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Microbial regrowth in premise plumbing is a threat to water safety. Disinfectant residuals are often diminished during water transportation and stagnation, leading to the regrowth of opportunistic pathogens. Although microbial regrowth potential is mostly determined by water treatment, little is known about how each treatment step affects two key factors that contribute to microbial regrowth potential: biodegradable organic matter and microbial abundance. In this study, we operated annular reactors to evaluate the microbial regrowth potential of water shaped after each treatment step in a full-scale drinking water treatment plant with ozonation and biological activated carbon filtration. The assimilable organic carbon and total cell count (TCC) were stable at all treatment steps during the sampling period from July to October 2015. The assimilable organic carbon consumption and TCC net increase in the annular reactors indicated that apparent growth yields (cell number base) of microbial communities were different in each reactor. Regrowth potential evaluated by indigenous microbial community in finished water was reduced to 22% of that in raw water, while 75% of assimilable organic carbon in raw water remained in finished water. It suggested that treatment performance evaluated by indigenous microbial communities was better than that evaluated by assimilable organic carbon. HIGHLIGHTS Changes in microbial regrowth potential in water during advanced drinking water treatment were evaluated.; The apparent growth yields of microbial communities growing in effluent from each treatment step were different.; Indigenous microbial community in finished water was used to assess the changes in specific regrowth-promoting substrates.

Escherichia coli normally cannot grow in the environment. One environmental stress that prevents E. coli growth may be the competition for growth substrates with co-existing micro-organisms. In this study, the growth substrates of E. coli were screened by high-resolution mass spectrometry and compared with those of indigenous bacteria in river water. In an incubation experiment, E. coli multiplied in sterilized river water, but did not multiply when indigenous micro-organisms were present in the water. By analysing dissolved organic matter in the river water before and after E. coli growth, 35 compounds were identified as putative growth substrates of E. coli. Among them, 33 compounds were also identified as putative growth substrates of indigenous bacteria. These results indicate that E. coli and indigenous bacteria compete for organic substrates in river water, which could suppress the growth of E. coli.

Nitrification is a key function of biological activated carbon (BAC) filters for drinking water treatment. It is empirically known that the nitrification activity of BAC filters depends on water temperature, potentially resulting in the leakage of ammonium from BAC filters when the water temperature decreases. However, the ammonium removal capacity of BAC filters and factors governing the capacity remain unknown. This study employed a bench-scale column assay to determine the volumetric ammonium removal rate (VARR) of BAC collected from a full-scale drinking water treatment plant. VARR was determined at a fixed loading rate under different conditions. Seasonal variations of the VARR as well as impacts of the water matrix and water temperature on ammonium removal were quantitatively analyzed. While the VARR in an inorganic medium at 25 °C was maintained even during low water temperature periods and during breakpoint chlorination periods, the water matrix factor reduced the VARR in ozonated water at 25 °C by 33% on average. The VARR in ozonated water was dependent on water temperature, indicating that the microbial activity of BAC did not adapt to low water temperature. The Arrhenius equation was applied to reveal the relationship between VARR and water temperature. The actual ammonium removal performance of a full-scale BAC filter was predicted. VARR is useful for water engineers to reexamine the loading and filter depth of BAC filters.

Dissolved organic matter (DOM) causes organic pollution in lakes, resulting in the occurrence of off-flavour etc. when lake water is used as a drinking water source. In this study, DOM in 11 major lakes in Japan was characterised by high-resolution Orbitrap mass spectrometry. Molecular formulas were assigned to 845–1,451 components per sample. Among them, 555 components were commonly found in all lakes. The DOM compositions of the 11 lakes were clustered into four groups. Correlation analysis could extract specific components whose relative intensities were associated with water quality indices such as specific ultraviolet absorbance, specific dissolved chemical oxygen demand (DCODMn), and specific trihalomethane formation potential (R = 0.80–0.93, p < 0.05). Although further molecular structural analyses of DOM components are necessary, these results could be informative for exploring key candidates related to specific water quality issues. Pre-treatment of samples with permanganate oxidation was applied to screen components which could contribute to DCODMn. DCODMn components accounted for only 7–30% of total peak intensities, indicating the limited performance of permanganate oxidation. Pre-treatment by permanganate coupled with Orbitrap MS revealed that components with higher molecular weight, higher oxygen-to-carbon ratios (O/C), and lower hydrogen-to-carbon ratios (H/C) could be responsible for DCODMn.

