Evaluation of the shear stability of aerobic granular sludge from a pilot-scale membrane bioreactor: Establishment of a quantitative method.

This work established a quantitative method to access the shear stability of aerobic granular sludge (AGS) and validated its feasibility by using the mature AGS from a pilot-scale (50 tons/day) membrane bioreactor (MBR) for treating real municipal wastewater. The results showed that the changing rate (ΔS) of the peak area (S) of granule size distribution (GSD) exhibited an exponential relationship (R2≥0.76) with the shear time (y=a-b·cx), which was a suitable indicative index to reflect the shear stability of different AGS samples. The limiting granule size (LGS) was defined and proposed to characterize the equilibrium size for AGS after being sheared for a period of time, whose value in terms of Dv50 showed high correlation (R2=0.92) with the parameter a. The free Ca2+ (28.44-34.21 mg/L) in the influent specifically interacted with polysaccharides (PS) in the granule's extracellular polymeric substance (EPS) as a nucleation site, thereby inducing the formation of Ca precipitation to enhance its Young's modulus, while Ca2+ primarily interacted with PS in soluble metabolic product (SMP) during the initial granulation process. Furthermore, the Young's modulus significantly affected the parameter a related to shear stability (R2=0.99). Since the parameter a was more closely related (R2=1.00) to ΔS than that of the parameter b or c, the excellent correlation (R2=0.99) between the parameter a and the wet density further verified the feasibility of this method.

Advanced temperature control in ethanol fermentation using a PSO-PID controller with split-range control strategy.

Global energy demand is experiencing a notable surge due to growing energy security. Renewable energy sources, like ethanol, are becoming more viable. In the present study, the application of a PSO-PID (Particle Swarm Optimization - Proportional Integral Derivative) controller with a split-range control strategy was suggested for the regulation of temperature within the fermentation system. To optimize performance, a POS-PID controller with a split-range arrangement utilizing two control valves for hot and cold utilities was constructed. The study began by examining the open-loop dynamic response of the system to inlet temperature and concentration disturbances during ethanol production fermentation. Subsequently, a transfer function model was developed through linearization at the steady-state operating point. The split-range controller structure, implemented by optimizing the PSO-PID controller parameters using PSO, effectively demonstrated temperature control in simulations of a nonlinear model. In this investigation, the ethanol fermentation system was modeled as a CSTR using a modified Monod equation for microbial growth kinetics. Various dynamic behavioral disturbances were explored and verified in the model with plant data in this study. The simulation model results were validated through plant data. The proposed method showed superior closed-loop performance with respect to errors, with the actuators proving to be effective than other reported methods for temperature control.

A high-titer scalable Chinese hamster ovary transient expression platform for production of biotherapeutics.

Transient gene expression (TGE) in Chinese hamster ovary (CHO) cells offers a route to accelerate biologics development by delivering material weeks to months earlier than what is possible with conventional cell line development. However, low productivity, inconsistent product quality profiles, and scalability challenges have prevented its broader adoption. In this study, we develop a scalable CHO-based TGE system achieving 1.9 g/L of monoclonal antibody in an unmodified host. We integrated continuous flow-electroporation and alternate tangential flow (ATF) perfusion to enable an end-to-end closed system from N-1 perfusion to fed-batch 50-L bioreactor production. Optimization of both the ATF operation for three-in-one application-cell growth, buffer exchange, and cell mass concentration-and the flow-electroporation process, led to a platform for producing biotherapeutics using transiently transfected cells. We demonstrate scalability up to 50-L bioreactor, maintaining a titer over 1 g/L. We also show comparable quality between both transiently and stably produced material, and consistency across batches. The results confirm that purity, charge variants and N-glycan profiles are similar. Our study demonstrates the potential of CHO-based TGE platforms to accelerate biologics process development timelines and contributes evidence supporting its feasibility for manufacturing early clinical material, aiming to strengthen endorsement for TGE's wider implementation.

Scalable process development of NK and CAR-NK expansion in a closed bioreactor.

