and 3 %) and then still dominated during the AOA period (13.9 % and 10.07 %) but contributed little to ED. Although apparent alternate orthophosphate variations existed in this reactor, no typical phosphorus accumulating organisms were abundant (< 2 %). More than that, within the long-term AOA operation (109 days), the nitrification weakened (merely 40.11 % of ammonium been oxidized) since the dual effects of low dissolved oxygen and long unaerated duration. This work reveals the necessity of developing practical strategies for starting and optimizing AOA, and then three aspects in future studying are pointed out.

Towards low carbon demand and highly efficient nutrient removal: Establishing denitrifying phosphorus removal in a biofilm-based system.

The combined denitrifying phosphorus removal (DPR) and Anammox process is expected to achieve advanced nutrient removal with low carbon consumption. However, exchanging ammonia/nitrate between them is one limitation. This study investigated the feasibility of conducting DPR in a biofilm reactor to solve that problem. After 46-day anaerobic/aerobic operation, high phosphorus removal efficiency (PRE, 83.15 %) was obtained in the activated sludge (AS) and biofilm co-existed system, in which the AS performed better. Phosphate-accumulating organisms might quickly adapt to the anoxic introduced nitrate, but the following aerobic stage ensured a low effluent orthophosphate (<1.03 mg/L). Because of waste sludge discharging and AS transforming to biofilm, the suspended solids dropped below 60 mg/L on Day 100, resulting in PRE decline (17.17 %) and effluent orthophosphate rise (4.23 mg/L). Metagenomes analysis revealed that Pseudomonas and Thiothrix had genes for denitrification and encoding Pit phosphate transporter, and Candidatus_Competibacter was necessary for biofilm formation.

Unexpected phosphorous removal in a Candidatus_Competibacter and Defluviicoccus dominated reactor.

Competition between polyphosphate- and glycogen-accumulating organisms (PAOs and GAOs) is problematic in the enhanced biological phosphorus removal (EBPR) process. Aiming at a high phosphorus removal efficiency (PRE), the phosphorus release amount (PRA) is considered an essential evaluating indicator. However, the correlations between PRE and PRA and the abundance of PAOs are not clear. In this study, the EBPR was established and optimized via adjusting influent carbon to phosphorus ratio (C/P). After 110-day operation, 17.67 mg/L of PRA and 75.86% of PRE simultaneously achieved with influent C/P of 40 mgCOD/mgP. As for PAOs, Candidatus_Accumulibacter and Tetrasphaera were absent, while Hypomicrobium (3.69%), Pseudofulvimonas (1.02%), and unclassified_f_Rhodobacteraceae (2.41%) were found at a low level. On the contrary, Candidatus_Competibacter and Defluviicoccus were unexpectedly enriched with high abundance (24.94% and 16.04%, respectively). These results also suggested that it was difficult to distinguish whether PAOs were enriched merely based on the variations of PRA and PRE.

Development of novel denitrifying nitrite accumulation and phosphorus removal (DNAPR) process for offering an alternative pretreatment to achieve mainstream Anammox.

For achieving mainstream anaerobic ammonium oxidation (Anammox), there is a need to achieve organic carbon and phosphorus removal meanwhile supplying nitrite (NO2--N). Based on this demand, a novel anaerobic/anoxic/aerobic operated denitrifying nitrite accumulation and phosphorus removal (DNAPR) process was proposed for treating synthetic municipal and nitrate (NO3--N) wastewaters simultaneously (volume ratio of 5:1). By adjusting influent composition, discharging anaerobic-end supernatant, shortening anoxic duration, and adding a short aerobic stage, DNAPR process achieved promising and stable nitrate-to-nitrite transformation (78.35%) and phosphorus removal (98.34%) performance. Moreover, effluent with chemical oxygen demand of 16.63 mg/L, nitrite of 54.16 mg/L, orthophosphate of 0.37 mg/L, and nitrite to ammonia ratio of 1.3 were finally obtained after 141-day operation. Microbiological analysis showed that Thauera (34.9%) and unclassified_f_Rhodobacteraceae (6.79%) were both responsible for DNAPR. Therefore, DNAPR, serving as promising alternative pretreatment, might possess significance for achieving mainstream Anammox.

