Membrane modification in enhancement of virus removal: A critical review.

Many waterborne diseases are related with viruses, and COVID-19 worldwide has raised the concern of virus security in water into the public horizon. Compared to other conventional water treatment processes, membrane technology can achieve satisfactory virus removal with fewer chemicals, and prevent the outbreaks of viruses to a maximal extent. Researchers developed new modification methods to improve membrane performance. This review focused on the membrane modifications that enhance the performance in virus removal. The characteristics of viruses and their removal by membrane filtration were briefly generalized, and membrane modifications were systematically discussed through different virus removal mechanisms, including size exclusion, hydrophilic and hydrophobic interactions, electronic interactions, and inactivation. Advanced functional materials for membrane modification were summarized based on their nature. Furthermore, it is suggested that membranes should be enhanced through different mechanisms mainly based on their ranks of pore size. The current review provided theoretical support regarding membrane modifications in the enhancement of virus removal and avenues for practical application.

A soft-sensor approach for predicting an indicator virus removal efficiency of a pilot-scale anaerobic membrane bioreactor (AnMBR).

The anaerobic membrane bioreactor (AnMBR) is a promising technology for not only water reclamation but also virus removal; however, the virus removal efficiency of AnMBR has not been fully investigated. Additionally, the removal efficiency estimation requires datasets of virus concentration in influent and effluent, but its monitoring is not easy to perform for practical operation because the virus quantification process is generally time-consuming and requires specialized equipment and trained personnel. Therefore, in this study, we aimed to identify the key, monitorable variables in AnMBR and establish the data-driven models using the selected variables to predict virus removal efficiency. We monitored operational and environmental conditions of AnMBR in Sendai, Japan and measured virus concentration once a week for six months. Spearman's rank correlation analysis revealed that the pH values of influent and mixed liquor suspended solids (MLSS) were strongly correlated with the log reduction value of pepper mild mottle virus, indicating that electrostatic interactions played a dominant role in AnMBR virus removal. Among the candidate models, the random forest model using selected variables including influent and MLSS pH outperformed the others. This study has demonstrated the potential of AnMBR as a viable option for municipal wastewater reclamation with high microbial safety.

A mass balance approach for quantifying the role of natural decay and fate mechanisms on SARS-CoV-2 genetic marker removal during water reclamation.

The recent SARS-CoV-2 outbreak yielded substantial data regarding virus fate and prevalence at water reclamation facilities (WRFs), identifying influential factors as natural decay, adsorption, light, pH, salinity, and antagonistic microorganisms. However, no studies have quantified the impact of these factors in full scale WRFs. Utilizing a mass balance approach, we assessed the impact of natural decay and other fate mechanisms on genetic marker removal during water reclamation, through the use of sludge and wastewater genetic marker loading estimates. Results indicated negligible removal of genetic markers during P/PT (primary effluent (PE) p value: 0.267; preliminary and primary treatment (P/PT) accumulation p value: 0.904; and thickened primary sludge (TPS) p value: 0.076) indicating no contribution of natural decay and other fate mechanisms toward removal in P/PT. Comparably, adsorption and decomposition was found to be the dominant pathway for genetic marker removal (thickened waste activated sludge (TWAS) log loading 9.75 log10 GC/day); however, no estimation of log genetic marker accumulation could be carried out due to high detections in TWAS. PRACTITIONER POINTS: The mass balance approach suggested that the contribution of natural decay and other fate mechanisms to virus removal during wastewater treatment are negligible compared with adsorption and decomposition in P/PT (p value: 0.904). During (P/PT), a higher viral load remained in the (PE) (14.16 log10 GC/day) compared with TPS (13.83 log10 GC/day); however, no statistical difference was observed (p value: 0.280) indicting that adsorption/decomposition most probably did not occur. In secondary treatment (ST), viral genetic markers in TWAS were consistently detected (13.41 log10 GC/day) compared with secondary effluent (SE), indicating that longer HRT and the potential presence of extracellular polymeric substance-containing enriched biomass enabled adsorption/decomposition. Estimations of total solids and volatile solids for TPS and TWAS indicated that adsorption affinity was different between solids sampling locations (p value: <0.0001).