Polymeric materials are widely used in premise plumbing. To assess the potential of pipe materials to promote microbial regrowth in drinking water, we incubated drinking water in a new cross-linked polyethylene pipe (N-PEX), a new steel pipe lined with powdered polyethylene (N-SPE), and an old steel pipe lined with powdered polyethylene (O-SPE). As a reference, we also incubated drinking water in a carbon-free glass bottle (REF). Free chlorine rapidly depleted to below the quantification limit in the pipe incubations. While the original drinking water contained 0.4 mg C/L of dissolved organic carbon (DOC), the DOC concentration in N-PEX, N-SPE and O-SPE increased to 5.2, 3.1 and 1.7 mg C/L, respectively, after 8 days of incubation. The total cell count (TCC) in N-PEX, N-SPE, and O-SPE increased from

Rapid urbanization induces organic pollution in closed water bodies and accelerates eutrophication. Dissolved organic matter (DOM), a major fraction of organic matter, causes various problems in water use. DOM sources in lakes are diverse. Moreover, DOM properties can be altered biologically and physicochemically during retention time. However, DOM in lakes consists of complex unknown substances. In the present study, high-resolution Orbitrap mass spectrometry (Orbitrap MS) was employed to determine the molecular-level composition of low-molecular-weight DOM in Lake Inba, the most polluted eutrophic lake in Japan. One-year monitoring demonstrated that the lake's DOM consisted of core components that were detected year-around and season-specific components. Although most DOM components extracted by solid phase extraction in the lake came from inflow rivers, lake-specific components were present, which could be associated with algal growth in the lake. Permanganate pretreatment was integrated with Orbitrap MS to screen for components contributing to dissolved chemical oxygen demand (DCODMn), an environmental parameter used in Japan to define the water quality of lakes and reservoirs. Results indicated that only minor and highly unsaturated components were responsible for DCODMn. With this approach, we were able to shed light on how the DOM composition in a eutrophic lake was shaped at molecular level, which cannot be achieved by conventional analysis.

In premise plumbing, microbial water quality may deteriorate under certain conditions, such as stagnation. Stagnation results in a loss of disinfectant residual, which may lead to the regrowth of microorganisms, including opportunistic pathogens. In the present study, microbial regrowth was investigated at eight faucets in a building over four seasons in one year. Water samples were obtained before and after 24 h of stagnation. In the first 100‍ ‍mL after stagnation, total cell counts measured by flow cytometry increased 14- to 220-fold with a simultaneous decrease in free chlorine from 0.17-0.36‍ ‍mg L

We investigated semiquantitative changes in almost 1000 dissolved organic matter (DOM) features during oxidation with 1 mg of O3 per liter (mg O3/L), 4 mg O3/L, or 4 mg O3/L + 2.5 mg of H2O2 per liter (advanced oxidation process, AOP) by unknown screening analysis with Orbitrap mass spectrometry. The consequential effects on formation of unknown disinfection by-products (DBPs) by chlorination were evaluated in laboratory-scale experiments. Several hundred unsaturated DOM features with positive oxygen-subtracted double bond equivalents per carbon ((DBE–O)/C) were decomposed by the ozone-only treatment and AOP. The AOP decomposed some saturated (negative (DBE–O)/C)) and reduced molecules, which had negative carbon oxidation states (Cos). Several hundred saturated oxidation by-products were detected after ozonation and the AOP. After chlorination, the samples pre-treated with ozone alone resulted in higher formation of unknown DBPs than the AOP pre-treated sample or the sample without oxidation. Over half of the DBP precursors, estimated by electrophilic substitution, were not totally decomposed by any oxidation process, but they were increased after the ozone-only process and AOP. DBP precursors produced by the ozone-only process or AOP formed unique unknown DBPs. Therefore, post-treatment processes after oxidation and before chlorination are important to minimize formation of unknown DBPs.

Kouleothrix (Type 1851) filamentous bacteria associated with bulking incidents were investigated for their quantitative correlations with sludge settleability in municipal wastewater treatment plants (WWTPs) around Japan. Factors affecting their abundance were also explored by statistical analyses with the aim of controlling bulking by suppressing their growth. Real-time quantitative PCR assays showed Kouleothrix spp. mainly ranged from 104−105 copies ng-DNA−1 in WWTPs. They were not always abundant in every WWTP, but their abundance (>105 copies ng-DNA−1) almost always coincided with low activated sludge settleability. Their abundance in nitrogen removal processes was always low (1.08 − 3.83 × 104 copies ng-DNA−1). Denitrification process at the feeding stage may be a useful selection pressure for Kouleothrix spp., leading to their suppression. A stepwise multiple regression analysis revealed that the increase of dissolved oxygen (DO) and pH in an aeration tank have a negative influence, while NH4+ increase in the influent has a positive influence on the Kouleothrix spp. abundance. The three-variable model was significant and accounted for approximately 45% of the variance in the dependent variable (i.e., Kouleothrix spp.). The DO increase had the greatest influence on the Kouleothrix spp. decrease, and thus, the DO control may be a promising way to suppress their growth in WWTPs.