Feasibility of reinforced post-endogenous denitrification coupling with synchronous nitritation, denitrification and phosphorus removal for high-nitrate sewage treatment using limited carbon source in municipal wastewater.

Post-endogenous denitrification (PED) process, utilizing internal rather than external carbons, has been proposed for nitrogen removal from wastewaters. However, its potential nitrogen removal capacity has not been approached, especially when facing simultaneous phosphorus removal. Here, the nitrogen removal ability of PED was further investigated by treating municipal and high-nitrate wastewaters in a novel process combined with synchronous nitritation, denitrification and phosphorus removal (SNiDPR). After optimization, the anoxic specific nitrite (and nitrate) reduction rate was increased from 0.41 to 1.13 mgN gVSS-1 h-1, accompanied with PED efficiency raising from 16.8% to 80.9%. It ensured that, by utilizing the limited organic carbons in municipal wastewater, deep-level nutrient removal could still be achieved (total nitrogen and phosphorus removal efficiencies were 93.1% and 99.9%, respectively). Nitrospira (0.1-0.4%) was outcompeted by Nitrosomonas (4.7-3.3%), which contributed to accumulation of nitrite in aerobic stage (99.6%) and dramatically reduced the carbons demand of following PED. Enriched Dechloromonas (8.5-5.6%) and Candidatus_Competibacter (9.1-11.3%) might play key roles in sufficient utilization of organic carbons in municipal wastewater anaerobically, and respectively facilitate aerobic phosphorus removal (100%) and anoxic PED (60.7% of overall nitrogen removal).

Achieving deep-level nutrient removal via combined denitrifying phosphorus removal and simultaneous partial nitrification-endogenous denitrification process in a single-sludge sequencing batch reactor.

The feasibility of coupling denitrifying phosphorus removal (DPR) with simultaneous partial nitrification-endogenous denitrification (SPNED) was investigated in a single-sludge sequencing batch reactor for deep-level nutrient removal from municipal and nitrate wastewaters. After 160-day operation, the DPR process simultaneously reduced most PO43--P and NO3--N anoxically, and the SPNED process achieved further total nitrogen (TN) removal at low dissolved oxygen condition with TN removal efficiency of 90.8%. The effluent NH4+-N, PO43--P and TN concentrations were 1.0, 0.1 and 7.2 mg/L, respectively. Microbial analysis revealed that Dechloromonas (6.7%) dominated DPR process, whereas the gradually enriched Nitrosomonas (4.5%) and Candidatus Competibacter (6.8%) conducted SPNED process accompanied with sharply eliminated Nitrospirae (1.4%). Based on these findings, a novel strategy was proposed to achieve further nutrient removal in conventional treatment through integrating the DPR-SPNED process. As a result, ∼100% of extra carbon and ∼10% of oxygen consumptions would be reduced with satisfactory effluent quality.

Evaluating the potential for sustaining mainstream anammox by endogenous partial denitrification and phosphorus removal for energy-efficient wastewater treatment.

This study demonstrated a novel process configuration for sustaining mainstream anammox by integrating the anammox and endogenous partial denitrification-and-phosphorus removal (EPDPR) in two-stage sequencing batch reactors (SBRs). In the EPDPR-SBR, high nitrate-to-nitrite transformation (68.2%) and P removal (99.3%) were achieved by adjusting the anaerobic/anoxic/aerobic durations and influent nitrate concentration, providing a suitable NO2--N/NH4+-N (∼1.37) for subsequent anammox reaction. In the Anammox-SBR, ∼95% of TN was removed without external carbon and oxygen demands. Satisfactory effluent quality (∼6 mgTN/L and 0.2 mgP/L) achieved in the integrated EPDPR/anammox opens a new window towards the energy-efficient wastewater treatment. Microbial analysis further revealed that Dechloromonas (1.6-9.6%) and Candidatus Competibacter (6.4-5.8%) respectively conducted P removal and NO2--N production (79.2%) from NO3--N denitrification in the EPDPR-SBR, whereas Candidatus Kuenenia (7.0-29.7%) dominated NO2--N and NH4+-N removal (91.3% and 99.5%) in the Anammox-SBR, with 10 genera identified as denitrifying bacteria (0.6-8.1%) further reduced 18.9% of the produced NO3--N.

