Seiya Hanamoto

Pepper mild mottle virus as an indicator and a tracer of fecal pollution in water environments: comparative evaluation with wastewater-tracer pharmaceuticals in Hanoi, Vietnam.

We analyzed pepper mild mottle virus (PMMoV) in 36 samples taken from surface water, wastewater, groundwater, tap water and bottled water in Hanoi, Vietnam. We then compared the occurrence and fates of PMMoV with pharmaceuticals and personal care products (PPCPs), which are known wastewater tracers. PMMoV was detected in 94% of the surface water samples (ponds, water from irrigated farmlands and rivers) and in all the wastewater samples. The PMMoV concentration ranged from 5.5×10(6)-7.2×10(6)copies/L in wastewater treatment plant (WWTP) influents, 6.5×10(5)-8.5×10(5)copies/L in WWTP effluents and 1.0×10(4)-1.8×10(6)copies/L in surface water. Among the sixty PPCPs analyzed, caffeine and carbamazepine had high detection rates in surface water (100% and 88%, respectively). In surface water, the concentration ratio of PMMoV to caffeine remained unchanged than that in WWTP influents, suggesting that the persistence of PMMoV in surface water was comparable to that of caffeine. The persistence and the large concentration ratio of PMMoV in WWTP influents to the method detection limit would account for its ubiquitous detection in surface water. In comparison, human enteric viruses (HEV) were less frequently detected (18-59%) than PMMoV in surface water, probably because of their faster decay. Together with the reported high human feces-specificity, our results suggested that PMMoV is useful as a sensitive fecal indicator for evaluating the potential occurrence of pathogenic viruses in surface water. Moreover, PMMoV can be useful as a moderately conservative fecal tracer for specifically tracking fecal pollution of surface water. PMMoV was detected in 38% of the groundwater samples at low concentrations (up to 19copies/L). PMMoV was not detected in the tap water and bottled water samples. In groundwater, tap water and bottled water samples, the occurrence of PPCPs and HEV disagreed with that of PMMoV, suggesting that PMMoV is not suitable as an indicator or a tracer in those waters.

Co-occurrence of estrogenic and antiestrogenic activities in wastewater: quantitative evaluation of balance by in vitro ERα reporter gene assay and chemical analysis.

Endocrine-disrupting chemicals are exogenous substances that alter the function of the endocrine system, with adverse health effects on organisms or their progeny. In vitro estrogen receptor (ER) reporter gene assays have long been used to measure estrogenic activity in wastewater. Nevertheless, there is still uncertainty about their usefulness in environmental monitoring on account of a discrepancy between the estrogenic response of the in vitro assay and concentrations of estrogenic compounds determined by chemical analysis. Here, we measured estrogenic and antiestrogenic activities in wastewater by ERα reporter gene assay. All samples were simultaneously analyzed for estrone, 17β-estradiol, estriol, and 17α-ethynylestradiol, and the concentrations were used to predict estrogenic activity. All samples in which measured estrogenic activity was significantly lower than predicted showed strong antiestrogenic activity. In addition, we confirmed that the fraction that did not have antiestrogenic activity showed stronger estrogenic activity than the unfractionated wastewater extract. These results indicate that antiestrogenic compounds in wastewater suppress the activity of natural estrogens, and the reporter gene assay represents the net activity.

Modeling the photochemical attenuation of down-the-drain chemicals during river transport by stochastic methods and field measurements of pharmaceuticals and personal care products.

Existing stochastic models for predicting concentrations of down-the-drain chemicals in aquatic environments do not account for the diurnal variation of direct photolysis by sunlight, despite its being an important factor in natural attenuation. To overcome this limitation, we developed a stochastic model incorporating temporal variations in direct photolysis. To verify the model, we measured 57 pharmaceuticals and personal care products (PPCPs) in a 7.6-km stretch of an urban river, and determined their physical and biological properties in laboratory experiments. During transport along the river, 8 PPCPs, including ketoprofen and azithromycin, were attenuated by >20%, mainly owing to direct photolysis and adsorption to sediments. The photolabile PPCPs attenuated significantly in the daytime but persisted in the nighttime. The observations were similar to the values predicted by the photolysis model for the photolabile PPCPs (i.e., ketoprofen, diclofenac and furosemide) but not by the existing model. The stochastic model developed in this study was suggested to be a novel and useful stochastic model for evaluating direct photolysis of down-the-drain chemicals, which occurs during the river transport.

