Autotrophic growth activity of complete ammonia oxidizers in an upflow biological contact filter for drinking water treatment.

Biological filters effectively remove ammonium from drinking water via nitrification. In a pilot-scale upflow biological contact filter (U-BCF), complete ammonia oxidizers (comammox), which are capable of oxidizing ammonia to nitrate in one cell, were more abundant than ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB). However, little information is available on the contribution of comammox to nitrification. In this study, we evaluated the autotrophic growth activity of comammox associated with biological activated carbon (BAC) in a U-BCF by DNA-stable isotope probing (DNA-SIP). BAC samples collected from the U-BCF were continuously fed mineral medium containing 0.14 mg N L-1 ammonium and 12C- or 13C-labeled bicarbonate for 20 days. DNA-SIP analysis revealed that comammox (clades A and B) as well as AOA assimilated bicarbonate after 10 days of incubation, proving that dominant comammox could contribute to nitrification. Contrarily, AOB remained inactive throughout the observation period. Amplicon sequencing of the 13C-labeled DNA fractions of comammox revealed that specific genotypes other than the most dominant genotype in the original sample were more enriched under the incubation condition for the DNA-SIP experiment. Thus, dominant genotypes of comammox in a U-BCF might utilize organic nitrogen to fuel nitrification in ammonia-limited environments.

Identification of new eligible indicator organisms for combined sewer overflow via 16S rRNA gene amplicon sequencing in Kanda River, Tokyo.

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.

Temperature-Dependent Ammonium Removal Capacity of Biological Activated Carbon Used in a Full-Scale Drinking Water Treatment Plant.

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.

Changes in Dissolved Organic Matter Composition and Disinfection Byproduct Precursors in Advanced Drinking Water Treatment Processes.

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 removed <300 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.

Abundance and diversity of ammonia-oxidizing archaea and bacteria on granular activated carbon and their fates during drinking water purification process.

Ammonia is a precursor to trichloramine, which causes an undesirable chlorinous odor. Granular activated carbon (GAC) filtration is used to biologically oxidize ammonia during drinking water purification; however, little information is available regarding the abundance and diversity of ammonia-oxidizing archaea (AOA) and bacteria (AOB) associated with GAC. In addition, their sources and fates in water purification process remain unknown. In this study, six GAC samples were collected from five full-scale drinking water purification plants in Tokyo during summer and winter, and the abundance and community structure of AOA and AOB associated with GAC were studied in these two seasons. In summer, archaeal and bacterial amoA genes on GACs were present at 3.7 × 10(5)-3.9 × 10(8) gene copies/g-dry and 4.5 × 10(6)-4.2 × 10(8) gene copies/g-dry, respectively. In winter, archaeal amoA genes remained at the same level, while bacterial amoA genes decreased significantly for all GACs. No differences were observed in the community diversity of AOA and AOB from summer to winter. Phylogenetic analysis revealed high AOA diversity in group I.1a and group I.1b in raw water. Terminal-restriction fragment length polymorphism analysis of processed water samples revealed that AOA diversity decreased dramatically to only two OTUs in group I.1a after ozonation, which were identical to those detected on GAC. It suggests that ozonation plays an important role in determining AOA diversity on GAC. Further study on the cell-specific activity of AOA and AOB is necessary to understand their contributions to in situ nitrification performance.

Sensitivity and quantitativeness of large-volume injection combined with liquid chromatography/high-resolution mass spectrometry for target screening analysis of emerging contaminants.