Nitrification is a major ecological process occurring in biological activated carbon (BAC) filters used for advanced drinking water treatment. Ammonia oxidation is a rate-limiting step in nitrification that depends on the activity of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) associated with BAC. However, the growth competition between AOA and AOB remains unclear. In this study, time-course DNA-stable isotope probing (DNA-SIP) was applied to compare the growth activities of AOA and AOB for ammonium and urea in a BAC filter, in which AOA were more abundant than AOB. When a low level of ammonium (0.14 mg N L−1) was continuously supplied to BAC, AOA demonstrated higher autotrophic growth activity. A similar trend was noticed in a microcosm supplied with a low concentration of urea (0.14 mg N L−1) and 13C-labeled bicarbonate (H13CO3−). However, the microcosm supplied with only 13C-urea as the sole nitrogen and carbon source indicated faster assimilation of carbon derived from urea by AOB. It is likely that AOB could directly incorporate and hydrolyze urea for nitrification. On the other hand, AOA demonstrated growth activity only when bicarbonate was added to urea, suggesting that they did not directly incorporate urea for nitrification. Thus, the different substrate affinities of AOA and AOB to ammonium and urea seem to be the reason for the niche separation on BAC. Understanding the different responses of ammonia oxidizers to nitrogen substrates may contribute to the improvement of BAC design and operation in actual advanced drinking water treatment.

Current challenges in the anaerobic bioremediation of benzene are the lack of capable cultures and limited knowledge on the biodegradation pathway. Under methanogenic conditions, benzene may be mineralized by syntrophic interactions between microorganisms, which are poorly understood. The present study developed an optimized formula for anoxic medium to successfully promote the growth of the putative benzene degrader Deltaproteobacterium Hasda-A and enhance the benzene degradation activity of methanogenic enrichment cultures. Within 70‍ ‍d of incubation, the benzene degradation activity and relative abundance of Hasda-A in cultures in the new defined medium increased from 0.5 to >3‍ ‍mg L–1 d–1 and from 2.5% to >17%, respectively. Together with Hasda-A, we found a strong positive relationship between the abundances of superphylum OD1 bacteria, three methanogens (Methanoregula, Methanolinea, and Methanosaeta) and benzene degradation activity. The syntrophic relationship between these microbial taxa and Hasda-A was then demonstrated in a correlation analysis of longitudinal data. The involvement of methanogenesis in anaerobic benzene mineralization was confirmed by inhibition experiments. The high benzene degradation activity and growth of Hasda-A were quickly recovered in successive dilutions of enrichment cultures, proving the feasibility of using the medium developed in the present study to produce highly capable cultures. The present results will facilitate practical applications in bioremediation and research on the molecular mechanisms underlying benzene activation and syntrophic interactions in benzene mineralization.

Fecal indicator bacteria (FIB) are commonly used to evaluate the pollution impact of combined sewer overflows (CSOs) in urban rivers. Although water quality assessment with FIB has a long tradition, recent studies demonstrated that FIB have a low correlation with pathogens and therefore are not accurate enough for the assessment of potential human hazards in water. Consequently, new eligible and more specific indicators have to be identified, which was done in this study via sequencing of genetic markers from total community DNA. To identify potential microbiome-based indicators, microbial communities in samples from an urban river in Tokyo under different climatic conditions (dry and rainy) were compared with the influent and effluent of three domestic wastewater treatment plants (WWTPs) by analyzing 16 S rRNA gene amplicon libraries. In the first part of this study, physicochemical parameters and FIB quantification with selective culture techniques facilitated the identification of samples contaminated with CSO, sewage, or both. This allowed the grouping of samples into CSO-contaminated and non-contaminated samples, an essential step prior to the microbiome comparison between samples. Increased turbidity, ammonia concentrations, and E. coli [up to (9.37 ± 0.95) × 102 CFU/mL after 11.5 mm of rainfall] were observed in CSO-contaminated river samples. Comparison of dry weather (including WWTP samples) and rainy weather samples showed a reduction in microbial diversity in CSO-contaminated samples. Furthermore, the results of this study suggest Bacteroides spp. as a novel indicator of sewage pollution in surface waters.