Production of large amounts of functional NK and CAR-NK cells represents one of the bottlenecks for NK-based immunotherapy. In this study, we developed a large-scale, reliable, and practicable NK and CAR-NK production using G-Rex 100M bioreactors, which depend on a gas-permeable membrane technology. This system holds large volumes of medium with enhanced oxygen delivery, creating conditions conducive to large-scale PBNK and CAR-NK expansions for cancer therapy. Both peripheral blood NK cells (PBNKs) and CAR-NKs expanded in these bioreactors retained similar immunophenotypes and exhibited comparable cytotoxicity towards hepatocellular carcinoma (HCC) cells akin to that of NK and CAR-NK cells expanded in G-Rex 6 well bioreactors. Importantly, cryopreservation minimally affected the cytotoxicity of NK cells expanded using the G-Rex 100M bioreactors, establishing a robust platform for scaled-up NK and CAR-NK cell production. This method is promising for the development of "off-the-shelf" NK cells, supporting the future clinical implementation of NK cell immunotherapy.

Treatment of Synthetic Wastewater Containing Polystyrene (PS) Nanoplastics by Membrane Bioreactor (MBR): Study of the Effects on Microbial Community and Membrane Fouling.

The persistent presence of micro- and nanoplastics (MNPs) in aquatic environments, particularly via effluents from wastewater treatment plants (WWTPs), poses significant ecological risks. This study investigated the removal efficiency of polystyrene nanoplastics (PS-NPs) using a lab-scale aerobic membrane bioreactor (aMBR) equipped with different membrane types: microfiltration (MF), commercial ultrafiltration (c-UF), and recycled ultrafiltration (r-UF) membranes. Performance was assessed using synthetic urban wastewater spiked with PS-NPs, focusing on membrane efficiency, fouling behavior, and microbial community shifts. All aMBR systems achieved high organic matter removal, exceeding a 97% COD reduction in both the control and PS-exposed reactors. While low concentrations of PS-NPs did not significantly impact the sludge settleability or soluble microbial products initially, a higher accumulation increased the carbohydrate concentrations, indicating a protective bacterial response. The microbial community composition also adapted over time under polystyrene stress. All membrane types exhibited substantial NP removal; however, the presence of nano-sized PS particles negatively affected the membrane performance, enhancing the fouling phenomena and increasing transmembrane pressure. Despite this, the r-UF membrane demonstrated comparable efficiency to c-UF, suggesting its potential for sustainable applications. Advanced characterization techniques including pyrolysis gas chromatography/mass spectrometry (Py-GC/MS) were employed for NP detection and quantification.

Fibronectin Functionalization: A Way to Enhance Dynamic Cell Culture on Alginate/Hydroxyapatite Scaffolds.

Bone defects are a global health concern; bone tissue engineering (BTE) is the most promising alternative to reduce patient morbidity and overcome the inherent drawbacks of autograft and allograft bone. Three-dimensional scaffolds are pivotal in this field due to their potential to provide structural support and mimic the natural bone microenvironment. Following an already published protocol, a 3D porous structure consisting of alginate and hydroxyapatite was prepared after a gelation step and a freezing-drying step. Despite the frequent use of alginate in tissue regeneration, the biological inertness of this polysaccharide hampers proper cell colonization and proliferation. Therefore, the purpose of this work was to enhance the biological properties by promoting the interaction and adhesion between cells and biomaterial with the use of Fibronectin. This extracellular matrix protein was physically adsorbed on the scaffold, and its presence was evaluated with environmental scanning electron microscopy (eSEM) and the Micro-Bicinchoninic Acid (µBCA) protein assay. The MG-63 cell line was used for both static and dynamic (i.e., in bioreactor) 3D cell culturing on the scaffolds. The use of the bioreactor allowed for a better exchange of nutrients and oxygen and a better removal of cell catabolites from the inner portion of the construct, mimicking the physiological environment. The functionalized scaffolds showed an improvement in cell proliferation and colonization compared to non-functionalized ones; the effect of the addition of Fibronectin was more evident in the dynamic culturing conditions, where the cells clearly adhered on the surface of functionalized scaffolds.

