Impact of harmful algal bloom severity on bacterial communities in a full-scale biological filtration system for drinking water treatment.

The occurrence of harmful algal blooms (HABs) in freshwater environments has been expanded worldwide with growing frequency and severity. HABs can pose a threat to public water supplies, raising concerns about safety of treated water. Many studies have provided valuable information about the impacts of HABs and management strategies on the early-stage treatment processes (e.g., pre-oxidation and coagulation/flocculation) in conventional drinking water treatment plants (DWTPs). However, the potential effect of HAB-impacted water in the granular media filtration has not been well studied. Biologically-active filters (BAFs), which are used in drinking water treatment and rely largely on bacterial community interactions, have not been examined during HABs in full-scale DWTPs. In this study, we assessed the bacterial community structure of BAFs, functional profiles, assembly processes, and bio-interactions in the community during both severe and mild HABs. Our findings indicate that bacterial diversity in BAFs significantly decreases during severe HABs due to the predominance of bloom-associated bacteria (e.g., Spingopyxis, Porphyrobacter, and Sphingomonas). The excitation-emission matrix combined with parallel factor analysis (EEM-PARAFAC) confirmed that filter influent affected by the severe HAB contained a higher portion of protein-like substances than filter influent samples during a mild bloom. In addition, BAF community functions showed increases in metabolisms associated with intracellular algal organic matter (AOM), such as lipids and amino acids, during severe HABs. Further ecological process and network analyses revealed that severe HAB, accompanied by the abundance of bloom-associated taxa and increased nutrient availability, led to not only strong stochastic processes in the assembly process, but also a bacterial community with lower complexity in BAFs. Overall, this study provides deeper insights into BAF bacterial community structure, function, and assembly in response to HABs.

Spatial and Temporal Variability of Saxitoxin-Producing Cyanobacteria in U.S. Urban Lakes.

Harmful cyanobacterial blooms (HCBs) are of growing global concern due to their production of toxic compounds, which threaten ecosystems and human health. Saxitoxins (STXs), commonly known as paralytic shellfish poison, are a neurotoxic alkaloid produced by some cyanobacteria. Although many field studies indicate a widespread distribution of STX, it is understudied relative to other cyanotoxins such as microcystins (MCs). In this study, we assessed eleven U.S. urban lakes using qPCR, sxtA gene-targeting sequencing, and 16S rRNA gene sequencing to understand the spatio-temporal variations in cyanobacteria and their potential role in STX production. During the blooms, qPCR analysis confirmed the presence of the STX-encoding gene sxtA at all lakes. In particular, the abundance of the sxtA gene had a strong positive correlation with STX concentrations in Big 11 Lake in Kansas City, which was also the site with the highest quantified STX concentration. Sequencing analysis revealed that potential STX producers, such as Aphanizomenon, Dolichospermum, and Raphidiopsis, were present. Further analysis targeting amplicons of the sxtA gene identified that Aphanizomenon and/or Dolichospermum are the primary STX producer, showing a significant correlation with sxtA gene abundances and STX concentrations. In addition, Aphanizomenon was associated with environmental factors, such as conductivity, sulfate, and orthophosphate, whereas Dolichospermum was correlated with temperature and pH. Overall, the results herein enhance our understanding of the STX-producing cyanobacteria and aid in developing strategies to control HCBs.

Assessing residential activity in a home plumbing system simulator: monitoring the occurrence and relationship of major opportunistic pathogens and phagocytic amoebas.

Opportunistic premise plumbing pathogens (OPPPs) have been detected in buildings' plumbing systems causing waterborne disease outbreaks in the United States. In this study, we monitored the occurrence of OPPPs along with free-living amoeba (FLA) and investigated the effects of residential activities in a simulated home plumbing system (HPS). Water samples were collected from various locations in the HPS and analyzed for three major OPPPs: Legionella pneumophila, nontuberculous mycobacterial species (e.g., Mycobacterium avium, M. intracellulare, and M. abscessus), and Pseudomonas aeruginosa along with two groups of amoebas (Acanthamoeba and Vermamoeba vermiformis). A metagenomic approach was also used to further characterize the microbial communities. Results show that the microbial community is highly diverse with evidence of spatial and temporal structuring influenced by environmental conditions. L. pneumophila was the most prevalent pathogen (86% of samples), followed by M. intracellulare (66%) and P. aeruginosa (21%). Interestingly, M. avium and M. abscessus were not detected in any samples. The data revealed a relatively low prevalence of Acanthamoeba spp. (4%), while V. vermiformis was widely detected (81%) across all the sampling locations within the HPS. Locations with a high concentration of L. pneumophila and M. intracellulare coincided with the highest detection of V. vermiformis, suggesting the potential growth of both populations within FLA and additional protection in drinking water. After a period of stagnation lasting at least 2-weeks, the concentrations of OPPPs and amoeba immediately increased and then decreased gradually back to the baseline. Furthermore, monitoring the microbial population after drainage of the hot water tank and partial drainage of the entire HPS demonstrated no significant mitigation of the selected OPPPs. This study demonstrates that these organisms can adjust to their environment during such events and may survive in biofilms and/or grow within FLA, protecting them from stressors in the supplied water.