[Effect of the Influent C/P Ratio on the Nutrient Removal Characteristics of the SNEDPR System].

This study focuses on the nitrogen (N) and phosphorus (P) removal characteristics in a simultaneous nitrification-endogenous denitrification and phosphorus removal (SNEDPR) system at different influent C/P ratios. An extended anaerobic/low aerobic (dissolved oxygen:0.5-1.0 mg·L-1) sequencing batch reactor (SBR) fed with municipal sewage was studied by adjusting different C/P ratios (10, 15, 20, 30, and 60). The experimental results show that the proper reduction of the influent C/P ratio (C/P ratio reduced from 60 to 30) enhances the competitive advantages of phosphorus-accumulating organisms (PAOs) in the SNEDPR system. The highest phosphorus removal efficiency was achieved at a C/P ratio of 30, with the anaerobic phosphorus release rate (PRR) and aerobic phosphorus uptake rate (PUR, used as P/MLSS) reaching 3.5 mg·(g·h)-1 and 4.2 mg·(g·h)-1 respectively, and an average effluent PO43--P concentration below 0.3 mg·L-1. The percentage of PAOs contributing to the storage of endogenesis carbon (PPAO, An) reached 88.1%. However, a poor phosphorus removal performance was observed with further reduction of the influent C/P ratios to 10; both the PO43--P removal efficiency and PPAO, An decreased from 38.1% and 82.4% to 3.1% and 5.3%, respectively. The PRR and PUR were 0.2 mg·(g·h)-1 and 0.24 mg·(g·h)-1, respectively. The COD removal performance was not affected by the decreasing influent C/P ratios; the average COD removal efficiency stabilized at 85%. In addition, the nitrification performance became worse with decreasing C/P ratios (from 60 to 20) because the effluent NH4+-N and NO2--N concentrations increased from 0 and 6.9 mg·L-1 to 5.1 mg·L-1 and 16.2 mg·L-1, respectively. The nitrificaton performance recovered when the C/P ratios further decreased to 10, but the nitrite accumulation was disturbed as both the effluent NH4+-N and NO2--N concentrations reduced to 0. The effluent NO3--N concentration increased from 0.08 mg·L-1 to 14.1 mg·L-1. The SNED efficiency first decreased from 62.1% to 36.4% and then increased to 56.4%. The advantageous competition of glycogen accumulating organisms (GAOs) improved when the influent C/P ratio was lower than 15. The enhancement of the endogenous denitrification ability of GAOs might explain the recovery denitrification performance of the system when the influent C/P ratios decreased from 20 to 10.

[Effect of Different Sludge Retention Time (SRT) Operations on the Nutrient Removal Characteristics of a SNEDPR System].

