In-line coagulation assessment for ultrafiltration fouling reduction to treat secondary effluent for water reuse.

Low pressure membranes are attracting attention for their potential to improve secondary effluent quality, but membrane fouling can limit their widespread applicability. In this study, in-line coagulation as pre-treatment to ultrafiltration (UF) was investigated using a bench-scale hollow fiber membrane at a constant flux of 33 L/m2 h. Membrane fouling was monitored by observing change in trans-membrane pressure when the membrane was fed with secondary effluent and in-line coagulated secondary effluent over a 24-h period. The impact of four coagulants at different dosages on reversible and irreversible membrane fouling and permeate quality was studied. It was found that in-line coagulation improved UF performance to varying degrees depending on coagulant type and dosage. Generally, higher reduction of fouling was achieved by increasing coagulant dosage within the 0.5-5.0 mg/L range investigated. Ferric-based coagulants were better than aluminum-based coagulants with respect to improving membrane performance for the secondary effluent investigated, even at low dosages (0.5 mg/L). Further investigations are required to determine how in-line coagulation affects removal of organic compounds through UF membranes.

Application of long amplicon propidium monoazide-PCR to assess the effects of temperature and background microbiota on pathogens in river water.

The decay rates of enteric waterborne pathogens were evaluated following the introduction of Yersinia enterocolitica, Salmonella enterica, Campylobacter jejuni and Arcobacter butzleri into river water at different temperatures (5, 15 and 25°C) for a period of 28 days. To improve the accuracy of the results a molecular viability assay, long amplicon propidium monoazide-polymerase chain reaction (PMA-PCR), was used to quantify the viable cell concentration and results from PCR with and without PMA were compared. As well, the effect of background microbiota was assessed for Y. enterocolitica and S. enterica by inoculating cells into sterile and non-sterile river water. Cell persistence was improved by up to 4 log for Y. enterocolitica and 4.5 log for S. enterica in sterile river water compared to natural river water, showing that the autochthonous biological activity in river water can accelerate the die-off of introduced bacteria. Results also showed that low temperature significantly improved the persistence of all four target bacteria in non-sterile river water. There was a more rapid decline in cell concentration in samples with PMA pretreatment; therefore using PMA-PCR analysis can provide more reliable data on viable/active enteric bacteria in aquatic microcosms and allows for improved assessment of pathogens in the environment.

Influence of water quality on nitrifier regrowth in two full-scale drinking water distribution systems.

The potential for regrowth of nitrifying microorganisms was monitored in 2 full-scale chloraminated drinking water distribution systems in Ontario, Canada, over a 9-month period. Quantitative PCR was used to measure amoA genes from ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA), and these values were compared with water quality parameters that can influence nitrifier survival and growth, including total chlorine, ammonia, temperature, pH, and organic carbon. Although there were no severe nitrification episodes, AOB and AOA were frequently detected at low concentrations in samples collected from both distribution systems. A culture-based presence-absence test confirmed the presence of viable nitrifiers. AOB were usually present in similar or greater numbers than AOA in both systems. As well, AOB showed higher regrowth potential compared with AOA in both systems. Statistically significant correlations were measured between several water quality parameters of relevance to nitrification. Total chlorine was negatively correlated with both nitrifiers and heterotrophic plate count (HPC) bacteria, and ammonia levels were positively correlated with nitrifiers. Of particular importance was the strong correlation between HPC and AOB, which reinforced the usefulness of HPC as an operational parameter to measure general microbiological conditions in distribution systems.

Assessment of Giardia and Cryptosporidium spp. as a microbial source tracking tool for surface water: application in a mixed-use watershed.