Removal of RNA viruses from swine wastewater using anaerobic membrane bioreactor: Performance and mechanisms.

The effective removal of viruses from swine wastewater using anaerobic membrane bioreactor (AnMBR) is vital to ecological safety. However, most studies have focused only on disinfectants, whereas the capabilities of the treatment process have not been investigated. In this study, the performance and mechanism of an AnMBR in the removal of porcine hepatitis E virus (HEV), porcine kobuvirus (PKoV), porcine epidemic diarrhea virus (PEDV), and transmissible gastroenteritis coronavirus (TGEV) are systematically investigated. The results show that the AnMBR effectively removes the four viruses, with average removal efficiencies of 1.62, 3.05, 2.41, and 1.34 log for HEV, PKoV, PEDV and TGEV, respectively. Biomass adsorption contributes primarily to the total virus removal in the initial stage of reactor operation, with contributions to HEV and PKoV removal exceeding 71.7 % and 68.2 %, respectively. When the membrane is fouled, membrane rejection dominated virus removal. The membrane rejection contribution test shows the significant contribution of membrane pore foulants (23-76 %). Correlation analysis shows that the surface characteristics and size differences of the four viruses contribute primarily to their different effects on biomass adsorption and membrane rejection. This study provides technical guidance for viral removal during the treatment of high-concentration swine wastewater using an AnMBR.

Viral clearance capability of monoclonal antibody purification.

Viral clearance steps are routinely included in monoclonal antibody purification processes to safeguard product from potential virus contamination. These steps are often experimentally studied using product-specific feeds and parameters for each project to demonstrate viral clearance capability. However, published evidence suggests that viral clearance capability of many of these steps are not significantly impacted by variations in feed material or process parameter within commonly used ranges. The current investigation confirms robust retrovirus inactivation by low pH treatment and parvovirus removal by second-generation virus filters, independent to individual antibody molecules. Our results also reveal robust retrovirus removal by flowthrough anion exchange chromatography, inside the limits of protein load and host cell protein content. The cumulative viral clearance capability from these steps leads to an excess clearance safety factor of 10,000-fold for endogenous retrovirus-like particles. These results further justify the use of prior knowledge-based modular viral clearance estimation as opposed to repetitive experimentation.

Size-based analysis of virus removal filter fouling using fractionated protein aggregates.

Fouling by protein aggregates reduces virus removal filter performance. In the present study, we investigated the effects of different-sized protein aggregates on fouling and aggregate retention in order to better understand the fouling mechanisms. Human immunoglobulin G was denatured by heating to produce aggregates of various sizes and then fractionated by size exclusion chromatography into different-sized aggregates with a narrow size distribution. The fractionated aggregates were filtered on Planova 20N, a virus removal filter known for its stable filtration capability. Analysis of flux behavior demonstrated different flux decrease patterns for different-sized aggregates. Observation of aggregate retention by staining revealed that larger aggregates were captured closer to the inner surface of the membrane while smaller aggregates penetrated farther into the membrane. These findings demonstrate that Planova 20N has a gradient structure with decreasing pore size from the inner to the outer surface of the membrane. This structure minimizes fouling and enables stable filtration by protecting the smaller pores located closer to the outer surface from clogging by large aggregates. Applying the predominant clogging models to the present filtrations revealed that clogging behavior transitioned from complete blocking to cake filtration as filtration progressed. In this combination model, after a certain number of pores are blocked by complete blocking, newly arrived aggregates begin to accumulate on previously captured aggregates, generating cake between capture layers within the membrane. Application of the approaches described here will facilitate elucidation of membrane fouling and virus removal mechanisms.

Integration of Planova filters in manufacturing processes of biologicals improve the virus safety effectively: A review of publicly available data.