Isolation and Identification of Naphthalene-Degrading Bacteria and its Application in a Two-phase Partitioning Bioreactor.

Naphthalene is a persistent environmental pollutant for its potential teratogenic, carcinogenic and mutagenic effects. In this study, 10 strains of bacteria capable of degrading naphthalene were isolated from crude-oil contaminated soil. Among them, Pseudomonas plecoglossicida 2P exhibited prominent growth with 1000 mg/L naphthalene as the sole carbon source and degraded 94.15% of naphthalene in 36 h. Whole genome sequencing analysis showed that P. plecoglossicida 2P had a total of 22 genes related to naphthalene degradation, of which 8 genes were related to the salicylic acid pathway only, 5 genes were related to the phthalic acid pathway only, 8 genes were common in both the salicylic acid and phthalic acid pathways, and 1 gene was related to the gentisic acid pathway. P. plecoglossicida 2P was applied in a two-phase partition bioreactor (TPPB) to degrade naphthalene in wastewater. The optimal operating conditions of the reactor were obtained through response surface optimization: initial naphthalene concentration (C0) =1600 mg/L, bacterial liquid concentration (OD600) = 1.3, and polymer-to-wastewater mass ratio (PWR) = 2%. Under these conditions, the naphthalene degradation rate was 98.36% at 24 h. The degradation kinetics were fitted using the Haldane equation with a high coefficient of determination (R2=0.94). The present study laid foundations for naphthalene degradation mechanism of genus Pseudomonas and its potential application in TPPB.

Analytical and computational studies predict negligible risk of cell death from eddy generation off flat surfaces in cell culture flow systems.

Cell-based therapies represent the current frontier of biomedical innovations, with the technologies required underpinning treatments as broad as CAR-T cell therapies, stem cell treatments, genetic therapies and mRNA manufacture. A key bottleneck in the manufacturing process for each of these lies in the expansion of cells within a bioreactor vessel, requiring by far the greatest share of time for what are often time-critical therapies. While various designs, culture feeding and mixing methods are employed in these bioreactors, a common concern among manufacturers and researchers lies in whether shear stresses generated by culture media flow will damage cells and inhibit expansion. This study develops an analytical tool to link macro-scale measures of flow to risk of cell death using relationships with eddy size and dissipation rates, from eddies generated off flat surfaces. This analytical tool was then employed using computational fluid dynamics (CFD) to replicate a range of generic bioreactor geometries and flow conditions. We found that no combination of flow condition or design parameter was predicted by the tool to cause cell death within eddies, indicating negligible risk of cell death due to eddy formation within cell culture systems. While this requires experimental validation, and does not apply when cells are expanded using microcarriers, this tool nonetheless provides reassurance and accessible prediction of bioreactor design parameters that could result in cell death. Finally, our findings show that bioreactor design can be tailored such that the shear stress stimulation of cells can be selectively altered through small changes in flow rate.

Simulation of anaerobic biodegradation process in a tubular bioreactor with a biofilm layer: Steady-state and unsteady-state conditions.

In this paper, anaerobic biodegradation process in a tubular bioreactor with an inner biofilm layer for steady-state and unsteady-state conditions are simulated. The effects of various parameters including bioreactor diameter, fraction of active biomass transferred to liquid phase, and residence time of the liquid on bioreactor performance are examined. Simulations indicate that decreasing diameter of bioreactor leads to increasing degree of conversion of the substrate in liquid phase and decreasing dimensionless concentration of the substrate in biofilm. With an increase in the fraction of active biomass transferred to liquid, substrate concentrations in liquid and biofilm slightly vary. Increased residence time of the liquid phase results in the degree of conversion of substrate goes up, but substrate concentration in biofilm lowers a little. In addition, it is found that biomass concentration of liquid phase is boosted with decreased bioreactor diameter and increased residence time of liquid. A proportional-integral controller is designed and the tuned parameters of K P = - 0.131 and K I = 0.02 are obtained using genetic algorithm. It is observed that controller regulate well the degree of conversion of the substrate within 120 s for both servo and regulatory modes.