An improved assessment of environmental risks to public water bodies requires screening a large number of micropollutants. This study reports the development of a novel target screening method based on solid-phase extraction (SPE), HPLC, and high-resolution Orbitrap MS for the analysis of micropollutants with diverse chemical properties. First, target compounds were screened for their detectability by Orbitrap MS. An optimized SPE cartridge and HPLC column maximized recovery and separated most target compounds. The sensitivity and repeatability of the method was validated by determining the detection limits and relative standard deviation (RSD). Eighty-four compounds with highly diverse properties were simultaneously detected with detection limits of 0.1–100 ng/L. Of these compounds, 52 were quantitated, with R2 ≥ 0.99 by linearity analysis and SPE recovery ratios of ≥50%. The remaining 32 compounds were qualitatively detected, with R2

Ammonia-oxidizing archaea (AOA) have recently been proposed as potential players for ammonia removal in wastewater treatment plants (WWTPs). However, there is little evidence directly showing the contribution of AOA to ammonia oxidation in these engineered systems. In this study, DNA-stable isotope probing (DNA-SIP) with labeled 13C-HCO3- was introduced to sludge from a municipal WWTP. Quantitative PCR demonstrated that AOA amoA genes outnumbered AOB amoA genes in this WWTP sludge. AOA amoA gene sequence analysis revealed that AOA present in this WWTP were specific to one subcluster within the group 1.1b Thaumarchaeota. When ammonia was supplied to DNA-SIP incubation, the DNA-SIP profiles demonstrated the incorporation of the 13C into AOA and AOB. However, the 13C was not found to be assimilated into both microorganisms in the incubation without ammonia. Specific primers were designed to target amoA genes of AOA belonging to the subcluster found in this WWTP. Applying the primers to DNA-SIP experiment revealed that AOA of this subcluter most likely utilized inorganic carbon during ammonia oxidation under the studied conditions.

Molecular changes in dissolved organic matter (DOM) from treatment processes at two drinking water treatment plants in Japan were investigated using unknown screening analysis by Orbitrap mass spectrometry. DOM formulas with carbon, hydrogen and oxygen (CHO-DOM) were the most abundant class in water samples, and over half of them were commonly found at both plants. Among the treatment processes, ozonation induced the most drastic changes to DOM. Mass peak intensities of less saturated CHO-DOM (positive (oxygen subtracted double bond equivalent per carbon (DBE-O)/C)) decreased by ozonation, while more saturated oxidation byproducts (negative (DBE-O)/C) increased and new oxidation byproducts (OBPs) were detected. By Kendrick mass analysis, ozone reactions preferred less saturated CHO-DOM in the same alkylation families and produced more saturated alkylation families of OBPs. Following ozonation, biological activated carbon filtration effectively removed300 Da CHO-DOM, including OBPs. Following chlorination, over 50 chlorinated formulas of disinfection byproducts (DBPs) were found in chlorinated water samples where at least half were unknown. Putative precursors of these DBPs were determined based on electrophilic substitutions and addition reactions. Ozonation demonstrated better decomposition of addition reaction-type precursors than electrophilic substitution-type precursors; over half of both precursor types decreased during biological activated carbon filtration.

A dissimilatory nitrate reduction to ammonium (DNRA) microbial community was developed under a high organic carbon to nitrate (C/NO3-) ratio in an anoxic semi-continuous sequencing batch reactor (SBR) fed with glucose as the source of carbon and NO3- as the electron acceptor. Activated sludge collected from a municipal wastewater treatment plant with good denitrification efficiency was used as the inoculum to start the system. The aim of this study was to examine the microbial populations in a high C/NO3- ecosystem for potential DNRA microorganisms, which are the microbial group with the ability to reduce NO3- to ammonium (NH4+). A low C/NO3- reactor was operated in parallel for direct comparisons of the microbial communities that developed under different C/NO3- values. The occurrence of DNRA in the high C/NO3- SBR was evidenced by stable isotope-labeled nitrate and nitrite (15NO3- and 15NO2-), which proved the formation of NH4+ from dissimilatory NO3-/NO2- reduction, in which both nitrogen oxides induced DNRA activity in a similar manner. An analysis of sludge samples with Illumina MiSeq 16S rRNA sequencing showed that the predominant microorganisms in the high C/NO3- SBR were related to Sulfurospirillum and the family Lachnospiraceae, which were barely present in the low C/NO3- system. A comparison of the populations and activities of the two reactors indicated that these major taxa play important roles as DNRA microorganisms under the high C/NO3- condition. Additionally, a beta-diversity analysis revealed distinct microbial compositions between the low and high C/NO3- SBRs, which reflected the activities observed in the two systems.