The capacity to remove viruses by Planova filters produced by Asahi Kasei, primarily by small virus-retentive filters, were compiled from data in peer-reviewed publications and, partly, publicly available data from presentations at conferences (Planova workshops). Data from more than 100 publications and presentations at conferences covering Planova filters were assessed. The data were grouped according to the different virus filters regarding mean pore sizes and viruses of different sizes for plasma and cell culture derived products. Planova 15N and 20N filters removed parvoviruses below the limit of detection of viruses in the filtrate in approx. 50% of all studies and mean LRFs (log reduction factors) for viruses detected in the filtrate were above 4, demonstrating effective parvovirus reduction. Parvovirus removal capacity increased for Planova BioEX filters as well as for 2 Planova 20N in series. Large viruses as retroviruses (e.g., HIV and MuLV), herpesviruses, flaviviruses and togaviruses were removed effectively by Planova 15N, 20N and BioEX filters and also by Planova 35N filters. Flow interruption, transmembrane pressure, volume and protein concentration per filter area had had no substantial impact on virus removal capacity at manufacturing specification. In conclusion, the incorporation of Planova filters in manufacturing processes of biologicals remove, depending on the filter pore size, small and large viruses from the feed stream reliably. This virus reduction step with an orthogonal mechanism integrated in the manufacturing processes of biologicals, based primarily on size exclusion of viruses, improves the virus safety of these biopharmaceutical products considerably.

Evaluating the viral clearance ability of continuous monoclonal antibody purification steps, in order to inactivate and/or remove four model viruses.

Background and Objectives: Viral clearance studies are an essential part of a manufacturer's plan to ensure the safety of an injectable biologic product. In this way, viral safety is a critical quality attribute for biologics such as monoclonal antibodies (Mabs). Evaluation of virus purification by downstream processes is a key component of risk mitigation. In this study, the capability of continuous monoclonal antibody purification steps was evaluated in the process of instant monoclonal antibody purification in different stages of purification, and the amount of reduction or inactivation of each step was determined. Materials and Methods: Four enveloped and non-enveloped viral models VSV, Reovirus, EMCV, and HSV1 were used for spiking in selected samples in the designated tests, to have a comprehensive examination of the ability to clear the virus such as the type of genetic material, chemical resistance, and particle size. A TCID50 and qPCR methods were used to measure viral reduction. Two cell lines, Vero (African green monkey kidney) and L929 (Mouse fibroblast) were used for 4 model viruses propagation. The steps that were evaluated included 4 steps monoclonal antibody purification; cation exchange chromatography, acidic pH treatment, affinity chromatography, and nanofiltration. Results: The nano-filter stage showed the highest viral reduction and cation exchange chromatography showed the lowest reduction. The cumulative decrease using TCID50 is equal to 19.27 [log10] for all steps and for the qPCR method is equal to 12.47 [log10] in three steps of nano-filter, affinity chromatography, and ion exchange chromatography. Conclusion: The overall average reduction coefficient for all four model viruses is significantly high, which indicates the high capacity of the monoclonal antibody production process in inactivating and removing viruses leads to reducing the load of all four model viruses.

Product safety aspects of plant molecular farming.

Plant molecular farming (PMF) has been promoted since the 1990s as a rapid, cost-effective and (most of all) safe alternative to the cultivation of bacteria or animal cells for the production of biopharmaceutical proteins. Numerous plant species have been investigated for the production of a broad range of protein-based drug candidates. The inherent safety of these products is frequently highlighted as an advantage of PMF because plant viruses do not replicate in humans and vice versa. However, a more nuanced analysis of this principle is required when considering other pathogens because toxic compounds pose a risk even in the absence of replication. Similarly, it is necessary to assess the risks associated with the host system (e.g., the presence of toxic secondary metabolites) and the production approach (e.g., transient expression based on bacterial infiltration substantially increases the endotoxin load). This review considers the most relevant host systems in terms of their toxicity profile, including the presence of secondary metabolites, and the risks arising from the persistence of these substances after downstream processing and product purification. Similarly, we discuss a range of plant pathogens and disease vectors that can influence product safety, for example, due to the release of toxins. The ability of downstream unit operations to remove contaminants and process-related toxic impurities such as endotoxins is also addressed. This overview of plant-based production, focusing on product safety aspects, provides recommendations that will allow stakeholders to choose the most appropriate strategies for process development.

Predicting microbial water quality in on-site water reuse systems with online sensors.