Solid phase extraction (SPE) has been widely used for pretreatment in target screening (TS) analysis. However, some compounds are difficult to recover by SPE or their recovery is unstable for environmental samples. In this study, we tested large-volume injection (LVI) without SPE for TS analysis of 103 compounds listed by the Ministry of the Environment (Japan)-so-called 'items to be surveyed'-using liquid chromatography high-resolution mass spectrometry. We evaluated the limit of quantification (LOQ) by LVI and compared this LOQ with the LOQ by SPE pretreatment using a hydrophilic-lipophilic balance (HLB) combined with activated carbon, which was found previously to afford the best SPE cartridges for target compounds recovery. The LOQ generally decreased as the injection volume increased, and the LOQ was at least 250 times lower for a 500-µL injection than for a 2-µL injection for half of the compounds. LVI provided LOQs lower than the predicted no effect concentration for more compounds than the SPE method. The average matrix effect (ME) by LVI was in the range 70%-130% for 69 out of 97 compounds. The ME was higher or lower for some of the remaining compounds, but the ME was in the range 10%-1000% for all 18 water samples for 84 of the 97 compounds. Comparing the ME by LVI and the recovery ratio by the SPE method showed that LVI achieved more accurate quantitation than the SPE method for a larger number of compounds. Therefore, LVI provides better sensitivity and quantitativeness than the SPE method using HLB and activated carbon for TS analysis of as many 'items to be surveyed' as possible.

Non-target liquid chromatography high-resolution mass spectrometry screening to prioritize unregulated micropollutants that persist through domestic wastewater treatment.

Efforts to regulate and monitor emerging contaminants are insufficient because new chemicals are continually brought to market, and many are unregulated and potentially harmful. Domestic wastewater treatment plants are not designed to remove micropollutants and are important sources of emerging contaminants in the aquatic environment. In this study, non-target screening, an unbiased method for analyzing compounds without prior information, was used to identify compounds that may be emitted in wastewater treatment plant effluent and should be monitored. Nine wastewater treatment plants using different treatment methods were studied, and a non-target screening data-processing method was used. The frequencies at which the contaminants were detected and contaminant persistence through the treatment processes were considered, and then the contaminants were prioritized. The predicted no-effect concentration of each prioritized contaminant was used to determine whether further analysis and monitoring of the contaminant was necessary. Quantitative analyses of five compounds (amantadine, atenolol, benzotriazole, diphenhydramine, and sulpiride) were performed using reference standards. Probable molecular formulae and structures were proposed for 17 contaminants, and the risks posed by the contaminants were estimated using predicted no-effect concentrations. The results provide valuable insights into how unregulated micropollutants can be identified and prioritized for monitoring in future studies.

Characterization of bacterial isolates from water reclamation systems on the basis of substrate utilization patterns and regrowth potential in reclaimed water.

Microbial regrowth causes problems during water reuse. Comprehensive understanding of the microorganisms that can regrow in reclaimed water and their substrate requirements are necessary. In this study, potential regrowth organisms were isolated from seven water reclamation plants in Japan. Based on 16S rDNA analysis, the isolates were grouped into 34 operational taxonomic units, belonging to Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Actinobacteria, Bacteroidetes, and Firmicutes. Substrate utilization profiling using Biolog microplate™ classified the isolates into four groups. Bacteria in Cluster 1 (e.g., Methylobacterium sp. and Acinetobacter sp.) mainly utilized polymers, esters, amides, and alcohol. Isolates in Cluster 2 (e.g., Flavobacterium sp. and Microbacterium sp.) preferred to utilize polymers, carbohydrates, and esters. Isolates in Cluster 3 (e.g., Pseudomonas sp. and Acidovorax sp.) mainly utilized esters, carboxylic acids, and amino acids. Isolates in Cluster 4 (e.g., Enterobacter sp. and Rhodococcus sp.) utilized carbohydrates, esters, and amino acids. All isolates grew in reclaimed water treated by sand filtration, whereas some isolates could not grow in reclaimed water treated by coagulation and ozonation. Most bacteria in the same Biolog clusters exhibited similar growth characteristics in water samples. The potential of bacteria to regrow in reclaimed water likely depended on substrate requirement.

Dynamics of the Microbial Community and Opportunistic Pathogens after Water Stagnation in the Premise Plumbing of a Building.

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-1 to <0.02| |mg L-1. After stagnation, total cell counts were not significantly different among seasons; however, the composition of the microbial community varied seasonally. The relative abundance of Pseudomonas spp. was dominant in winter, whereas Sphingomonas spp. were dominant in most faucets after stagnation in other seasons. Opportunistic pathogens, such as Legionella pneumophila, Mycobacterium avium, Pseudomonas aeruginosa, and Acanthamoeba spp., were below the quantification limit for real-time quantitative PCR in all samples. However, sequences related to other opportunistic pathogens, including L. feeleii, L. maceachernii, L. micdadei, M. paragordonae, M. gordonae, and M. haemophilum, were detected. These results indicate that health risks may increase after stagnation due to the regrowth of opportunistic pathogens.