Innovative fixed bed bioreactor platform: Enabling linearly scalable adherent cell biomanufacturing with real-time biomass prediction from nutrient consumption.

Scalable single-use adherent cell-based biomanufacturing platforms are essential for unlocking the full potential of cell and gene therapies. The primary objective of this study is to design and develop a novel fixed bed bioreactor platform tailored specifically for scaling up adherent cell culture. The bioreactor comprises a packed bed of vertically stacked woven polyethylene terephthalate mesh discs, sandwiched between two-fluid guide plates. Leveraging computational fluid dynamics modeling, we optimized bioreactor design to achieve uniform flow with minimal shear stress. Residence time distribution measurements demonstrated excellent flow uniformity with plug flow characteristics. Periodic media sampling coupled with offline analysis revealed minimal gradients of crucial metabolites (glucose, glutamine, lactate, and ammonia) across the bioreactor during cell growth. Furthermore, the bioreactor platform demonstrated high performance in automated cell harvesting, with ≈96% efficiency and ≈98% viability. It also exhibited linear scalability in both operational parameters and performance for cell culture and adeno-associated virus vector production. We developed mathematical models based on oxygen uptake rates to accurately predict cell growth curves and estimate biomass in real-time. This study demonstrates the effectiveness of the developed fixed-bed bioreactor platform in enabling scalable adherent cell-based biomanufacturing with high productivity and process control.

Stratified structure of membrane clogs observed in full scale cross-flow tubular ultrafiltration (UF) system: Fouling characterization and a proposed two-stage formation mechanism.

Membrane fouling remains a significant challenge in wastewater treatment, hindering both efficiency and lifespan. This study reports a distinct phenomenon of stratified membrane clogging observed in a full-scale cross-flow tubular ultrafiltration (UF) system treating sludge anaerobic digestion (AD) reject water. The distinct stratified structure, comprising inner and outer layers within the cake layer, has not been previously described. This research involved characterizing the filtration performance, analyzing membrane clog composition, and proposing a two-stage formation mechanism for the stratified clogs. It was revealed that higher inorganic and lower organic content in the outer layer compared to the inner layer. Acid and alkali treatments demonstrated the effectiveness of combined cleaning strategies. A mathematical model was developed to determine the critical conditions for stratified clog formation, influenced by membrane flux and cross-flow velocity (CFV). It is proposed that outer layer forms through long-term selective deposition, while the inner layer results from short-term dewatering within limited tubular space. High CFV (>2.5 m/s) prevents inner layer formation. Critical conditions for stratification occur at a flux of 18 L/m2/h with a CFV of 0.1 m/s or 65 L/m2/h with a CFV of 0.35 m/s. This study contributes a novel understanding of stratified membrane clogging, proposing a two-stage formation mechanism and identifying critical conditions, which provides insights for effective fouling control strategies and maintenance of operational efficiency for membrane systems.

Study on membrane fouling mechanisms and mitigation strategies in a pilot-scale anaerobic membrane bioreactor (P-AnMBR) treating digestate.

Anaerobic Membrane Bioreactor (AnMBR) are employed for solid-liquid separation in wastewater treatment, enhancing process efficiency of digestion systems treating digestate. However, membrane fouling remains a primary challenge. This study operated a pilot-scale AnMBR (P-AnMBR) to treat high-concentration organic digestate, investigating system performance and fouling mechanisms. P-AnMBR operation reduced acid-producing bacteria and increased methane-producing bacteria on the membrane, preventing acid accumulation and ensuring stable operation. The P-AnMBR effectively removed COD and VFA, achieving removal rates of 82.3 % and 92.0 %, respectively. Higher retention of organic nitrogen and lower retention of ammonia nitrogen were observed. The membrane fouling consisted of organic substances (20.3 %), predominantly polysaccharides, and inorganic substances (79.7 %), primarily Mg ions (10.1 %) and Ca ions (4.5 %). To reduce the increased transmembrane pressure (TMP) caused by fouling (a 10.6-fold increase in filtration resistance), backwash frequency experiment was conducted. It revealed a 30-min backwash frequency minimized membrane flux decline, facilitating recovery to higher flux levels. The water produced amounted to 70.3 m³ over 52 days. The research provided theoretical guidance and practical support for engineering applications, offering practical insights for scaling up P-AnMBR.