Comprehensive characterization of aerobic groundwater biotreatment media.

Aerobic biotreatment systems can treat multiple reduced inorganic contaminants in groundwater, including ammonia (NH3), arsenic (As), iron (Fe), and manganese (Mn). While individual systems treating multiple contaminants simultaneously have been characterized and several systems treating one contaminant have been compared, a comparison of systems treating co-occurring contaminants is lacking. This study assessed the treatment performance and microbial communities within 7 pilot- and full-scale groundwater biotreatment systems in the United States that treated waters with pH 5.6-7.8, 0.1-2.0 mg/L dissolved oxygen, 75-376 mg CaCO3/L alkalinity, < 0.03-3.79 mg NH3-N/L, < 4-31 µg As/L, < 0.01-9.37 mg Fe/L, 2-1220 µg Mn/L, and 0.1-5.6 mg/L total organic carbon (TOC). Different reactor configurations and media types were represented, allowing for a broad assessment of linkages between water quality and microbial communities via microscopy, biofilm quantification, and molecular methods. Influent NH3, TOC, and pH contributed to differences in the microbial communities. Mn oxidase gene copy numbers were slightly negatively correlated with the influent Mn concentration, but no significant relationships between gene copy number and influent concentration were observed for the other contaminants. Extracellular enzyme activities, community composition, and carbon transformation pathways suggested heterotrophic bacteria may be important in nitrifying biofilters. Aerobic groundwater biofilters are complex, and improved understanding could lead to engineering enhancements.

Depth profiles of biological aerated contactors: Characterizing microbial activity treating reduced contaminants.

The biological treatment process consisting of an aerated contactor and filter is effective for groundwaters containing elevated ammonia and other reduced contaminants, including iron, manganese, arsenic, and methane. Depth profiles characterizing microbial activity across aerated contactors are lacking. A 1-year pilot study comparing gravel- and ceramic-packed contactors was conducted, and media depth profile samples were collected at the conclusion of the study. Media and water samples also were collected from pilot-scale aerated contactors at 4 other water systems. Water quality, media surface metals concentrations, and a suite of biofilm parameters were analyzed. Media surface metals concentrations were greatest at the influent end. ATP concentrations, extracellular polymeric substances, and extracellular enzyme activities tended to be similar across depth. Bacteria and functional genes involved in contaminant oxidation co-occurred and tended to decrease across depth, but were not correlated to the media metals concentration. Microbial community composition changed with depth, and the diversity either decreased or remained similar. The microbial activity profiles through aerated contactors differed from what is typically reported for groundwater biofilters, suggesting that the different reactor flow and dissolved oxygen profiles impacted the microbial community.

Recycling urine for bioelectrochemical hydrogen production using a MoS2 nano carbon coated electrode in a microbial electrolysis cell.

In this study, a novel molybdenum disulfide (MoS2) nano-carbon (NC) coated cathode was developed for hydrogen production in a microbial electrolysis cell (MEC), while treating simulated urine with 2-6 times dilution (conductivity <20 mS cm-1). MoS2 nanoparticles were electrodeposited on the NC coated cathodes at -100, -150 and -200 µA cm-2 and their performances were evaluated in the MEC. The chronopotentiometry (CP) tests showed the improved catalytic activity of MoS2-NC cathodes with much lower cathode overpotential than non-MoS2 coated electrodes. The MoS2-NC200 cathode, electrodeposited at -200 µA cm-2, showed the maximum hydrogen production rate of 0.152 ± 0.002 m3 H2 m-2 d-1 at 0.9V of Eap, which is comparable to the previously reported Pt electrodes. It was found that high solution conductivity over 20 mS cm-1 (>600 mg L-1 NH3-N) can adversely affect the biofilm architecture and the bacterial activity at the anode of the MEC. Exoelectrogenic bacteria for this system at the anode were identified as Tissierella (Clostridia) and Bacteroidetes taxa. Maximum ammonia-nitrogen (NH3-N) and phosphorus (PO4 3--P) removal were 68.7 and 98.6%, respectively. This study showed that the newly fabricated MoS2-NC cathode can be a cost-effective alternative to the Pt cathode for renewable bioelectrochemical hydrogen production from urine.