This study focuses on the nitrogen (N) and phosphorus (P) removal characteristics in a simultaneous nitrification endogenous denitrification and phosphorus removal (SNEDPR) system operating at different sludge retention time (SRT). Four extended anaerobic/low aerobic (dissolved oxygen:0.5-1.0 mg·L-1)-operated sequencing batch reactors (SBRs) fed with municipal sewage were studied at different SRT of 5, 10, 15, and 25 d. The experimental results show that a shorter SRT at an SRT ≥ 10 d enhances the competitive advantage of PAOs in the system and an efficient phosphorus removal performance of the SNEDPR system was achieved at a SRT of 10 d and 15 d. Especially at an SRT of 10 d; the average PPAOs, An was 68.4%, the PRA and PUA reached 31.9 and 34.3 mg·L-1, respectively. The nitrification performance of the system was not affected by SRT changes. The most efficient nitrogen removal performance was achieved at a SRT of 15 d, with a high average TN removal and SNED efficiencies reaching 89.6% and 71.8%, respectively. At a SRT ≥ 10 d, the COD removal performance of the SNEDPR system was also not affected by SRT changes. The COD removal efficiencies were higher than 78%. However, when the SRT was shortened to 5 d, the C, N, and P performances of the system worsened due to the loss of biomass; the SNED and PO43--P removal efficiencies were as low as 5.7% and 0.5%, respectively. In addition, at an SRT=15 d, the sludge-settling performance of the system was the best. The SV and SVI were 20% and 64 mL·g-1, respectively, and the sludge concentration increased with the extension of the SRT. Under long SRT (25 d) operation, the system showed a good resistance to shock loads, but the sedimentation performance of the sludge deteriorated.

[Denitrification and Phosphorus Removal from Low C/N Urban Sewage Based on a Post-Partial Denitrification AOA-SBR Process].

This study focuses on the investigation of the nitrogen (N) and phosphorus (P) removal characteristics of a combination of enhanced phosphorus removal (EBPR) with simultaneous partial nitrification endogenous denitrification (SPND) and post-partial denitrification process. An anaerobic/aerobic/anoxic (A/O/A) operated sequencing batch reactor (SBR) fed with urban sewage was optimized by regulating the aeration rate and anoxic time. Based on this optimization, deep-level nitrogen and phosphorus removals from low C/N urban sewage could be realized. The experimental results show that the effluent PO43--P concentration decreased from 0.06 mg·L-1 to 0 mg·L-1, the effluent NH4+-N, NO2--N, and NO3--N concentrations gradually decreased from 0.18, 18.79, and 0.08 mg·L-1 to 0, 16.46, and 0.05 mg·L-1, respectively, and the TN removal efficiency increased from 72.69% to 77.97% when the aeration rate decreased from 1.0 L·min-1 to 0.6 L·min-1 and the anoxic duration was 180 min. With the reduction of the aeration rate, the SPND phenomenon became notable and the SND rate increased from 19.18% to 31.20%. When the anoxic duration was extended from 180 min to 420 min, the effluent PO43--P, NH4+-N, and NO3--N concentrations stabilized at~0, 0, and 0.03 mg·L-1, respectively. The effluent NO2--N concentration was as low as 3.06 mg·L-1, the SND rate was~32.21%, the TN removal performance gradually improved, and the TN removal efficiency was as high as 99.42%. Thus, deep-level nitrogen and phosphorus removals could be realized with the SPNDPR-PD system.

[Effect of Influent C/N Ratio on the Nutrient Removal Characteristics of SNEDPR Systems].

To determine the performance of nitrogen and phosphorus removal within a simultaneous nitrification endogenous denitrification system (SNEDPR), an extended anaerobic/low aerobic (dissolved oxygen:0.5-2.0 mg·L-1)-operated sequencing batch reactor (SBR) was fed with simulation wastewater. The SBR was initiated under a constant influent C/N ratio of 10, with the simultaneous enrichment of polyphosphate-accumulating organisms (PAOs). It was then investigated at different influent C/N ratios of 10, 7.5, 5, and 2.5. The experimental results indicated that, when the influent C/N ratio was 10, SNEDPR could be successfully started up. The effluent PO43--P and total nitrogen (TN) concentrations were 0.1 mg·L-1 and 8.1 mg·L-1. PO43--P efficiency, TN efficiency, and SNED efficiency were 99.79%, 89.38%, and 58.0%, respectively. When the influent C/N ratio increased from 5 to 10, the nitrogen and phosphorus removal performance of the system improved with PRA, and SNED efficiency increased from 16.0 m·L-1 and 48.0% to 24.4 mg·L-1 and 69.2%, respectively. When the C/N ratio was 10, the TN and PO43--P removal efficiencies increased to 94.5% and 100%, respectfully. When the C/N ratio was decreased to 2.5, the nitrogen and phosphorus removal performance of the system decreased. The PRA and SNED efficiencies were only 1.36 mg·L-1 and 10%, respectively. During the stable phase of the system (C/N ratio were 10, 7.5 and 5), SNED efficiency reached to 85.9%, with the average effluent concentration of NH4+-N, x--N, and PO43--P being 0.0, 8.1, and 0.1 mg·L-1, respectively.