Widespread implementation of on-site water reuse is hindered by the limited availability of monitoring approaches that ensure microbial quality during operation. In this study, we developed a methodology for monitoring microbial water quality in on-site water reuse systems using inexpensive and commercially available online sensors. An extensive dataset containing sensor and microbial water quality data for six of the most critical types of disruptions in membrane bioreactors with chlorination was collected. We then tested the ability of three typological machine learning algorithms - logistic regression, support-vector machine, and random forest - to predict the microbial water quality as "safe" or "unsafe" for reuse. The main criteria for model optimization was to ensure a low false positive rate (FPR) - the percentage of safe predictions when the actual condition is unsafe - which is essential to protect users health. This resulted in enforcing a fixed FPR ≤ 2%. Maximizing the true positive rate (TPR) - the percentage of safe predictions when the actual condition is safe - was given second priority. Our results show that logistic-regression-based models using only two out of the six sensors (free chlorine and oxidation-reduction potential) achieved the highest TPR. Including sensor slopes as engineered features allowed to reach similar TPRs using only one sensor instead of two. Analysis of the occurrence of false predictions showed that these were mostly early alarms, a characteristic that could be regarded as an asset in alarm management. In conclusion, the simplest algorithm in combination with only one or two sensors performed best at predicting the microbial water quality. This result provides useful insights for water quality modeling or for applications where small datasets are a common challenge and a general advantage might be gained by using simpler models that reduce the risk of overfitting, allow better interpretability, and require less computational power.

Sensor setpoints that ensure compliance with microbial water quality targets for membrane bioreactor and chlorination treatment in on-site water reuse systems.

Widespread implementation of on-site water reuse systems is hindered by the limited ability to ensure the level of treatment and protection of human health during operation. In this study, we tested the ability of five commercially available online sensors (free chlorine (FC), oxidation-reduction potential (ORP), pH, turbidity, UV absorbance at 254 nm) to predict the microbial water quality in membrane bioreactors followed by chlorination using logistic regression-based and mechanism-based models. The microbial water quality was assessed in terms of removal of enteric bacteria from the wastewater, removal of enteric viruses, and regrowth of bacteria in the treated water. We found that FC and ORP alone could predict the microbial water quality well, with ORP-based models generally performing better. We further observed that prediction accuracy did not increase when data from multiple sensors were integrated. We propose a methodology to link online sensor measurements to risk-based water quality targets, providing operation setpoints protective of human health for specific combinations of wastewaters and reuse applications. For instance, we recommend a minimum ORP of 705 mV to ensure a virus log-removal of 5, and an ORP of 765 mV for a log-removal of 6. These setpoints were selected to ensure that the percentage of events where the water is predicted to meet the quality target but it does not remains below 5%. Such a systematic approach to set sensor setpoints could be used in the development of water reuse guidelines and regulations that aim to cover a range of reuse applications with differential risks to human health.

Microporous Polyethersulfone Membranes Grafted with Zwitterionic Polymer Brushes Showing Microfiltration Permeance and Ultrafiltration Bacteriophage Removal.

Virus removal from water using microfiltration (MF) membranes is of great interest but remains challenging owing to the membranes' mean pore sizes typically being significantly larger than most viruses. We present microporous membranes grafted with polyzwitterionic brushes (N-dimethylammonium betaine) that combine bacteriophage removal in the range of ultrafiltration (UF) membranes with the permeance of MF membranes. Brush structures were grafted in two steps: free-radical polymerization followed by atom transfer radical polymerization (ATRP). Attenuated total reflection Fourier transform infrared (ATR-FTIR) and X-ray photoelectron (XPS) verified that grafting occurred at both sides of the membranes and that the grafting increased with increasing the zwitterion monomer concentration. The log reduction values (LRVs) of the pristine membrane increased from less than 0.5 LRV for T4 (∼100 nm) and NT1 (∼50 nm) bacteriophages to up to 4.5 LRV for the T4 and 3.1 LRV for the NT1 for the brush-grafted membranes with a permeance of about 1000 LMH/bar. The high permeance was attributed to a high-water fraction in the ultra-hydrophilic brush structure. The high measured LRVs of the brush-grafted membranes were attributed to enhanced bacteriophages exclusion from the membrane surface and entrapment of the ones that penetrated the pores due to the membranes' smaller mean pore-size and cross-section porosity than those of the pristine membrane, as seen by scanning electron microscopy (SEM) and measured using liquid-liquid porometry. Micro X-ray fluorescence (µ-XRF) spectrometry and nanoscale secondary ion mass spectrometry showed that 100 nm Si-coated gold nanospheres accumulated on the surface of the pristine membrane but not on the brush-coated membrane and that the nanospheres that penetrated the membranes were entrapped in the brush-grafted membrane but passed the pristine one. These results corroborate the LRVs obtained during filtration experiments and support the inference that the increased removal was due to a combined exclusion mechanism and entrapment. Overall, these microporous brush-grafted membranes show potential for use in advanced water treatment.