Knowledge of host specificity, combined with genomic sequencing of Giardia and Cryptosporidium spp., has demonstrated a microbial source tracking (MST) utility for these common waterborne microbes. To explore the source attribution potential of these pathogens, water samples were collected in a mixed rural-urban watershed in the Township of Langley, in southwestern British Columbia (BC), Canada, over a 2-year period. Cryptosporidium was detected in 63% of surface water samples at concentrations ranging from no positive detection (NPD) to 20,600 oocysts per 100 liters. Giardia was detected in 86% of surface water samples at concentrations ranging from NPD to 3,800 cysts per 100 liters of water. Sequencing at the 18S rRNA locus revealed that 50% of Cryptosporidium samples and 98% of Giardia samples contained species/genotypes (Cryptosporidium) or assemblages (Giardia) that are capable of infecting humans, based on current knowledge of host specificity and taxonomy. Cryptosporidium genotyping data were more promising for source tracking potential, due to the greater number of host-adapted (i.e., narrow-host-range) species/genotypes compared to Giardia, since 98% of Giardia isolates were zoonotic and the potential host could not be predicted. This report highlights the benefits of parasite genomic sequencing to complement Method 1623 (U.S. Environmental Protection Agency) and shows that Cryptosporidium subtyping for MST purposes is superior to the use of Giardia subtyping, based on better detection limits for Cryptosporidium-positive samples than for Giardia-positive samples and on greater host specificity among Cryptosporidium species. These additional tools could be used for risk assessment in public health and watershed management decisions.

Removal of the cyanotoxin anatoxin-a by drinking water treatment processes: a review.

Anatoxin-a (ANTX-a) is a potent alkaloid neurotoxin, produced by several species of cyanobacteria and detected throughout the world. The presence of cyanotoxins, including ANTX-a, in drinking water sources is a potential risk to public health. This article presents a thorough examination of the cumulative body of research on the use of drinking water treatment technologies for extracellular ANTX-a removal, focusing on providing an analysis of the specific operating parameters required for effective treatment and on compiling a series of best-practice recommendations for owners and operators of systems impacted by this cyanotoxin. Of the oxidants used in drinking water treatment, chlorine-based processes (chlorine, chloramines and chlorine dioxide) have been shown to be ineffective for ANTX-a treatment, while ozone, advanced oxidation processes and permanganate can be successful. High-pressure membrane filtration (nanofiltration and reverse osmosis) is likely effective, while adsorption and biofiltration may be effective but further investigation into the implementation of these processes is necessary. Given the lack of full-scale verification, a multiple-barrier approach is recommended, employing a combination of chemical and non-chemical processes.

Formation kinetics of gemfibrozil chlorination reaction products: analysis and application.

Aqueous chlorination kinetics of the lipid regulator gemfibrozil and the formation of reaction products were investigated in deionized water over the pH range 3 to 9, and in two wastewater matrices. Chlorine oxidation of gemfibrozil was found to be highly dependent on pH. No statistically significant degradation of gemfibrozil was observed at pH values greater than 7. Gemfibrozil oxidation between pH 4 and 7 was best represented by first order kinetics. At pH 3, formation of three reaction products was observed. 4'-C1Gem was the only reaction product formed from pH 4-7 and was modeled with zero order kinetics. Chlorine oxidation of gemfibrozil in two wastewater matrices followed second order kinetics. 4'-C1Gem was only formed in wastewater with pH below 7. Deionized water rate kinetic models were applied to two wastewater effluents with gemfibrozil concentrations reported in literature in order to calculate potential mass loading rates of 4'C1Gem to the receiving water.

Practical Framework for Evaluation and Improvement of Drinking Water Treatment Robustness in Preparation for Extreme-Weather-Related Adverse Water Quality Events.

Robustness is the ability of a drinking water treatment plant (DWTP) to achieve the desired finished water quality even during adverse raw water quality events. Increasing the robustness of a DWTP is beneficial for regular operations and especially for extreme weather adaptation. This paper proposes three robustness frameworks: (a) a general framework outlining the main steps and methodology for systematic assessment and improvement of the robustness of a DWTP, (b) a parameter-specific framework applying the general framework to a water quality parameter (WQP), and (c) a plant-specific framework applying the parameter-specific framework to a DWTP. A parameter-specific framework for turbidity is presented using the turbidity robustness index (TRI) for evaluation and applied to a full-scale DWTP in Ontario, Canada. This evaluation was conducted with historical plant data, as well as bench-scale experimental data simulating extremely high-turbidity scenarios. The framework application is capable of identifying (i) less robust processes which are likely to be vulnerable during climate extremes, (ii) operational responses to increasing short-term robustness, and (iii) a critical WQP threshold beyond which capital improvements are necessary. The proposed framework provides insights into the current state of robustness of a DWTP and serves as a tool for climate adaptation planning.

Identifying host sources, human health risk and indicators of Cryptosporidium and Giardia in a Canadian watershed influenced by urban and rural activities.