Understanding Microbial Loads in Wastewater Treatment Works as Source Water for Water Reuse.

Facing challenges in water demands and population size, particularly in the water-scarce regions in the United States, the reuse of treated municipal wastewater has become a viable potential to relieve the ever-increasing demands of providing water for (non-)potable use. The objectives of this study were to assess microbial quality of reclaimed water and to investigate treatability of microorganisms during different treatment processes. Raw and final treated effluent samples from three participating utilities were collected monthly for 16 months and analyzed for various microbial pathogens and fecal indicator organisms. Results revealed that the detectable levels of microbial pathogens tested were observed in the treated effluent samples from all participating utilities. Log10 reduction values (LRVs) of Cryptosporidium oocysts and Giardia cysts were at least two orders of magnitude lower than those of human adenovirus and all fecal indicator organisms except for aerobic endospores, which showed the lowest LRVs. The relatively higher LRV of the indicator organisms such as bacteriophages suggested that these microorganisms are not good candidates of viral indicators of human adenovirus during wastewater treatment processes. Overall, this study will assist municipalities considering the use of wastewater effluent as another source of drinking water by providing important data on the prevalence, occurrence, and reduction of waterborne pathogens in wastewater. More importantly, the results from this study will aid in building a richer microbial occurrence database that can be used towards evaluating reuse guidelines and disinfection practices for water reuse practices.

Comparison of two culture methods for the enumeration of Legionella pneumophila from potable water samples.

Legionella infections have steadily increased in the United States over the last 20 years, and most of these infections have been attributed to contaminated water. The gold standard for confirmation of Legionella presence in water is culturing with Buffered Charcoal Yeast Extract (BCYE) agar. Following many modifications, this method is still time-consuming, expensive, and can take longer than 10 days for full confirmation. The Legiolert is a newer and simpler culture product that is claimed to be able to quantify Legionella pneumophila in 7 days with high sensitivity and specificity and does not need further confirmation for the presence of L. pneumophila. This study compared the culturability of L. pneumophila occurring in a simulated home plumbing system using both Legiolert and BCYE agar methods. Out of 185 water samples, Legiolert and BCYE method detected L. pneumophila in 83 and 85% of the samples, respectively. The two methods were determined to be statistically equivalent for culturability of L. pneumophila, though the detected levels by Legiolert were slightly higher than the BCYE method. The molecular confirmation of positive (n = 254) and negative wells (n = 82) with Legiolert also showed a high specificity of 96.5% (i.e., 3.5% false positives (9/254) and 0% false negatives (0/82)).

Bacterial diversity and predicted enzymatic function in a multipurpose surface water system - from wastewater effluent discharges to drinking water production.

BACKGROUND: Rivers and lakes are used for multiple purposes such as for drinking water (DW) production, recreation, and as recipients of wastewater from various sources. The deterioration of surface water quality with wastewater is well-known, but less is known about the bacterial community dynamics in the affected surface waters. Understanding the bacterial community characteristics -from the source of contamination, through the watershed to the DW production process-may help safeguard human health and the environment. RESULTS: The spatial and seasonal dynamics of bacterial communities, their predicted functions, and potential health-related bacterial (PHRB) reads within the Kokemäenjoki River watershed in southwest Finland were analyzed with the 16S rRNA-gene amplicon sequencing method. Water samples were collected from various sampling points of the watershed, from its major pollution sources (sewage influent and effluent, industrial effluent, mine runoff) and different stages of the DW treatment process (pre-treatment, groundwater observation well, DW production well) by using the river water as raw water with an artificial groundwater recharge (AGR). The beta-diversity analysis revealed that bacterial communities were highly varied among sample groups (R = 0.92, p <  0.001, ANOSIM). The species richness and evenness indices were highest in surface water (Chao1; 920 ± 10) among sample groups and gradually decreased during the DW treatment process (DW production well; Chao1: 320 ± 20). Although the phylum Proteobacteria was omnipresent, its relative abundance was higher in sewage and industrial effluents (66-80%) than in surface water (55%). Phyla Firmicutes and Fusobacteria were only detected in sewage samples. Actinobacteria was more abundant in the surface water (≥13%) than in other groups (≤3%). Acidobacteria was more abundant in the DW treatment process (≥13%) than in others (≤2%). In total, the share of PHRB reads was higher in sewage and surface water than in the DW treatment samples. The seasonal effect in bacterial communities was observed only on surface water samples, with the lowest diversity during summer. CONCLUSIONS: The low bacterial diversity and absence of PHRB read in the DW samples indicate AGR can produce biologically stable and microbiologically safe drinking water. Furthermore, the significantly different bacterial communities at the pollution sources compared to surface water and DW samples highlight the importance of effective wastewater treatment for protecting the environment and human health.