Perfused In Vitro Intestine Tissue Model to Evaluate the Role of Stromal and Immune Cells in Epithelial Response to Inflammatory Cues and Drug Therapies.

We establish an in vitro perfusion intestinal tissue bioreactor system tailored to study drug responses related to inflammatory bowel disease (IBD). The system includes key components including multiple human intestinal cell types (colonoids, myofibroblasts, and macrophages), a three-dimensional (3D) intestinal architecture, and fluid flow. Inclusion of myofibroblasts resulted in increased secretion of cytokines such as glypican-1 (GCP-1), granulocyte-macrophage colony-stimulating factor (GM-CSF), and interleukin 1-α (IL-1α), whereas inclusion of macrophages resulted in increased secretion of monocyte chemoattractant proteins (MCPs) demonstrating a significant role of both stromal and immune cell types in intestinal inflammation. The system is responsive to drug treatments, as reflected in the reduction of pro-inflammatory cytokine production in tissue in some treatment scenarios. While future studies are needed to evaluate more nuanced responses in an IBD context, the present study demonstrates the ability to establish a 3D intestinal model with multiple relevant cell types and flow that is responsive to both inflammatory cues and various drug treatment options.

Enhancing the membrane bioreactor efficiency: The impact of rotating membrane modules and aeration strategies on the transmembrane pressure.

The study analyses the performance of a pilot plant using a rotating hollow fibre (HF) membrane bioreactor system. The experiments evaluated the effect of operational parameters such as rotational speed, aeration strategies, and maintenance cleaning (MC) procedures on the efficiency of the system, in particular transmembrane pressure (TMP) and filtrate quality. The results indicate that the rotating membrane module reduces TMP increase and can operate for 48 days with satisfactory performance, even without aeration. This has the potential to significantly improve efficiency, resulting in significant energy savings. In addition, two MC methods, clean in air and clean in place, were tested and found to be efficient for weekly MC. It was observed that operating without aeration during colder seasons may not be effective. Therefore, adaptive strategies are needed to address seasonal temperature variations.

Development of Stephania tetrandra S. MOORE hairy root culture process for tetrandrine production.

Tetrandrine, a bioactive active compound mainly found in the roots of Stephania tetrandra, exhibits various pharmacological properties. In vitro hairy root (HR) culture may serve as a promising solution for the extraction of tetrandrine, overcoming the limitations of natural cultivation. The present study describes the consistent production of tetrandrine from S. tetrandra hairy roots induced by different strains of Agrobacterium rhizogenes. Cultivation in woody plant medium (WPM) resulted in the highest HR biomass (0.056 g/petri-dish) and tetrandrine content (7.28 mg/L) as compared to other media. The maximum HR biomass (6.95 g dw/L) and tetrandrine production (68.69 mg/L) were obtained in the fifth week of cultivation. The presence of ammonium nitrate (800 mg/L), calcium nitrate (1156 mg/L), sucrose (20 g/L) and casein (2 g/L) enhanced the tetrandrine production. Moreover, the fed-batch cultivation demonstrated that the NH4NO3 (1200 mg/L) was an important growth limiting factor that yielded the highest tetrandrine amount (119.59 mg/L). The cultivation of hairy roots in a mist trickling bioreactor for eight weeks was less (26.24 mg/L) than in the flask. Despite a lower tetrandrine yield observed in bioreactors compared to flask cultures, refining the growth medium and fine-tuning bioreactor operations hold promise for boosting tetrandrine yield.

Dynamic three-dimensional coculture model: The future of tissue engineering applied to the peripheral nervous system.