Cryptosporidium and Giardia were characterized in a watershed in southern Ontario, Canada, over a 2½ year period. River samples were collected every two weeks, primarily near a municipal drinking water treatment plant intake. Cryptosporidium and Giardia were frequently detected with an overall occurrence rate of 88 and 97%, respectively. Giardia concentrations were higher than Cryptosporidium, with median values of 80 cysts 100 L(-1) and 12 oocysts 100 L(-1), respectively. Although pathogens rarely show a significant relationship with fecal or water quality indicators, this study determined that Cryptosporidium, but not Giardia, was significantly correlated with Escherichia coli, turbidity and river flow. There was no correlation between the two types of protozoa, and only Giardia showed a seasonal trend with higher concentrations at cold water temperatures. Cryptosporidium genotyping of all samples found that farm animals and wildlife were an important contributor of oocysts in the watershed, and that Cryptosporidium strains/genotypes of medium to high risk for human infection (C. hominis, C. parvum and C. ubiquitum) were detected in 16% of samples. This study was able to identify Cryptosporidium host sources and human health risk, and to identify differences between Cryptosporidium and Giardia occurrence in the watershed.

Ion-Exchange Treatment of Perfluorinated Carboxylic Acids in Water: Comparison of Polystyrenic and Polyacrylic Resin Structures and Impact of Sulfate on Their Performance.

The removal of three perfluorinated carboxylic acids (PFCAs)-PFHpA, PFOA, and PFNA-in ultrapure and river water was evaluated using two anion-exchange resins-previously unreported macroporous polystyrenic A-500P and a more widely studied macroporous polyacrylic A-860. Both resins had similar properties, allowing direct comparison of PFCA removal performance between the two resin structures/matrices. This study also presents a new gas chromatography-mass spectrometry (GC/MS) method developed for PFCA analysis in water. In ultrapure water, A-500P exhibited higher removal capacity and faster removal compared to A-860, suggesting greater effectiveness of the polystyrenic structure compared to the polyacrylic structure. In the Grand River water, the target PFCAs were well removed by A-500P but not A-860. However, both resins achieved similarly high overall reductions of dissolved organic carbon (∼75%), suggesting, later confirmed in ultrapure water experiments, that inorganic anions (sulfate particularly) were the dominant competitors for the A-860 resin. The uncharged styrenic and acrylic beads (base materials) of the two tested resins were unable to remove PFOA, implying that the dominant removal mechanism involves charge interactions between the negatively charged PFCA and the positively charged anion-exchange functional groups.

Pilot-scale comparison of sodium silicates, orthophosphate and pH adjustment to reduce lead release from lead service lines.

Sodium silicate is thought to mitigate lead release via two mechanisms: by increasing pH and by forming a protective silica film. A pilot-scale study using an excavated lead service line (LSL) fed with water from a Great Lakes source was undertaken to: (1) clearly distinguish the pH effect and the silica effect; (2) compare sodium silicate to orthophosphate and pH adjustment; (3) determine the nature of silica accumulation in the pipe scale. The LSL was cut into segments and acclimated with water at pH 7.1. Median dissolved lead was 197 µg/L in the last 8 weeks of acclimation and dropped to 16 µg/L, 54 µg/L, and 85 µg/L following treatment with orthophosphate (dose: 2.6 mg-PO4/L, pH: 7.9), pH adjustment (pH: 7.9) and sodium silicate (dose: 20 mg-SiO2/L, pH: 7.9), respectively. When silica dose was increased from 20 mg-SiO2/L to 25 mg-SiO2/L (pH: 8.1), lead release destabilized and increased (median dissolved lead: 141 µg/L) due to formation of colloidal dispersions composed mainly of lead- and aluminum-rich phases as detected by field flow fractionation used with inductively coupled plasma mass spectrometry. Si was present in the scale at a maximum of 2.2 atomic % after 17 weeks of silica dosing at 20 mg- SiO2/L. Under the conditions tested, sodium silicate did not offer any benefits for reducing lead release from this LSL other than increasing pH. However, sodium silicate resulted in lower levels of biofilm accumulation on pipe walls, as measured by heterotrophic plate counts, when compared to orthophosphate.

The detection of Yersinia enterocolitica in surface water by quantitative PCR amplification of the ail and yadA genes.