A strategy for power generation from bilgewater using a photosynthetic microalgal fuel cell (MAFC).

Microbial fuel cells (MFCs) have recently been applied to generate electricity from oily wastewater. Although MFCs that utilize microalgae to provide a self-supporting oxygen (O2) supply at the cathode have been well discussed, those with microalgae at the anode as an active biomass for treating wastewater and producing electrons are still poorly studied and understood. Here, we demonstrated a bilgewater treatment using single- and double-chamber microalgal fuel cells (SMAFC and DMAFC) capable of generating energy with a novel microalgal strain (Chlorella sorokiniana) that was initially isolated from oily wastewater. Compared to previous MFC studies using green algae, relatively high voltage output (151.3-160.1 mV, 71.3-83.4 mV m-2 of power density) was observed in the SMAFC under O2 controlled systems (i.e., acetate addition or light/dark cycle). It was assumed that, under the O2 depletion, alternative electron acceptors such as bicarbonate may be utilized for power generation. A DMAFC showed better power density (up to 23.9%) compared to the SMAFC due to the separated cathode chamber which fully utilizes O2 as an electron acceptor. Both SMAFC and DMAFC removed 67.2-77.4% of soluble chemical oxygen demands (SCOD) from the synthetic bilgewater. This study demonstrates that the application of algae-based MFCs is a feasible strategy to treat oil-in-water emulsion while generating electricity.

Ecological insights into assembly processes and network structures of bacterial biofilms in full-scale biologically active carbon filters under ozone implementation.

To address the adverse effects of harmful algal blooms, there are increased demands over the implementation of ozone coupled with biologically active carbon (BAC) filters in the drinking water treatment plants. Although the microbial biofilms are vital elements to support the proper performance of BAC filters, except for taxonomic affiliations, little is known about the assembly mechanisms of microbial communities in the full-scale BAC filters. This study aimed to examine how the assembly processes and their associated factors (e.g., influent characteristics, biological interactions) drive the temporal dynamics of bacterial communities in full-scale BAC filters, which underwent ozone implementation (five consecutive seasons from 2017 to 2018). The results revealed that along with the increase of bacterial taxonomic richness and evenness, stochastic processes became more crucial to determine the bacterial community assembly in the summer and autumn after ozone implementation (relative contribution: 61.23% and 83.75%, respectively). Moreover, their corresponding networks possessed simple network structures with lower modularity than other seasons, which implied lesser biological interactions among bacterial populations. The correlation between taxonomic and predicted functional diversities using functional redundancy index indicated that relatively high levels of bacterial functional redundancy (>0.83) were generally present in BAC filters. However, compared to other seasons, significantly higher degrees of functional redundancy existed in the summer and autumn after ozone implementation (0.85 ± 0.01 and 0.86 ± 0.01, respectively). Overall, this work improves our understanding of the microbial ecology of full-scale BAC filters by providing a conceptual framework that characterizes bacterial biofilm assembly processes relevant to performance optimization of full-scale BAC filters.

Algal softening followed by ozonation: The fate of persistent micropollutants and natural organic matter in groundwater.