Ions play different roles in virus removal caused by different NOMs in UF process: Removal efficiency and mechanism analysis.

In this study, we investigated the effect of different compositions of aquatic natural organic matter (NOM) and ions on virus removal by ultrafiltration (UF). MS2 bacteriophage was used as a surrogate. Humic acid (HA) improved the MS2 removal rate from 1.95 ± 0.09 LRV to 2.40 ± 0.03 LRV at the HA dosage of 9 mg/L through the combined mechanisms of size exclusion, electrostatic repulsion and hydrophobicity. MS2 removal rate further increased to 3.10 ± 0.05 LRV by 10 mmol/L Na+ dosage and 3.19 ± 0.12 LRV by Ca2+ 1 mmol/L in the HA-containing UF system. Size exclusion turned into the dominant virus removal mechanism according to the results of the fouling model fitting and the weakening of electrostatic repulsion and hydrophobicity. The complexation of Ca2+ also played a role in MS2 removal based on the analysis of interaction force. MS2 removal rate by bovine serum albumin (BSA) was poor, which was 2.07 ± 0.06 LRV at the BSA dosage of 9 mg/L. Hydrophobicity was greatly reduced and the dominant virus removal mechanisms were size exclusion and electrostatic repulsion. 10 mmol/L Na+ in the presence of BSA deteriorated MS2 removal rate to 2.02 ± 0.07 LRV by the weakening of electrostatic repulsion, hydrophobicity and size exclusion. Electrostatic repulsion severely decreased by 1 mmol/L Ca2+ and the enhanced adsorption barrier represented competitive adsorption of Ca2+ by BSA and MS2 contributed for MS2 removal further decline (1.99 ± 0.05 LRV). Complex components in water will have different effects on virus removal due to their properties and interactions. This study can provide references for selecting more efficient water treatment methods according to the different compositions of raw water in actual water treatment applications during the UF process. Moreover, the retention of virus by UF can be predicted based on our study results.

Boronic acid-modified bacterial cellulose microspheres as packing materials for enveloped virus removal.

Viruses are the most abundant microorganisms on the earth, their existence in contaminated waters possesses a significant threat to humans. Waterborne viral infections could be fatal to sensitive population including young child, the elderly, and the immune-compromised. It is imperative to remove viruses during water treatment to better protect public health, especially in the light of evidence of detection of coronaviruses genetic fragments in raw sewage. We reported bench-scale experiments evaluating the extent and mechanisms of removal of a model virus (spring viremia of carp virus, SVCV) in water by adsorption. Microspheres made by boronic acid-modified bacterial cellulose with excellent mechanical strength were successfully fabricated as packing materials for the column to remove glycoproteins and enveloped viruses from water. The synthesized adsorbent was characterized by attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, Atomic Force Microscopy (AFM), Scanning Electron Microscopy (SEM), and Brunauer Emmett Teller (BET) measurement. The adsorption efficiency of glycoproteins was investigated by SDS-PAGE and the Broadford protein assay, while the binding capacity with the virus (spring viremia of carp virus) was monitored by cell culture to calculate the viral cytopathic effect and viral titer caused by the virus. The data obtained from the above experiments showed that ∼3-log removal of SVCV in 3 h, which significantly reduced the virus concentration from microspheres packed column. The present study provides substantial evidence to prove beyond doubt that material based on bacterial cellulose seems to have the potential for virus removal from water which can be extended to systems of significant importance.