Yersinia enterocolitica has been detected in surface water, and drinking untreated water is a risk factor for infection. PCR-based methods have been used to detect Y. enterocolitica in various sample types, but quantitative studies have not been conducted in water. In this study, quantitative PCR (qPCR)-based methods targeting the Yersinia virulence genes ail and yadA were used to survey the Grand River watershed in southern Ontario, Canada. Initial testing of reference strains showed that ail and yadA PCR assays were specific for pathogenic biotypes of Y. enterocolitica; however the genes were also detected in one clinical Yersinia intermedia isolate. A survey of surface water from the Grand River watershed showed that both genes were detected at five sampling locations, with the ail and yadA genes detected in 38 and 21% of samples, respectively. Both genes were detected more frequently at colder water temperatures. A screening of Yersinia strains isolated from the watershed showed that the ail gene was detected in three Y. enterocolitica 1A/O:5 isolates. Results of this study show that Yersinia virulence genes were commonly detected in a watershed used as a source of drinking water, and that the occurrence of these genes was seasonal.

An evaluation of methods for the isolation of Yersinia enterocolitica from surface waters in the Grand River watershed.

Yersinia enterocolitica is a foodborne pathogen, but the importance of water as a route of exposure for human infection is not well known. Y. enterocolitica isolation methods were developed primarily for food and clinical samples, and may not be effective for use with environmental samples. The objective of this study was to assess the recovery of Y. enterocolitica from surface water used for drinking water treatment. Four enrichment broths and an alkaline treatment protocol were compared for the isolation of Y. enterocolitica bioserogroup 4/O:3 spiked into surface water samples. Results showed that the methods tested were not effective for the recovery of Y. enterocolitica, primarily due to inadequate inhibition of interfering background microorganisms. Using one method that showed the most potential for recovery, Yersinia spp. were isolated from rivers in southwestern Ontario, Canada, over a 17-month period. Of 200 samples analysed, Yersinia spp. were isolated from 52 samples. All river isolates belonged to non-pathogenic sub-groups, including Y. enterocolitica biotype 1A, Y. aldovae, Y. bercovieri, Y. frederiksenii, Y. intermedia, Y. kristensenii and Y. mollaretii. Results of this study show that method improvements are required to more fully understand the role of water as a source of clinically important Yersinia strains.

Development of a direct DNA extraction protocol for real-time PCR detection of Giardia lamblia from surface water.

Giardia lamblia is one of the most recognized waterborne protozoan parasites causing gastrointestinal disease. A simple but effective DNA extraction protocol for real-time PCR detection from surface water samples was developed in this study. Eleven protocols were compared, which consisted of freeze-thaw treatments (liquid N(2) and boiling water) and purification using the Qiagen DNeasy kit, together with different combinations of proteinase K, PVP360, GITC and Chelex 100 incubation. Using concentrated surface water samples spiked with G. lamblia cysts, the necessary steps for high DNA recovery were shown to be freeze-thaw, DNeasy purification and Chelex 100 incubation. Multiple rounds of freeze-thaw treatment (five cycles per round) were reported for the first time in this study to significantly increase the DNA yield from G. lamblia cysts, from ~20% after one round of freeze-thaw to 40 and 70% after two and three-rounds of freeze-thaw, respectively. More than three rounds of freeze-thaw treatment did not promote additional DNA recovery. The final protocol included three-three-rounds of freeze-thaw treatment, DNeasy purification and Chelex 100 incubation. This method was simpler, more cost-effective, and had a comparable DNA recovery to methods involving immunomagnetic separation.

Selection and evaluation of water pretreatment technologies for managed aquifer recharge (MAR) with reclaimed water.

Managed aquifer recharge with reclaimed water is a promising strategy for indirect potable reuse. However, residual contaminants in the treated wastewater effluent could potentially have adverse effects on human health. Hence, adequate water pretreatment is required. A multi-criteria approach was used to select and evaluate suitable water pretreatment technologies that can remove these critical contaminants in wastewater effluent for MAR identified in a previous study (Yuan et al., 2017). The treatment efficiency targets were calculated based on the concentrations and the suggested limits of critical contaminants. Treatment efficiency credits were then assigned to each treatment option for the removal of critical contaminants based on literature data. Treatment units that resulted in the highest efficiency credit scores were selected and combined into treatment train options, which were evaluated in terms of treatability, cost, and sustainability. This paper proposes an approach for the selection and evaluation of water treatment options, which will be helpful to guide the future implementation of MAR projects with reclaimed water.

Bench-scale column evaluation of factors associated with changes in N-nitrosodimethylamine (NDMA) precursor concentrations during drinking water biofiltration.