Traumatic injuries to the peripheral nervous system (PNI) can lead to severe consequences such as paralysis. Unfortunately, current treatments rarely allow for satisfactory functional recovery. The high healthcare costs associated with PNS injuries, worker disability, and low patient satisfaction press for alternative solutions that surpass current standards. For the treatment of injuries with a deficit of less than 30 mm to bridge, the use of synthetic nerve conduits (NGC) is favored. However, to develop such promising therapeutic strategies, in vitro models that more faithfully mimic nerve physiology are needed. The absence of a clinically scaled model with essential elements such as a three-dimension environment and dynamic coculture has hindered progress in this field. The presented research focuses on the development of an in vitro coculture model of the peripheral nervous system (PNS) involving the use of functional biomaterial which microstructure replicates nerve topography. Initially, the behavior of neuron-derived cell lines (N) and Schwann cells (SC) in contact with a short section of biomaterial (5 mm) was studied. Subsequent investigations, using fluorescent markers and survival assays, demonstrated the synergistic effects of coculture. These optimized parameters were then applied to longer biomaterials (30 mm), equivalent to clinically used NGC. The results obtained demonstrated the possibility of maintaining an extended coculture of SC and N over a 7-day period on a clinically scaled biomaterial, observing some functionality. In the long term, the knowledge gained from this work will contribute to a better understanding of the PNS regeneration process and promote the development of future therapeutic approaches while reducing reliance on animal experimentation. This model can be used for drug screening and adapted for personalized medicine trials. Ultimately, this work fills a critical gap in current research, providing a transformative approach to study and advance treatments for PNS injuries.

A novel Anaerobic Cathodic Dynamic Membrane Bioreactor (AnCDMBR) for efficient mitigating fouling and recovering bioenergy from municipal wastewater.

Concerns regarding membrane fouling and suboptimal bioenergy recovery have constrained the implementation of anaerobic membrane bioreactor (AnMBR) for treating low-strength municipal wastewater. This study presents a novel anaerobic cathodic dynamic membrane bioreactor (AnCDMBR) designed to address these challenges. A self-formed cathodic dynamic membrane (CDM) on inexpensive carbon cloth was developed to function as both a membrane and biocathode to achieve dual-function effects of mitigating membrane fouling and accelerating organics conversion. Compared with common dynamic membrane (1.52 kPa/d) and commercial membranes (7.52 kPa/d), the developed CDM presented a significantly reduced fouling rate (1.02 kPa/d), exhibiting the potential as a substitute for high-cost conductive membranes. Furthermore, efficient and stable biomethanation occurred in AnCDMBR with a superior methane yield rate of 0.26 L-CH4/g-COD (CH4 content > 95 %), which was 1.42 times higher than the control, linked to the higher activities of microbial metabolism and methanogenic-related key enzymes. Further analysis revealed that electrostimulation-induced niche differentiation of microbiota regulated interspecies interactions between electroactive microorganisms and complex anaerobic digestion microbiomes, facilitating organic matter conversion to methane and leading to superior bioenergy recovery. This study offered a new strategy for effectively mitigating fouling and recovering bioenergy from low-strength wastewater, potentially expanding the application of AnMBRs.

A Column Study: Impact of Redox, Substrate Composition and Exposure Order on Toluene and Benzene Biodegradation and Microbial Communities.

Aromatic compounds persist as hazardous contaminants in both aquatic and terrestrial environments, needing rapid and effective remediation strategies. This study evaluated toluene and benzene biodegradation under sulfate and nitrate-reducing conditions in column experiments, utilizing aquifer sediments from a contaminated site. Over a period of 36 weeks, four glass columns were operated simultaneously in an alternating flow-batch regime. Each column received either nitrate or sulfate as an electron acceptor while being exposed to different substrate compositions in varied exposure orders. A redox dependent contaminant removal efficiency was observed, with toluene removal efficiency at 81% under sulfate and 55% under nitrate-reducing conditions, and benzene removal efficiency approximately at 44% and 59%, respectively, within 4 to 6 weeks. The rapid removal under anaerobic conditions was attributed to the alternating flow-batch regime, allowing biomass growth in batch mode, and applying selection pressure to non-specific biodegraders during flow regime. Toluene removal remained unaffected by benzene's presence but exhibited slight inhibition in the presence of an aromatic mixture composed of BTEX, indene, indane, and naphthalene. Benzene removal efficiency dropped to 8% in the presence of toluene but remained unaffected by the mixture. Pre-exposure to a single compound enhanced breakdown efficiency when further faced with a more complex mixture. Additionally, beta-diversity analysis conducted on the four columns revealed distinct microbial community clustering between sulfate and nitrate-reducing conditions, emphasizing the determining role of redox conditions. Findings of this study can be used to develop more effective pollution cleanup strategies, specifically targeting parameters like redox conditions, substrate interactions, and pollution history, thus improving our ability to mitigate contamination across diverse environments.