Viral removal by column chromatography in downstream processing of monoclonal antibodies.

For monoclonal antibodies (mAbs) produced in mammalian cells, viral safety is a critical concern. The downstream process, in addition to removing other impurities, needs to ensure robust clearance (removal or inactivation) of potential endogenous and adventitious viruses. In general, Protein A and polishing chromatography steps all can provide certain level of virus removal. Chromatographic removal combined with virus inactivation and nanofiltration usually provides adequate virus clearance across the overall downstream process. This article reviews the virus clearance capability of commonly used column chromatography, with attention to possible interference of virus-mAb interaction on virus removal. In addition, the potential of using viral surrogate as a safe alternative to live virus for assessing viral clearance is briefly discussed.

Recent advances in aqueous virus removal technologies.

The COVID-19 outbreak has triggered a massive research, but still urgent detection and treatment of this virus seems a public concern. The spread of viruses in aqueous environments underlined efficient virus treatment processes as a hot challenge. This review critically and comprehensively enables identifying and classifying advanced biochemical, membrane-based and disinfection processes for effective treatment of virus-contaminated water and wastewater. Understanding the functions of individual and combined/multi-stage processes in terms of manufacturing and economical parameters makes this contribution a different story from available review papers. Moreover, this review discusses challenges of combining biochemical, membrane and disinfection processes for synergistic treatment of viruses in order to reduce the dissemination of waterborne diseases. Certainly, the combination technologies are proactive in minimizing and restraining the outbreaks of the virus. It emphasizes the importance of health authorities to confront the outbreaks of unknown viruses in the future.

Removal of MS2 and fr Bacteriophages Using MgAl2O4-Modified, Al2O3-Stabilized Porous Ceramic Granules for Drinking Water Treatment.

Point-of-use ceramic filters are one of the strategies to address problems associated with waterborne diseases to remove harmful microorganisms in water sources prior to its consumption. In this study, development of adsorption-based ceramic depth filters composed of alumina platelets was achieved using spray granulation (calcined at 800 °C). Their virus retention performance was assessed using cartridges containing granular material (4 g) with two virus surrogates: MS2 and fr bacteriophages. Both materials showed complete removal, with a 7 log10 reduction value (LRV) of MS2 up to 1 L. MgAl2O4-modified Al2O3 granules possessed a higher MS2 retention capacity, contrary to the shortcomings of retention limits in pure Al2O3 granules. No significant decline in the retention of fr occurred during filtration tests up to 2 L. The phase composition and morphology of the materials were preserved during filtration, with no magnesium or aluminum leakage during filtration, as confirmed by X-ray diffractograms, electron micrographs, and inductively coupled plasma-optical emission spectrometry. The proposed MgAl2O4-modified Al2O3 granular ceramic filter materials offer high virus retention, achieving the criterion for virus filtration as required by the World Health Organization (LRV ≥ 4). Owing to their high thermal and chemical stability, the developed materials are thus suitable for thermal and chemical-free regeneration treatments.

Moringa oleifera-derived coagulants for water treatment: Floc structure, residual organics, and performance trade-offs.

The study explored the suitability of unfractionated extracts from the seeds of the Moringa oleifera tree as a coagulant for water treatment. The coagulant was obtained by soaking crushed and sieved seeds in a low salinity aqueous solution: a simple and inexpensive alternative to conventional coagulants in settings where specialized expertise and equipment are lacking. The performance of M. oleifera-derived coagulants was quantified in terms of turbidity removal, bacteriophage clearance, concentration of residual organics, as well as meta-parameters such as floc size and fractal dimension. Treating high turbidity clay suspensions at the optimal coagulant dosage (14.7 mg(DOC)/L) and flocculation mixing conditions ([Formula: see text]= 22.4 s-1) removed > 94% of turbidity, similar to that recorded in reference tests with alum. Floc size distribution shifted to larger sizes during the first 10 min of flocculation with no change afterwards, while the floc fractal dimension, [Formula: see text], continued to increase, pointing to the gradual formation of denser ([Formula: see text]= 2.1 to 2.2), more settleable flocs. Preliminary tests with MS2 bacteriophage showed that coagulation with M. oleifera decreased the viable MS2 titre by ~ 1.3 log, which was significantly above the turbidity removal (~ 1 log). The extraction process, however, allowed a large amount of residual organics (> 78% of extracted DOC) into the treated water. Combining the coagulants with downstream filtration and adsorption, employing UV or solar disinfection, or limiting applications to non-potable reuse is suggested for mitigating the concerns related to residual DOC.