Biofiltration has been observed to increase or decrease the concentrations of N-nitrosodimethylamine (NDMA) precursors in the effluents of full-scale drinking water facilities, but these changes have been inconsistent over time. Bench-scale tests comparing biofiltration columns side-by-side exposed to different conditions were employed to characterize factors associated with changes in NDMA precursor concentrations, as measured by application of chloramines under uniform formation conditions (UFC). Side-by-side comparisons of biofiltration media from different facilities fed with water from each of these facilities demonstrated that differences in source water quality were far more important than any original differences in the microbial communities on the biofiltration media for determining whether NDMA precursor concentrations increased, decreased or remained constant across biofilters. Additional tests involving spiking of specific constituents hypothesized to promote increases in NDMA precursor concentrations demonstrated that inorganic nitrogen species associated with nitrification, including ammonia, hydroxylamine and chloramines, and biotransformation of known precursors (i.e., municipal wastewater and the cationic polymer, polyDADMAC) to more potent forms were not important. Biotransformation of uncharacterized components of source waters determined whether NDMA precursor concentrations increased or decreased across biofilters. These uncharacterized source water component concentrations varied temporally and across locations. Where biotransformation of source water precursors increased NDMA precursor concentrations, ∼30-60% of the levels observed in column effluents fed with biofiltration influent water remained associated with the media and could be rinsed therefrom in either the dissolved or particulate form. Ozone pre-treatment significantly reduced NDMA precursor concentrations at one facility, suggesting that pre-oxidation could be an effective technique to mitigate the increase in NDMA precursor concentrations during biofiltration. Biofiltration decreased the concentrations of halogenated disinfection byproduct precursors.

Adsorption characteristics of selected pharmaceuticals and an endocrine disrupting compound-Naproxen, carbamazepine and nonylphenol-on activated carbon.

The adsorption of two representative pharmaceutically active compounds (PhACs) (naproxen and carbamazepine) and one endocrine disrupting compound (nonylphenol) were evaluated on two types of activated carbon. When determining their isotherms at environmentally relevant concentration levels, it was found that at this low concentration range (10-800 ng/L), removals of the target compounds were contrary to expectations based on their hydrophobicity. Nonylphenol (log K(ow) 5.8) was most poorly adsorbed, whereas carbamazepine (log K(ow) 2.45) was most adsorbable. Nonylphenol Freundlich isotherms at this very low concentration range had a much higher 1/n compared to isotherms at much higher concentrations. This indicates that extrapolation from an isotherm obtained at a high concentration range to predict the adsorption of nonylphenol at a concentration well below the range of the original isotherm, leads to a substantial overestimation of its removals. Comparison of isotherms for the target compounds to those for other conventional micropollutants suggested that naproxen and carbamazepine could be effectively removed by applying the same dosage utilized to remove odorous compounds (geosmin and MIB) at very low concentrations. The impact of competitive adsorption by background natural organic matter (NOM) on the adsorption of the target compounds was quantified by using the ideal adsorbed solution theory (IAST) in combination with the equivalent background compound (EBC) approach. The fulfilment of the requirements for applying the simplified IAST-EBC model, which leads to the conclusion that the percentage removal of the target compounds at a given carbon dosage is independent of the initial contaminant concentration, was confirmed for the situation examined in the paper. On this basis it is suggested that the estimated minimum carbon usage rates (CURs) to achieve 90% removal of these emerging contaminants would be valid at concentrations of less than 500 ng/L in natural water.

Optimizing gas chromatographic-mass spectrometric analysis of selected pharmaceuticals and endocrine-disrupting substances in water using factorial experimental design.