Emergence of mobile tigecycline resistance gene tet(X4)-harbouring Shewanella xiamenensis in a water environment.

OBJECTIVES: Tigecycline resistance mediated by the mobile tigecycline-inactivating enzyme gene tet(X) in Gram-negative bacteria is an emerging concern for global public health. However, limited information is available on the distribution of tet(X) in the natural environment. In this study, we investigated the presence of tet(X) in environmental Gram-negative bacteria. METHODS: A carbapenem- and tigecycline-resistant Shewanella xiamenensis isolate (NUITM-VS1) was obtained from an urban drainage in Hanoi, Vietnam, in March 2021. Whole-genome sequencing analysis was performed by long- and short-read sequencing, resulting in a complete genome sequence. Antimicrobial resistance genes (ARGs) in the genome were detected based on the custom ARG database, including all known tigecycline resistance genes. RESULTS: Shewanella xiamenensis isolate NUITM-VS1 harboured the tet(X4) gene and the blaOXA-48 carbapenemase gene on the chromosome. tet(X4) was flanked by IS91 family transposase genes, suggesting that the acquisition of tet(X4) was mediated by this mobile gene element (MGE), whereas no MGE was found surrounding blaOXA-48, consistent with previous findings that blaOXA-48-like ß-lactamase genes are species-specific intrinsic ARGs in Shewanella spp. CONCLUSION: To the best of our knowledge, this is the first report of a tet(X4)-harbouring Shewanella sp. isolate. Our results provide genetic evidence of the complexity of the dynamics of clinically important ARGs among bacteria in the water environment.

Shotgun isotope array for rapid, substrate-specific detection of microorganisms in a microbial community.

The shotgun isotope array method has been proposed to be an effective new tool for use in substrate-specific microbe exploration without any prior knowledge of the community composition. Proof of concept was demonstrated by detection of acetate-degrading microorganisms in activated sludge and further verified by independent stable isotope probing (SIP).

Changes in dissolved organic matter during water treatment by sequential solid-phase extraction and unknown screening analysis.

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.

Non-target screening of dissolved organic matter in raw water, coagulated water, and chlorinated water by Orbitrap mass spectrometry.

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.

A Transferable IncC-IncX3 Hybrid Plasmid Cocarrying blaNDM-4, tet(X), and tmexCD3-toprJ3 Confers Resistance to Carbapenem and Tigecycline.

Tigecycline is a last-resort antimicrobial against carbapenemase-producing Enterobacterales (CPE). However, mobile tigecycline resistance genes, tet(X) and tmexCD-toprJ, have emerged in China and have spread possibly worldwide. Tet(X) family proteins function as tigecycline-inactivating enzymes, and TMexCD-TOprJ complexes function as efflux pumps for tigecycline. Here, to the best of our knowledge we report a CPE isolate harboring both emerging tigecycline resistance factors for the first time. A carbapenem- and tigecycline-resistant Klebsiella aerogenes strain, NUITM-VK5, was isolated from an urban drainage in Vietnam in 2021, and a plasmid, pNUITM-VK5_mdr, cocarrying tet(X) and tmexCD3-toprJ3 along with the carbapenemase gene blaNDM-4 was identified in NUITM-VK5. pNUITM-VK5_mdr was transferred to Escherichia coli by conjugation and simultaneously conferred high-level resistance against multiple antimicrobials, including carbapenems and tigecycline. An efflux pump inhibitor reduced TMexCD3-TOprJ3-mediated tigecycline resistance, suggesting that both tigecycline resistance factors independently and additively contribute to the high-level resistance. The plasmid had the IncX3 and IncC replicons and was estimated to be a hybrid of plasmids with different backbones. Unlike IncX3 plasmids, IncC plasmids are stably maintained in an extremely broad range of bacterial hosts in humans, animals, and the environment. Thus, the future global spread of multidrug resistance plasmids such as pNUITM-VK5_mdr poses a public health crisis. IMPORTANCE Tigecycline is important as a last-resort antimicrobial and effective against antimicrobial-resistant bacteria, such as carbapenem-producing Enterobacterales (CPE), whose infections are difficult to treat with antimicrobials. Since 2019, mobile tigecycline resistance genes, tet(X) and tmexCD-toprJ, and their variants have been reported mainly from China, and it has become important to understand their epidemiological situation and detailed genetic mechanisms. In this study, we identified a bacterial isolate coharboring tet(X) and tmexCD-toprJ on the same plasmid. A Klebsiella aerogenes isolate in Vietnam carried both these tigecycline resistance genes on a transferable plasmid leading to high-level resistance to multiple clinically important antimicrobials, including carbapenem and tigecycline, and could actually transfer the plasmid to other bacteria. The spread of such a multidrug resistance plasmid among bacterial pathogens should be of great concern because there are few antimicrobials to combat bacteria that have acquired the plasmid.