Bioelectrochemical reactors using AMI-7001S and CMI-7000S membranes were investigated for silver recovery and power generation. High silver removal (83.73-92.50%) and columbic efficiency (11.50-19.89%) were obtained in CMI-based reactor after 24h, although, some diffusion of Ag+ ions was observed. In contrast, substrate loss was found in AMI-based reactor, which caused low overall performance. At an initial Ag+ concentration 2000mg/L, a maximum power density of 5396mW/m3 and 3385mW/m3 were obtained in CMI and AMI-based reactor, respectively. Under SEM analysis, different morphologies of metallic silver deposits were detected on the cathode surfaces, and was confirmed by EDX and XRD technique. High diversity of anodic microbial communities were also found, in which Firmulates, Proteobacteria, and Bateroidetes were dominant phyla. Geobacter sp. and other exoelectrogens could be detected while no Shewanella-like sequences were retrieved. The study suggests that CMI-7000S showed better performance when compared to AMI-7001S at the same operational conditions.

Nitrification, the microbial oxidation of ammonia to nitrate via nitrite, occurs in a wide range of acidic soils. However, the ammonia-oxidizing bacteria (AOB) that have been isolated from soil to date are acid-sensitive. Here we report the isolation and characterization of an acid-adapted AOB from an acidic agricultural soil. The isolated AOB, strain TAO100, is classified within the Gammaproteobacteria based on phylogenetic characteristics. TAO100 can grow in the pH range of 5-7.5 and survive in highly acidic conditions until pH 2 by forming cell aggregates. Whereas all known gammaproteobacterial AOB (γ-AOB) species, which have been isolated from marine and saline aquatic environments, are halophiles, TAO100 is not phenotypically halophilic. Thus, TAO100 represents the first soil-originated and non-halophilic γ-AOB. The TAO100 genome is considerably smaller than those of other γ-AOB and lacks several genes associated with salt tolerance which are unnecessary for survival in soil. The ammonia monooxygenase subunit A gene of TAO100 and its transcript are higher in abundance than those of ammonia-oxidizing archaea and betaproteobacterial AOB in the strongly acidic soil. These results indicate that TAO100 plays an important role in the nitrification of acidic soils. Based on these results, we propose TAO100 as a novel species of a new genus, Candidatus Nitrosoglobus terrae.

The dominant filamentous bacteria associated with bulking incidents in Japanese activated sludge plants with nutrient removal were identified and their quantitative correlations with sludge settleability were assessed, with the aim of controlling bulking incidents by specifically suppressing bacterial growth. Fluorescence in situ hybridization (FISH) analyses using existing oligonucleotide FISH probes indicated that the presence of Eikelboom type 1851 filamentous bacteria belonging to the phylum Chloroflexi is correlated with biomass settleability in the municipal wastewater treatment plants examined. Real-time quantitative PCR (qPCR) assays developed in this study also showed a linear correlation between type 1851 filament members and sludge settleability, with the exception of some winter samples. The real-time qPCR assays and 16S ribosomal RNA gene amplicon sequencing to reveal the microbial community of activated sludge showed that the abundance of type 1851 at 200 mL g−1 of sludge volume index was estimated to be about 1.9% of the total microbial cells. The abundance of type 1851 served as a bulking indicator in plants where type 1851 was dominant.

This study reveals that the abundance of the filament Kouleothrix (Eikelboom type 1851) correlated positively with poor settleability of activated sludge biomass in a Japanese full-scale nutrient removal wastewater treatment plant sampled over a one-year period. 16S rRNA amplicon sequence data confirmed that Kouleothrix was the dominant filament in the plant, with a relative abundance of 3.06% positively correlated with sludge volume index (SVI) (R = 0.691). Moreover, Kouleothrix (type 1851) appeared to form interfloc bridges, typical of bulking sludge, regardless of season. Together with earlier studies that indicated the responsibility of Kouleothrix (type 1851) on bulking events, these data suggest that their high relative abundances alone may be responsible for sludge bulking. 16S rRNA qPCR data for this filament showed changes in its relative abundance correlated with changes in several operational parameters, including mixed liquor temperature, sludge retention time, and suspended solids concentration, and it may be that manipulating these may help control Kouleothrix bulking.