Virus removal during sewage treatment by anaerobic membrane bioreactor (AnMBR): The role of membrane fouling.

Anaerobic membrane bioreactor (AnMBR) is a low-energy and promising solution for sewage treatment. During the treatment, the fouled membrane of AnMBR is recognized as an important barrier against pathogenic viruses. Here, the role of membrane fouling of an AnMBR at room temperature in the virus removal was investigated using MS2 bacteriophage as a virus surrogate. Results revealed that the virus removal efficiency of AnMBR was in the range of 0.2 to 3.6 logs, gradually increasing with the course of AnMBR operation. Virus removal efficiency was found to be significantly correlated with transmembrane pressure (R2=0.92, p<0.01), especially in the rapid fouling stage, indicating that membrane fouling was the key factor in the virus removal. The proportion of virus decreased from 52.03% to 15.04% in the membrane foulants when membrane fouling was aggravating rapidly, yet increased from 0.74% to 21.52% in the mixed liquor. Meanwhile, the permeate flux dramatically dropped. These imply that the primary rejection mechanism of virus by membrane in the slow fouling stage is the virus adsorption onto membrane, while the sieving effect is the main reason in the rapid fouling stage. Ex-situ virus rejection test unveiled that the cake layer was the main contributor to the overall virus rejection, while the greatest resistance-specific virus rejection was provided by the organic pore blocking. This paper provides operation strategies to balance enhanced virus removal and high permeate flux by regulating the membrane fouling process.

Reduction and liquid-solid partitioning of SARS-CoV-2 and adenovirus throughout the different stages of a pilot-scale wastewater treatment plant.

Investigating waterborne viruses is of great importance to minimizing risks to public health. Viruses tend to adsorb to sludge particles from wastewater processes by electrostatic and hydrophobic interactions between virus, aquatic matrix, and particle surface. Sludge is often re-used in agriculture; therefore, its evaluation is also of great interest to public health. In the present study, a pilot scale system treating real domestic wastewater from a large city in Brazil was used to evaluate the removal, the overall reduction, and liquid-solid partitioning of human adenovirus (HAdV), the novel coronavirus (SARS-CoV-2) and fecal indicators (F-specific coliphages and E. coli). The system consists of a high-rate algal pond (HRAP) post-treating the effluent of an upflow anaerobic sludge blanket (UASB) reactor. Samples were collected from the influent and effluent of each unit, as well as from the sludge of the UASB and from the microalgae biomass in the HRAP. Pathogens and indicators were quantified by quantitative polymerase chain reaction (qPCR) (for HAdV), qPCR with reverse transcription (RTqPCR) (for SARS-CoV-2), the double agar plaque assay (for coliphages), and the most probable number (MPN) method (for E. coli). The removal and overall reduction of HAdV and SARS-CoV-2 was greater than 1-log10. Almost 60% of remaining SARS-CoV-2 RNA and more than 70% of remaining HAdV DNA left the system in the sludge, demonstrating that both viruses may have affinity for solids. Coliphages showed a much lower affinity to solids, with only 3.7% leaving the system in the sludge. The system performed well in terms of the removal of organic matter and ammoniacal nitrogen, however tertiary treatment would be necessary to provide further pathogen reduction, if the effluent is to be reused in agriculture. To our knowledge, this is the first study that evaluated the reduction and partitioning of SARS-CoV-2 and HAdV through the complete cycle of a wastewater treatment system consisting of a UASB reactor followed by HRAPs.