Abundance and diversity of ammonia-oxidizing archaea and bacteria on biological activated carbon in a pilot-scale drinking water treatment plant with different treatment processes.

The effects of different placements of rapid sand filtration on nitrification performance of BAC treatment in a pilot-scale plant were evaluated. In this plant, rapid sand filtration was placed after ozonation-BAC treatment in Process (A), while it preceded ozonation-BAC treatment in Process (B). Analysis of amoA genes of ammonia-oxidizing archaea (AOA) and bacteria (AOB) combined with nitrification potential test was conducted. BAC from Process (A) demonstrated slightly higher nitrification potential at every sampling occasion. This might be due to higher abundances of AOB on BAC from Process (A) than those on BAC from Process (B). However, AOA rather than AOB could be predominant ammonia-oxidizers in BAC treatment regardless of the position of rapid sand filtration. The highest nitrification potential was observed for BAC from both processes in February when the highest abundances of AOA-amoA and AOB-amoA genes were detected. Since rapid sand filtration was placed after BAC treatment in Process (A), residual aluminum concentration in BAC influent was higher in Process (A). However, adverse effects of aluminum on nitrification activity were not observed. These results suggest that factors other than aluminum concentration in different treatment processes could possibly have some influence on abundances of ammonia-oxidizing microorganisms on BAC.

Evaluating the mobile heavy metal pool in soak-away sediment, road dust and soil through sequential extraction and isotopic exchange.

Selective sequential dissolution (SSD) and isotopic dilution are two most commonly applied techniques for the measurement of mobile fraction of heavy metal present in the urban environment. This work examined the compliance between SSD proposed by the Community Bureau of Reference (BCR) and isotopic dilution technique (IDT) for determination of mobile pool of heavy metal contained in soakaway sediment, road dust, and soil sample. Heavy metals (Cu, Zn, Cd and Pb) were fractionated using the three-stage BCR protocol, while isotopically exchangeable metal concentrations (E-value) were investigated through isotopic tracers (¹¹¹Cd, 65Cu, ²°7Pb and 66Zn). In general, total contamination level, E-value and BCR exchangeable fractions of different samples followed the similar order of road dust > soakaway sediment > soil. Results revealed that the E-value exceeded the BCR exchangeable fraction in all samples. In addition, the first three fractions of BCR which have potential to become mobile under certain environmental conditions were collectively termed as "potential mobile pool" and compared with E-value. It was concluded that metal extracted by weak acid underestimates the exchangeable fraction while the potential mobile pool extracted by three reagents overestimates the real mobile forms of heavy metals. However, better mobility characteristics of heavy metals can be assessed by coupling information obtained through sequential extraction and isotopic exchange.

Comparison of metal (Zn and Cu) complexation characteristics of DOM in urban runoff, domestic wastewater and secondary effluent.