We investigated semiquantitative changes in almost 1000 dissolved organic matter (DOM) features during oxidation with 1 mg of O

The increasing organic contamination of surface water hinders the conventional tap water treatment process. Membrane bioreactors (MBRs) are a promising alternative technology for recovering water from polluted surface water. In this study, the composition changes of dissolved organic matters (DOMs) in MBR and ultraviolet/ozone (UV/O3)-MBR systems for polluted surface water treatment were investigated using Orbitrap mass spectrometry analysis with unknown screening. The intense DOM ions within a mass-to-charge ratio range of 100–500 was detected, and 2340 molecular formulae from 5743 peaks were assigned to the two systems. The most abundant components were formulae with C, H, O, N, and CHO only classes. The highest formulae decrease including CHO, CHON, CHOS, and CHONS were attributed to the bio-carrier used in both systems. Results showed that bioprocess was the main contributor in the DOM reduction, and the integration of UV/O3 into the MBR improved the DOM composition changes. Biodegradable components with low O/C ratio in the CHO and CHON classes remarkably increased in the UV/O3-MBR system. The integration of UV/O3 as a polishing step in the recirculation stream of MBR system was effective in improve the DOM removal.

This study aimed to classify the possible molecular formulas of precursors for disinfection by-products (DBPs) in raw, coagulated, and chlorinated water samples from the U-Tapao Canal, Songkhla, Thailand. The molecular formulas of DBPs in chlorinated water were investigated. Polyaluminum chloride (PACl) was employed as a coagulant. Orbitrap Fourier transform-mass spectrometry was able to estimate the composition of dissolved organic matter (DOM) with the carbon (C), hydrogen (H), oxygen (O), nitrogen (N), and sulfur (S) elements contained and DBPs at the molecular level. The molecular formulas of the DOM in the raw water primarily consisted of CHO and CHON when extracted by lichrolut EN. The CHO and CHON species were rich in lignin-, tannin-, and condensed aromatic-like substances. The DOM with high-molecular-weight from 300 to 500 Da were preferentially removed by coagulation. The PACl coagulation decreased the abundances of lignin-, tannin-, and condensed aromatic-like substances in the CHO formulas, while lignin- and condensed aromatic-like substances in the CHON formulas remained. The remaining precursors corresponded to CHON molecules in the coagulated water, which may result in the formation of some chlorine (Cl)-containing molecules. Several DBPs among the CHOCl and CHONCl species were produced in the chlorinated water through the addition reaction of chlorine. New chlorinated N-DBPs of 21 formulas were detected.

Dissolved organic matter (DOM) have been reported as precursors of disinfection byproducts (DBPs) and its molecular characteristics are rarely investigated due to its complexity. In this study, changes in the characteristics of DOM were investigated in the lower Phong River in Thailand in dry season and after the first rain in rainy season, using a non-targeted analysis with Orbitrap mass spectrometry. The river was rich with CHO features dominated by lignin-like molecules, while lipid-like molecules increase after domestic wastewater discharges. Wastewater discharge released DOM with higher molecular weight (MW) that was less oxygenated (low O/C) and less oxidized (low carbon oxidation state [Cos]). A lake affected by anthropogenic activities contributed more oxidized DOM into the river, while surface runoff carried DOM that is more oxygenated (high O/C), less hydrogenated (low H/C), and more oxidized (high Cos) to the stream. Water treatment further modified DOM to be lower MW. Approximately three hundred Cl-containing features (CHOCl) detected upstream were also found downstream. Disinfection by chlorine (Cl2) or chlorine dioxide (ClO2) formed both CHO and CHOCl DBPs. Low chlorine dosage applied to upstream and downstream samples resulted in many common unknown DBPs while increasing chlorine dosage resulted in more unique DBPs. At the same dosage, Cl2 reacted with DOM more than ClO2, including more oxidized molecules that are refractory to ClO2. Both Cl2 and ClO2 produced chlorinated and non-chlorinated DBPs, and some DBPs were commonly found by both disinfections. Cl2-produced DBPs were more unsaturated (higher [DBE-O]/C) and oxidized (higher Cos) than ClO2-DBPs.