An analytical method based on gas chromatography-mass spectrometry (GC-MS) has been developed to simultaneously determine selected acidic and neutral pharmaceuticals and endocrine-disrupting substances in surface and tap water. Solid-phase extraction (SPE) with Oasis HLB cartridges is followed by derivatization of the target analytes in the eluted extract. Derivatization was systematically optimized by employing a factorial experimental design. More specifically a central composite design was applied to search for the optimal conditions of the derivatization process and it was demonstrated that N-methyl-N-tert-butyl-dimethysilyl-trifluoroacetamide (MTBSTFA) had a better overall performance compared to N,O-bis(trimethylsilyl) trifluoroacetamide (BSTFA). The influence of solvent ratios and elution volumes while using SPE were also studied using a factorial design. The method was developed successfully for most of the selected compounds [i.e. ibuprofen, salicylic acid, gemfibrozil, naproxen, triclosan, propranolol, diclofenac, carbamazepine, 4-octylphenol (OP), 4-nonylphenol (NP), nonylphenol-monoethoxylate (NP1EO), nonylphenoxyacetic acid (NP1EC), estrone (E1), and 17alpha-ethinyloestradiol (EE2)]. Relative recoveries for spiked river and tap water ranged from 47 to 130% and 60-109%, respectively. Typical limits of detection were less than 5 ng/L in tap water and less than 10 ng/L in river water. Twelve target compounds were detected in river and tap water samples using the developed method. This method is currently used in bench-scale drinking water treatment studies.

Pathogen and indicator variability in a heavily impacted watershed.

Water samples were collected from 36 locations within the Grand River Watershed, in Southwestern Ontario, Canada from July 2002 to December 2003 and were analyzed for total coliforms, fecal coliforms, Escherichia coli, Escherichia coli O157:H7, and thermophilic Campylobacter spp. A subset of samples was also analyzed for Cryptosporidium spp., Giardia spp., culturable human enteric viruses, and Clostridium perfringens. Storm and snowmelt events were sampled at two locations including a drinking water intake. For the majority of the events, the Spearman rank correlation test showed a positive correlation between E. coli levels and turbidity. Peaks in pathogen numbers frequently preceded the peaks in numbers of indicator organisms and turbidity. Pathogen levels sometimes decreased to undetectable levels during an event. As pathogen peaks did not correspond to turbidity and indicator peaks, the correlations were weak. Weak correlations may be the result of differences in the sources of the pathogens, rather than differences in pathogen movement through the environment. Results from this investigation have implications for planning monitoring programs for water quality and for the development of pathogen fate and transport models to be used for source water risk assessment.

Identification of critical contaminants in wastewater effluent for managed aquifer recharge.

Managed aquifer recharge (MAR) using highly treated effluent from municipal wastewater treatment plants has been recognized as a promising strategy for indirect potable water reuse. Treated wastewater effluent can contain a number of residual contaminants that could have adverse effects on human health, and some jurisdictions have regulations in place to govern these. For those that do not, but where reuse may be under consideration, it is of crucial importance to develop a strategy for identifying priority contaminants, which can then be used to understand the water treatment technologies that might be required. In this study, a multi-criteria approach to identify critical contaminants in wastewater effluent for MAR was developed and applied using a case study site located in southern Ontario, Canada. An important aspect of this approach was the selection of representative compounds for each group of contaminants, based on potential for occurrence in wastewater and expected health or environmental impacts. Due to a lack of MAR regulations in Canada, the study first proposed potential recharge water quality targets. Predominant contaminants, potential additional contaminants, and potential emerging contaminants, which together comprise critical contaminants for MAR with reclaimed water, were then selected based on the case study wastewater effluent monitoring data and literature data. This paper proposes an approach for critical contaminant selection, which will be helpful to guide future implementation of MAR projects using wastewater treatment plant effluents.

Application of flow cytometry to monitor assimilable organic carbon (AOC) and microbial community changes in water.

Flow cytometry is an efficient monitoring tool for rapid cell counting, and can be applied to research on water quality and treatment. In this study, a method that employs flow cytometry and a natural microbial inoculum to determine assimilable organic carbon (AOC) was adapted for use with challenging surface waters that have a high organic and particle content, and subsequently applied in a long term river water study. AOC method optimization showed that river water bacteria could pass through a 0.2µm membrane filter, and therefore membrane filtration combined with heat treatment was required for sample sterilization. Preparation of the natural river inoculum with an acceptable yield value could only be achieved when grown using the natural water source, since growth was limited on different types of inorganic minimal media and in natural spring water. The resulting flow cytometry AOC method was reliable and reproducible, and results were comparable to the standard plate count AOC method. Size exclusion chromatography showed that both high and low molecular weight organic matter fractions were utilized by the natural AOC inoculum. Flow cytometry was used to measure both AOC levels and total cell counts in a long term study to monitor the water quality of a river which was used as a drinking water source. The method could distinguish between high nucleic acid (HNA) and low nucleic acid (LNA) groups of bacteria, and HNA bacteria were found to respond faster than LNA bacteria to seasonal changes in nutrients and water temperature.