Exploring Aeration Strategies for Enhanced Simultaneous Nitrification and Denitrification in Membrane Aerated Bioreactors: A Computational Approach.

In this study we employ computational methods to investigate the influence of aeration strategies on simultaneous nitrification-denitrification processes. Specifically, we explore the impact of periodic and intermittent aeration on denitrification rates, which typically lag behind nitrification rates under identical environmental conditions. A two-dimensional deterministic multi-scale model is employed to elucidate the fundamental processes governing the behavior of membrane aerated biofilm reactors (MABRs). We aim to identify key factors that promote denitrification under varying aeration strategies. Our findings indicate that the concentration of oxygen during the off phase and the duration of the off interval play crucial roles in controlling denitrification. Complete discontinuation of oxygen is not advisable, as it inhibits the formation of anaerobic heterotrophic bacteria, thereby impeding denitrification. Extending the length of the off interval, however, enhances denitrification. Furthermore, we demonstrate that the initial inoculation of the substratum (membrane in this study) influences substrate degradation under periodic aeration, with implications for both nitrification and denitrification. Comparison between continuous and periodic/intermittent aeration scenarios reveals that the latter can extend the operational cycle of MABRs. This extension is attributed to relatively low biofilm growth rates associated with non-continuous aeration strategies. Consequently, our study provides a comprehensive understanding of the intricate interplay between aeration strategies and simultaneous nitrification-denitrification in MABRs. The insights presented herein can contribute significantly to the optimization of MABR performance in wastewater treatment applications.

Orthophosphate uptake onto woodchips in denitrifying bioreactors is enhanced by anoxic-(anaerobic-)oxic cycling.

Woodchips are widely used as a low-cost and renewable organic carbon source for denitrifying biofilms in passive nutrient removal systems. One limitation of wood-based biofiltration systems is their relatively poor removal of phosphorus (P) from subsurface drainage and stormwaters, necessitating the use of additional filter media when co-treatment of nitrogen (N) and P is required. Here, we show that anoxic-oxic cycling of woodchip media, which enhances nitrate (NO3-) removal by increasing the mobilization of organic carbon from wood, also improves orthophosphate (Pi) uptake onto woodchips. Orthophosphate removal rates in flow-through woodchip columns ranged from 0 to 34.9 µg PO43- L-1 h-1 under continuously-saturated (anoxic) conditions, and increased to 17.5 to 71.9 µg PO43- L-1 h-1 in columns undergoing drying-rewetting (oxic-anoxic) cycles. The highest Pi removal efficiencies were observed in the first 20 h after reactors were re-flooded, and were concurrent with maxima in polyphosphate kinase (ppk) gene expression by the polyphosphate accumulating organisms (PAOs) Accumulibacter spp. and Pseudomonas spp. Batch experiments confirmed that anoxic-anaerobic-oxic pre-incubation conditions led to orthophosphate uptake onto woodchips as high as 74.9 ± 0.8 mg PO43-/kg woodchip, and batch tests with autoclaved woodchips demonstrated that Pi uptake was due to biological processes and not adsorption. NO3- removal in batch tests was also greatest under oxic incubation conditions, attributed to greater carbon availability in hypoxic to anoxic zones in woodchip biofilms. While further research is needed to elucidate the mechanisms controlling enhanced Pi uptake by woodchip biofilms under anoxic-(anaerobic-)oxic cycling, these results suggest a role for enhanced Pi uptake by PAOs in a nature-based system for treatment of nonpoint source nutrients.