Isolation of complex dissolved organic matter (DOM) from environmental water is a major challenge for unknown screening analysis by high-resolution mass spectrometry. In this study, DOM in process water during advanced drinking water treatment was fractionated sequentially by three solid-phase extraction (SPE) cartridges based on the polarity and charge of DOM molecules. By sequential SPE with unknown screening analysis, over 3000 DOM features were found in raw water, whereas around 2000 were obtained by a single SPE. The hydrophobic neutral (HPON) fraction contained CHO features with highest averaged molecular weight followed by hydrophobic acid (HPOA) and then hydrophilic acid (HPIA). The average degree of carbon double bond equivalents and carbon oxidation states indicated that the HPON fraction contained molecules that were more unsaturated and less oxidized than those of the HPOA and HPIA fractions. Ozone selectively decomposed (1) more unsaturated and less oxidized HPON features, (2) more unsaturated HPOA compounds, and (3) less oxidized HPIA molecules. Oxidation by-products were mostly HPON and HPIA compounds that were more oxidized than the decomposed molecules. During biological activated carbon (BAC) filtration, less oxidized HPON were preferentially removed, whereas HPOA were removed without selectivity. HPON and HPIA molecules with more oxidized character were found to be refractory to BAC treatment. HPON with more unsaturated and HPIA with more oxidized characters were decomposed by chlorine. Many types of HPIA decomposed during chlorination were the oxidation by-products of ozonation that were refractory to BAC treatment. Sequential SPE with unknown screening analysis provided previously unknown details of the molecular characteristics of DOM and its changes during advanced water treatment.

Reclaimed water has recently become an important water source for urban use, but the composition of dissolved organic matter (DOM) in reclaimed water has rarely been characterized at the compound level because of its complexity. In this study, the transformation and changes in composition of low molecular weight DOM in water reclamation processes, where secondary effluent of the municipal wastewater treatment plant was further treated by biofiltration, ozonation and chlorination, were investigated by “unknown” screening analysis using Orbitrap mass spectrometry (Orbitrap MS). The intense ions were detected over an m / z range from 100 to 450. In total, 2412 formulae with various heteroatoms were assigned, and formulae with carbon (C), hydrogen (H) and oxygen (O) only and C, H, O and sulfur (S) were the most abundant species. During biofiltration, CHO-only compounds with relatively high hydrogen to carbon (H/C) ratio or with saturated structure were preferentially removed, while CHOS compounds were mostly removed. Ozonation induced the greatest changes in DOM composition. CHOS compounds were mostly decreased after ozonation while ozone selectively removed CHO compounds with relatively unsaturated structure and produced compounds that were more saturated and with a higher degree of oxidation. After chlorination, 168 chlorine-containing formulae, chlorinated disinfection by-products (DBPs), were additionally detected. Candidate DBP precursors were determined by tracking chlorinated DBPs formed via electrophilic substitution, half of which were generated during the ozonation.

Microbial regrowth, microbial growth after disinfection, is an important problem that deteriorates water quality during the storage and distribution of reclaimed water. Biodegradable organic matter (BOM) that remains after water reclamation processes directly promotes microbial regrowth. In this study we propose a novel assay called the ‘bacterial growth fingerprint (BGF)’ to characterise BOM based on the maximum growth of bacterial strains, which is the extension of the conventional assimilable organic carbon assay for drinking water. Nine bacterial strains were selected from nearly 200 isolates from various reclaimed water systems. These selected bacterial strains exhibited unique substrate utilisation patterns. The BGF assay clearly reflected the difference in the quantity and quality of BOM between six different reclamation plants and the changes in BOM during a full-scale reclamation process. The information on BOM revealed by the BGF assay is useful to optimise the treatment processes or operational conditions for biologically stable reclaimed water.

The nitrogen load was determined in road runoff during rainfall events. Moreover, nitrate isotopes analysis was conducted to determine the contribution of nitrates from atmospheric deposition and leaching from road dust. The concentrations of NO3-N in road runoff were higher than those in atmospheric deposits for each rainfall event, except one event with a long antecedent dry weather period. The δ18O-NO3 in road runoff was lower than in atmospheric deposits and higher than in leachate from road dust; however, no difference in δ15N-NO3 was observed. By using δ18O-NO3 as an indicator for evaluating NO3-N sources in road runoff, contribution ratios of NO3-N from road dust were estimated to be 14–22%, 23–25%, and 22–34% for Event 1 to Event 3, respectively. These results indicated that the NO3-N from the atmosphere accounts for more than half of the NO3-N in road runoff.