[Operating Characteristics of a DPR-SNED System Treating Low C/N Municipal Wastewater and Nitrate-containing Sewage].

In order to realize the simultaneous treatment of low C/N municipal wastewater and high nitrate wastewater, a sequencing batch reactor (SBR), inoculated with activated sludge, was used to initiate the denitrifying phosphorus removal coupled with simultaneous nitrification and endogenous denitrification (DPR-SNED). The anaerobic/anoxic/hypoxic durations and dissolved oxygen (DO) concentration were appropriately controlled, and the nitrogen and phosphorus removal characteristics were examined. The experimental results demonstrated that, in the anaerobic/hypoxia operation mode, with an anaerobic duration of 3 h and DO concentration of 0.5-1.0 mg·L-1, the simultaneous nitrification of phosphorus removal (SNEDPR) system successfully began in 60 d. The effluent PO43--P concentration was below 0.5 mg·L-1, the nutrient and COD removal efficiencies were stably maintained above 90% and 80%, respectively, and the SNED efficiency and CODins efficiency reached 70% and 95%, respectively. When the operation mode was anaerobic/anoxic/hypoxic and nitrate-containing sewage was added at the beginning of the anoxic stage, DPR-SNED was achieved with the effluent PO43--P concentration<0.5 mg·L-1, nutrient and COD removal efficiencies above 88% and 90%, respectively, and SNED efficiency and CODins efficiency maintained at 62% and 90%, respectively. After the successful initiation of DPR-SNED, enhanced intracellular carbons storage was achieved by phosphorus-and glycogen-accumulating organisms using the limited carbons in raw municipal wastewater to provide sufficient carbon sources for subsequent nutrient removal. In addition, the endogenous partial denitrification ensured the efficient nitrogen removal performance of the DPR-SNED system at low C/N conditions (average 4).

[Simultaneous Nitrogen and Phosphorus Removal Characteristics of An Anaerobic/Aerobic Operated SPNDPR System Treating Low C/N Urban Sewage].

This study focused on the nitrogen (N) and phosphorus (P) removal performance optimization of simultaneous partial nitrification-endogenous denitrification and phosphorus removal (SPNDPR) systems. An anaerobic (180 min)/aerobic operated sequencing batch reactor (SBR) fed with domestic wastewater was used for investigating the startup and optimization of SPNDPR by regulating the aeration rate and aerobic duration time. The experimental results showed that at an aerobic aeration rate of 0.8 L·min-1 and aerobic duration time of 150 min, the effluent PO43--P concentration was about 1.5 mg·L-1, with the effluent NH4+-N and NO3--N concentrations gradually decreasing from 10.28 and 8.14 mg·L-1 to 0 and 2.27 mg·L-1, respectively, and effluent NO2--N concentration increasing to 1.81 mg·L-1. When the aeration rate was increased to 1.0 L·min-1 and the aerobic duration time was shortened to 120 min, the phosphorus removal and partial nitrification-endogenous performance of the system gradually increased, but the total nitrogen (TN) removal performance initially decreased and then gradually increased. The final effluent PO43--P and NH4+-N were stably below 0.5 and 1.0 mg·L-1, respectively, aerobic nitrite accumulation and simultaneous nitrification-endogenous denitrification (SND) efficiencies were 98.65 and 44.20%, respectively, and TN removal efficiency was 79.78%. The concurrence of aerobic phosphorus absorption, denitrifying phosphorus removal, partial nitrification, and nitrification-endogenous in the aerobic stage of the SPNDPR system ensured the simultaneous removal of N and P from low C/N wastewater.