Assessing the population equivalent and performance of wastewater treatment through the ratios of pharmaceuticals and personal care products present in a river basin: Application to the River Thames basin, UK

The quality of surface waters in lowland rivers is largely dependent on the efficiency of wastewater treatment. Even in the developed countries, there have been difficulties in evaluating the effectiveness of wastewater management and the proportion of wastewater content (WWC) in the river, as well as in estimating the contributing human population. This study aimed to develop a wastewater quality and quantity assessment based on the occurrence of pharmaceuticals in the receiving waters. A survey of 53 pharmaceuticals in 324 samples (river water and influent and effluent of sewage (wastewater) treatment plants) was carried out in southern England in the River Thames catchment over four years. Carbamazepine was selected as stable marker and from its concentration WWC in the rivers and cumulative human populations along the catchment were estimated. The estimated population had a strong relationship (R2=0.94) with that reported by the local water company. The concentration ratio of the labile marker caffeine to carbamazepine indicated the efficiency of wastewater treatment in the different treatment systems (i.e. trickling filter or activated sludge) and in the receiving waters. The ratio in some river samples revealed unexpected discharges of untreated or poorly treated wastewater, with a total concentration of the analytes (up to 20μg/L) five times higher than that in treated wastewater. Such information could be valuable to estimate the discharge or occurrence of not only non-targeted chemicals, but also pathogens within the basin.

Elevated risk from estrogens in the Yodo River basin (Japan) in winter and ozonation as a management option

A simple model was set up to predict estrogen concentrations and endocrine disruption risk in the Yodo River, Japan. This catchment spans the conurbations of Kyoto and Osaka and is the main source of drinking water for Osaka City, Japan. From the river survey data (5 separate occasions between 2005 and 2008), a maximum of 32 g per day estrone (E1) load was observed in the most downstream site of the river. Predicted E1 concentrations were in reasonable agreement with the measurements taken at several points within the basin from a series of sampling campaigns. The predicted concentrations exceeded a net estradiol (E2) equivalent of 1 ng L(-1) on only a few occasions, suggesting that only limited endocrine disruption phenomena in fish along the Yodo River is likely. The model was then used to examine the impact on estrogen concentrations and endocrine disruption of a number of different scenarios. It was found that in-river biodegradation had little effect on predicted concentrations and the outcome of endocrine disruption along the catchment. However, reduced sewage treatment removal, as can be experienced in winter in Japan, led to levels of 3.1 ng L(-1) E2 equivalents being possible. The reduced river flow in winter in Japan exacerbates the situation as it offers less dilution. It was found that the application of the ozonation process as a tertiary sewage treatment in winter could prevent this higher risk endocrine disruption situation.

Quantitative Distribution of Infectious F-Specific RNA Phage Genotypes in Surface Waters

ABSTRACT F-specific RNA phages (FRNAPHs) are considered potential viral indicators of water pollution due to their occurrence and stability in water environments. However, their suitability as viral indicators is not fully elucidated because the characteristics of FRNAPHs are variable depending on the genotype. In this study, for the characterization of infectious FRNAPH genotypes, integrated culture reverse transcription-PCR coupled with the most probable number approach was applied to surface water samples. Further, to recover low concentrations of FRNAPH genotypes, an FRNAPH recovery method was developed. The novel FRNAPH recovery method using a noncharged microfiltration membrane could effectively recover FRNAPH strains without inactivation, while a method using an electronegative microfiltration membrane resulted in the inactivation of some strains. Infectious FRNAPH genotypes in surface water samples were successfully quantified with an efficiency comparable to that of the conventional plaque assay. Genotype I (GI) and GII FRNAPHs tended to be predominant at locations impacted by treated and untreated municipal wastewater, respectively. The numbers and proportions of infectious FRNAPHs tended to be higher during the winter season when water temperature decreased. IMPORTANCE Properties of FRNAPHs are highly variable depending on their genotypes. Previous typing methods for FRNAPHs are not quantitative and/or are based on molecular assays, which cannot differentiate infective strains from inactive strains. Due to the reasons mentioned above, the utility of FRNAPHs as viral indicators of water pollution has not been fully validated. In this study, a quantitative genotyping method for infectious FRNAPHs was developed and applied to surface water samples. The method enabled characterization of infectious FRNAPH genotypes in terms of their occurrence and seasonality. Moreover, comparison of the method to a conventional molecular assay (reverse transcription-quantitative PCR) enabled characterization of their stability. Our approach can provide novel findings for further validation of FRNAPHs as viral indicators of water pollution.

Source estimation of pharmaceuticals based on catchment population and in-stream attenuation in Yodo River watershed, Japan