A proof-of-concept study evaluates the performance of a novel strategy using photosynthetic microorganisms to soften groundwater instead of using caustic chemicals. The microalga Scenedesmus quadricauda was used to increase the pH of the groundwater via natural photosynthesis. This work applied softening as a pretreatment to ozonation of hard groundwater and mainly focused on investigating the multiple effects of algal softening on the degradation of persistent micropollutants upon subsequent ozonation. The algae-induced alkaline conditions (pH > 10) were favorable to catalyze the formation of OH radicals directly from O3 molecules. Moreover, algal softening removed the strong radical-scavenging carbonate species (HCO3- and CO32-) to a much greater extent than that achieved by chemical softening, which was attributed to the combination of mineral carbonation and metabolic CO2 reduction. The fate of the natural organic matter (NOM) was characterized with spectroscopy, chromatography, and bioassay, which indicates that algal treatment decomposed the NOM to be less susceptible to attack by OH radicals. Consequently, the ozonation of alkaline groundwater achieved a better removal of the micropollutant residues in groundwater. Carbamazepine and diclofenac were used as model chemicals of persistent groundwater contaminants and were almost completely removed with an addition of 1.25 mg O3 L-1 (0.63 mg-O3 mg-C-1).

Design and evaluation of degassed anaerobic membrane biofilm reactors for improved methane recovery.

Anaerobic treatment of domestic wastewater (DWW) produces dissolved methane that needs to be recovered for use as an energy product. Membrane-based recovery systems have been reported in the literature but are often limited by fouling. The objective of this study was to develop a methane producing biofilm on the shell side surface a membrane to allow for immediate recovery of methane as it was produced, negating mass transfer resistance caused by fouling. Between 89 and 96% of total methane produced was recovered via in-situ degassing without the need for fouling control or cleaning throughout 72 weeks of operation. High methane recovery efficiencies led to predictions of net positive energy yield in one reactor and a 32-61% reduction in energy demand in the others compared to the control. This research demonstrates the feasibility and usefulness of combining attached growth anaerobic wastewater treatment processes with hollow fiber membrane methane recovery systems for improved operation.

Impact of algal organic matter on the performance, cyanotoxin removal, and biofilms of biologically-active filtration systems.

The occurrence of harmful algal blooms dominated by toxic cyanobacteria has induced continuous loadings of algal organic matter (AOM) and toxins in drinking water treatment plants. However, the impact of AOM on the active biofilms and microbial community structures of biologically-active filtration (BAF), which directly affects the contaminant removal, is not well understood. In this study, we systematically examined the effects of AOM on BAF performance and bacterial biofilm formation over 240 days, tracing the removal of specific AOM components, a cyanotoxin [microcystin-LR (MC-LR)], and microbial community responses. The component analysis (excitation and emission matrix analysis) results for AOM revealed that terrestrial humic-like substances showed the highest removal among all the identified components and were strongly correlated to MC-LR removal. In addition, reduced empty bed contact time and deactivation of biofilms significantly decreased BAF performances for both AOM and MC-LR. The active biofilm, bacterial community structure, and mlrA gene (involved in microcystin degradation) abundance demonstrated that bacterial biofilm composition responded to AOM and MC-LR, in which Rhodocyclaceae, Saprospiraceae, and Comamonadaceae were dominant. In addition, MC-LR biodegradation appeared to be more active at the top than at the bottom layer in BAF. Overall, this study provides deeper insights into the role of biofilms and filter operation on the fate of AOM and MC-LR in BAF.

Legionellosis and Recent Advances in Technologies for Legionella Control in Premise Plumbing Systems: A Review.

This review discusses Legionella, among the most prolific and publicly well-known waterborne pathogens, and advances in potential treatment technologies. The number of cases associated with Legionella continues to rise, as does its public awareness. Currently, cases associated with premise plumbing account for the largest number of legionellosis cases in the United States. So, while it is important to understand Legionella as such, it is also important to investigate how to treat drinking water in premise plumbing for Legionella and other waterborne pathogens. While there are currently several methods recognized as potential means of inactivating waterborne pathogens, several shortcomings continue to plague its implementation. These methods are generally of two types. Firstly, there are chemical treatments such as chlorine, chlorine dioxide, monochloramine, ozone, and copper-silver ionization. Secondly, there are physical treatments such as thermal inactivation and media filtration. Their shortcomings range from being labor-intensive and costly to having negative health effects if not properly operated. Recently developed technologies including ultraviolet (UV) irradiation using light emitting diodes (LEDs) and innovative carbon nanotube (CNT) filters can better control waterborne pathogens by allowing for the simultaneous use of different treatment measures in plumbing systems.

Assessing the chemical compositions and disinfection byproduct formation of biofilms: Application of fluorescence excitation-emission spectroscopy coupled with parallel factor analysis.