Isotope dilution techniques (IDT) and sequential extraction procedures (SEPs) were compared to apprehend the differences between two techniques in determining metal exchangeability and vulnerability to pollute the urban groundwater. For this purpose, soil (n = 2), “soakaway” sediment deposited in the artificial infiltration facilities (AIF) (n = 4), and road dust (n = 2) were sampled from Tokyo metropolitan. Sorption coefficients of four metals (Cu, Zn, Cd and Pb) were assessed through isotopic exchangeability (E-value) and potential mobile pool i.e. addition of exchangeable, reducible and oxidizable fraction obtained by Community Bureau of Reference (BCR)-procedures. The E-value for the three samples were found smaller than the potential mobile pool but were higher than BCR-exchangeable fractions. The use of strong extractants are likely to play an active role in the disagreement between SEPs and IDT. IDT accounts for the isotopic exchangeability while BCR provides information of vulnerability of metals associated with different fractions that can leach under different environmental conditions. Sorption coefficients measured in soakaway sediment was found comparable to soil thus likely to retain metals. However, as variability in environmental conditions is likely to affect Kd, the soakaway sediment may become an active metal source in future rather than acting as the permanent sink. The study concludes that there is the possibility of errors while predicting metal vulnerability to groundwater with both techniques and thus a model compliance integrating the virtue of both techniques will be a way forward.

Aims A continuous quench-flow (CQF) reactor was developed to collect samples at the reaction times of less than one second. The reactor is applied to determine ozone disinfection kinetics of poliovirus and to study whether EMA-qPCR can assess the viral infectivity after ozone disinfection. Methods Ozone disinfection of poliovirus was conducted in the developed CQF, and the disinfection kinetics were tested in the range of 0·7-5·0 s at ozone concentration of 0·08 and 0·25 mg l-1 . Inactivation, damage on viral genome and damage on capsid integrity were determined by plaque assay, quantitative reverse transcription polymerase chain reaction (RT-qPCR) and ethidium monoazide treatment coupled with RT-qPCR (EMA-qPCR), respectively. Results By using CQF, 2·18 and 2·76 log10 reductions were observed at the reaction time of 0·7 s and ozone concentration of 0·08 and 0·25 mg l-1 , respectively, followed by tailing. Ozone disinfection kinetics of poliovirus 1 were better fit by the efficiency factor Hom model than by the Chick-Watson model, or the modified Chick-Watson model. Kinetics observed were similar between RT-qPCR and EMA-qPCR assays at the reaction times of 5 s, viral concentration evaluated by EMA-qPCR was reduced in comparison to stable RT-qPCR results. Both assays still underestimated the virus inactivation. Conclusion The simple developed reactor can be used to investigate viral ozone disinfection kinetics and to elucidate inactivation characteristics or mechanisms at very short exposure times. Significance and impact of the study The developed CQF reactor is beneficial for better understanding of virus inactivation by ozone, and the reactor can be used to better elucidate disinfection kinetics and mechanisms for future research. This work constitutes an important contribution to the existing knowledge of the application and limitation of the EMA/PMA-qPCR to assess virus infectivity after ozone disinfection.

Granular activated carbon (GAC) has been widely introduced to advanced drinking water purification plants to remove organic matter and ammonium. Backwashing, which is the routine practice for GAC maintenance, is an important operational factor influencing the performance of GAC and its microbial biomass. In this study, the effects of backwashing on the ammonium removal potential of GAC were evaluated. In addition, abundances of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) on GAC were analyzed. GAC samples before and after backwashing were collected from a full-scale drinking water purification plant. Samplings were conducted before and after implementation of prechlorination of raw water. The results showed that the ammonium removal potential of the GAC increased by 12% after backwashing before prechlorination (p < 0.01). After implementing the prechlorination, the ammonium removal potential of the GAC decreased by 12% even after backwashing (p < 0.01). The AOA was predominant on the GAC in the two samplings. Regardless of prechlorination, the amounts of the AOA and the AOB remained at the same level before and after backwashing. Analysis of the backwashing water indicated that the amounts of the AOA and AOB washed out from the GAC were negligible (0.08%⁻0.26%) compared with their original amounts on the GAC. These results revealed the marginal role of backwashing on the biomass of ammonia oxidizers on GAC. However, the results also revealed that backwashing could have a negative impact on the ammonium removal potential of GAC during prechlorination.