This study was aimed at comparing the Zn and Cu binding parameters with dissolved organic matter (DOM) in road runoff, wastewater treatment plant (WWTP) effluent and influent. Conditional stability constant (K') and binding site concentration ([L]) are important to predict free or labile metal concentration and toxicity in the water environment. The values of K' and [L] of three DOMs were determined by metal titration and Scatchard linearization. The Zn and Cu titration data for DOMs in WWTP effluent and influent fitted to a two-ligand model, while DOM in road runoff fitted to a single-ligand model. The order of the values of K' was WWTP influent > WWTP effluent > road runoff both for Zn and Cu. Total ambient binding site concentration ([L(T)]) was lower in DOM of road runoff (1.4-1.5 µM) than those in DOM of WWTP effluent (1.6-2.3 µM) and influent (17-18 µM), suggesting lower metal complexation capacity in DOM of road runoff. DOM in WWTP influent was expected to bind to both Zn and Cu more strongly than that of the effluent DOM.

Simultaneous screening for chemically diverse micropollutants in public water bodies in Japan by high-performance liquid chromatography-Orbitrap mass spectrometry.

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 < 0.99 or SPE recovery ratio of <50%. Satisfactory repeatability was obtained (RSD < 13.5%). This method was applied to the surveillance of the Arakawa River in Japan in 2019. Thirty-two compounds, including pesticides, surfactants, plasticizers, adhesives, and industrial solvents, were detected in the river. The measured concentrations of 13 compounds were compared with their predicted no effect concentrations (PNECs). Decanoic acid showed a higher concentration than the corresponding PNEC value, suggesting that its risk to the Arakawa water environment required further evaluation. The concentrations of dicyclohexylamine, 1,3-diphenylguanidine, and 2,4-dichlorophenoxyacetic acid were higher than their corresponding PNEC/10 values, demonstrating that these compounds were of higher priority than other compounds.

Metal partitioning and leaching vulnerability in soil, soakaway sediments, and road dust in the urban area of Japan.

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.

Consecutive ultrafiltration and silica adsorption for recovery of extracellular antibiotic resistance genes from an urban river.

The dissemination of antibiotic resistance (AR) has attracted global attention because of the increasing antibiotic treatment failure it has caused. Through natural transformation, a live bacterium takes up extracellular DNA (exDNA), which facilitates AR dissemination. However, recovery of exDNA from water samples is challenging. In this study, we validated a consecutive ultrafiltration-based protocol to simultaneously recover intracellular DNA (inDNA), dissolved exDNA (Dis_exDNA, dissolved in the bulk water), and adsorbed exDNA (Ads_exDNA, adsorbed to the surfaces of suspended particles). Using hollow fiber ultrafiltration (HFUF), all DNA fractions were concentrated from environmental water samples, after which Dis_exDNA (supernatant) was separated from inDNA and Ads_exDNA (pellets) using centrifugation. Ads_exDNA was washed off from the pellets with proteinase K and sodium phosphate buffer. Dis_exDNA and Ads_exDNA were further concentrated using centrifugal ultrafiltration, from which silica binding was performed. inDNA was extracted from washed pellets with a commercial kit. For inDNA, HFUF showed recovery efficiencies of 96.5 ± 18.5% and 88.0 ± 2.0% for total cells and cultured Escherichia coli, respectively (n = 3). To represent all possible DNA fragments in water environment, exDNA with different lengths (10.0, 4.0, 1.0, and 0.5 kbp) were spiked to test the recovery efficiencies for Dis_exDNA. The whole process achieved 62.2%-62.9% recovery for 10 and 4 kbp exDNA, and 38.8%-44.5% recovery for 1.0 and 0.5 kbp exDNA. Proteinase K treatment enhanced the recovery of Ads_exDNA by 4.0-10.7 times. The protocol was applied to water samples from an urban river in Tokyo, Japan. The abundance of AR genes (ARGs) in inDNA, Dis_exDNA, and Ads_exDNA increased downstream of wastewater treatment plants. ARGs in Ads_exDNA and Dis_exDNA accounted for 1.8%-26.7% and 0.03%-20.9%, respectively, of the total DNA, implying that Ads_exDNA and Dis_exDNA are nonnegligible potential pools for the horizontal transfer of ARGs.