There are increased concerns over the contributions of biofilms to disinfection byproduct (DBP) formation in engineered water systems (EWS). However, monitoring the biomolecular characteristics of biofilms to understand their impacts on DBP formation has been a great challenge as it requires complex analytical techniques. This study aimed to examine the applicability of fluorescence excitation-emission matrices (EEMs) coupled with parallel factor analysis (PARAFAC) to assess the chemical compositions and DBP formation of biofilms. Biofilms were collected from reactors grown on R2A media, as well as two drinking water-related organic substrates such as humic substances and algal organic matter. The chemical composition and formation of carbonaceous and nitrogenous DBPs of biofilms were continuously monitored every 21 days for 168 days and correlated with the derived EEM-PARAFAC components. Results indicated that all biofilm samples comprised mostly of protein-like components (∼90%), and to a lesser extent, humic-like components (∼10%). Strong correlations were generally found between tryptophan-like substances and the studied DBP formation (R2min ≥ 0.76, P < 0.05), indicating that they play a major role in producing biofilm-derived DBPs upon chlorination. Moreover, significant discrepancies between the chemical compositions and DBP formation of biofilms and their corresponding feed solutions were observed, likely due to biotransformation and biosorption processes. Overall, this work highlights that EEM-PARAFAC analysis is a promising tool to monitor the biomolecular characteristics of biofilm components and to predict the subsequent DBP formation in optimizing disinfection protocols for EWS.

Electrically heatable carbon nanotube point-of-use filters for effective separation and in-situ inactivation of Legionella pneumophila.

Despite municipal chlorination and secondary disinfection, opportunistic waterborne pathogens (e.g., Legionella spp.) persist in public and private water distribution systems. As a potential source of healthcare-acquired infections, this warrants development of novel pathogen removal and inactivation systems. In this study, electrically heatable carbon nanotube (CNT) point-of-use (POU) filters have been designed to remove and inactivate Legionella pneumophila in water. The CNT/polymer composite membranes effectively removed Legionella (> 99.99%) (i.e., below detection limit) and were able to inactive them on the membrane surface at 100% efficiency within 60 s using ohmic heating at 20 V. The novel POU filters could be used as a final barrier to provide efficient rejection of pathogens and thereby simultaneously eliminate microorganisms in public and private water supplies.

Efficacy of inactivation of human enteroviruses by dual-wavelength germicidal ultraviolet (UV-C) light emitting diodes (LEDs).

The efficacy of germicidal ultraviolet (UV-C) light emitting diodes (LEDs) was evaluated for inactivating human enteroviruses included on the United States Environmental Protection Agency (EPA)'s Contaminant Candidate List (CCL). A UV-C LED device, emitting at peaks of 260 nm and 280 nm and the combination of 260∣280 nm together, was used to measure and compare potential synergistic effects of dual wavelengths for disinfecting viral organisms. The 260 nm LED proved to be the most effective at inactivating the CCL enteroviruses tested. To obtain 2-log10 inactivation credit for the 260 nm LED, the fluences (UV doses) required are approximately 8 mJ/cm2 for coxsackievirus A10 and poliovirus 1, 10 mJ/cm2 for enterovirus 70, and 13 mJ/cm2 for echovirus 30. No synergistic effect was detected when evaluating the log inactivation of enteroviruses irradiated by the dual-wavelength UV-C LEDs.

Treatment of reverse osmosis concentrate using an algal-based MBR combined with ozone pretreatment.

Algal treatment was combined with ozone pretreatment for treatment of synthetic reverse osmosis concentrate (ROC) prior to microfiltration. The research mainly focused on minimizing the fouling of polyvinylidene-fluoride membranes and maximizing the restoration of membrane permeability. The algal treatment alone was only moderately effective for the mitigation of fouling in microfiltration, while a markedly improved performance was achieved when the algal treatment followed ozonation. The combination of ozonation and algal treatment reduced membrane permeability decline and significantly (p < 0.05) increased the reversibility of fouling after hydraulic washing. A longitudinal evaluation was also performed with a goal of achieving a robust removal of contaminants. Ozonation followed by algal treatment was very effective in attenuating both caffeine and carbamazepine, as well as removing organic matter and inorganic nutrients from ROC in a single bioreactor. In this study, an alkaline condition (∼pH 12), produced by microalgae in the light without supplemental aeration was applied for in-situ cleaning of fouled membranes. The result showed that the algal-induced cleaning successfully restored the permeability of organic-fouled membranes during the filtration of both raw and algal-treated ROC. This in-situ strategy offers a novel option for periodic cleaning of fouled membranes while maintaining operational simplicity, especially for existing submerged membrane filtration facilities.