Fifty-five pharmaceuticals were monitored at four rivers and inlets and/or outlets of three sewage treatment plants (STPs) in Yodo River watershed, Japan over 17 sampling events. Twenty-six quantified pharmaceuticals were classified by source and fate. The load per person (LPP) of nine pharmaceuticals, including six with observed mass balance in studied river stretch of <80%, was appreciably lower in river water (RW) than in the effluent (EF) of STPs (RW/EF <0.5), indicating that they were susceptible to in-stream attenuation in the study area, while the others were relatively conservative. The LPP of 12 pharmaceuticals in RW were within ±50% of that in EF. Because their mass loadings in rivers were correlated with human population in the catchment and most people use the sewer system, the major source of the 12 pharmaceuticals was considered to be STPs. The LPP of the three most labile pharmaceuticals in STPs (caffeine, theophylline, and acetaminophen) was >1.5 in RW/EF and <1.0 in RW/influent (IF) of STPs. Poorly treated sewage discharged from households without using the sewer system was considered to be influential source of the three pharmaceuticals. The LPP (RW/EF) of caffeine, a pharmaceutical contained in food and beverage, was considerably higher than that of the other two, and this is attributable to untreated gray water discharged at households using the night-soil treatment system. The LPP of two veterinary drugs (sulfamonomethoxine and lincomycin) were >1.5 (RW/EF) and >1.0 (RW/IF). Their mass loadings in rivers showed a positive correlation with swine population in the catchment, although sulfamonomethoxine is equally used in both cattle and swine farming. This was attributable to application of cattle excrement as manure, and lability of sulfamonomethoxine during composting processes. The major source of the two veterinary drugs was considered to be on-site treatment facilities of swine urine.

Modeling the fate of a photoproduct of ketoprofen in urban rivers receiving wastewater treatment plant effluent

Photoproducts of pharmaceuticals have been studied in order not to overlook their potential risks to aquatic organisms. However, no studies have verified an equation for predicting the fate of photoproducts in aquatic environment (Poiger equation) by field measurements, leaving uncertainties in its practical utility. Therefore, we conducted this study to test the applicability of the Poiger equation to 3-ethylbenzophenone (EBP), a photoproduct of ketoprofen (KTP). Photolysis experiments determined the fraction of KTP transformed into EBP as 0.744±0.074 and the quantum yield of EBP degradation as 0.000418±0.000090. Field studies in urban rivers and wastewater treatment plants (WWTPs) revealed that EBP was produced by sunlight, mainly in the rivers, but also appreciably in outdoor primary and secondary clarifiers in the WWTPs. We developed a model in the secondary clarifiers, disinfection tanks, and rivers by incorporating the Poiger equation, which was effective at predicting the concentrations of EBP in the river waters and wastewaters. Thus, our first trial of verification by field measurements enhanced the practical utility of the Poiger equation, though further study including several photoproducts should be conducted.

Detection of Physiological Activities of G Protein-Coupled Receptor-Acting Pharmaceuticals in Wastewater

Although pharmaceuticals are generally found at very low levels in aquatic environments, concern about their potential risks to humans and aquatic species has been raised because they are designed to be biologically active. To resolve this concern, we must know whether the biological activity of pharmaceuticals can be detected in waters. Nearly half of all marketed pharmaceuticals act by binding to the G protein-coupled receptors (GPCRs). In this study, we measured the physiological activity of pharmaceuticals in wastewater. We applied the in vitro transforming growth factor-α (TGFα) shedding assay, which accurately and sensitively detect GPCR activation, to investigate the agonistic/antagonistic activities of wastewater extracts against receptors for angiotensin (AT1), dopamine (D2, D4), adrenergic family members (α1B, α2A, β1, β3), acetylcholine (M1, M3), cannabinoid (CB1), vasopressin (V1A, V2), histamine (H1, H2, H3), 5-hydroxytryptamine (5-HT1A, 5-HT2C), prostanoid (EP3), and leukotriene (BLT1). As a result, antagonistic activity against AT1, D2, α1B, β1, M1, M3, H1, and V2 receptors was detected at up to several μg/L for the first time. Agonistic activity against α2A receptor was also detected. The TGFα shedding assay is useful for measuring the physiological activity of GPCR-acting pharmaceuticals in the aquatic environment.

Detection of Physiological Activity of Pharmaceuticals in Wastewater and River Water

　Pharmaceuticals are widely found in aquatic environments worldwide. Concern about their potential risks to aquatic species has been raised because they are designed to be biologically active. To address this concern, we must know whether biological activity of pharmaceuticals can be detected in waters. Nearly half of all marketed pharmaceuticals act by binding to the G protein-coupled receptor (GPCR). In this study, we measured the physiological activity of GPCR-acting pharmaceuticals in effluent from a wastewater treatment plant (WWTP) and upstream and downstream of its outfall in Japan during 2 years. We used the in vitro transforming growth factor-α (TGFα) shedding assay, which accurately and sensitively detects GPCR activation, to investigate the antagonistic activities of water extracts against receptors for dopamine (D2) and histamine (H1). Activities detected in waters were quantified as antagonist equivalent quantities (EQs). In WWTP effluent extracts, antagonistic activity was detected at several hundred ng/L of sulpiride-EQ (D2) and several μg/L of diphenhydramine (DIP)-EQ (H1). In downstream river water extracts, antagonistic activity against H1 was around several hundred ng/L of DIP-EQ, higher than that upstream owing to the WWTP effluent. This review discusses the research needed to resolve the concern about potential risks of pharmaceuticals in waters to aquatic species.