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Phthalates are compounds used in polymers to increase their flexibility and are now ubiquitous in the environment as a result of widespread use. Because few studies have focused on the adverse effects of these chemicals in aquatic species, the present study aimed to determine the effects of phthalate diesters in amphibians. Western clawed frog (Silurana tropicalis) tadpoles were acutely exposed to water spiked with monomethyl phthalate (MMP; 1.3-1595.5 mg/L), dimethyl phthalate (DMP; 0.03-924.0 mg/L), or dicyclohexyl phthalate (DCHP; 0.3-99.3 mg/L). Because few studies have addressed the toxicity of these specific phthalates in most organisms, the present study used higher concentrations of these chemicals to determine their toxicity pathways in amphibians and at the same time investigate a suite of genes known to be altered by the well-studied phthalates. Both DMP and DCHP increased larval mortality (9.1-924.0 mg/L DMP and 4.1-99.3 mg/L DCHP), increased frequency of malformations in tadpoles (0.1-34.1 mg/L DMP and 4.1-19.0 mg/L DCHP), and up-regulated cellular stress-related messenger-RNA (mRNA) levels (4.1 mg/L DCHP). To characterize the molecular toxicity pathway of these phthalates in tadpoles, transcriptome analysis was conducted using a custom microarray. Parametric analysis of gene set enrichment revealed important changes in the expression of genes related to drug metabolism and transport, liver metabolism, xenobiotic clearance, and xenobiotic metabolism after DMP and DCHP treatments, although these responses were less pronounced with MMP (the metabolite of DMP). The present study is one of the few studies that demonstrated complementarity between gene expression analysis and organismal effects. Environ Toxicol Chem 2016;35:2511-2522. © 2016 SETAC.

The concentrations of 17 drugs of abuse, including cocaine, several amphetamines, opioid drugs, and 2 metabolites--benzoylecgonine, a metabolite of cocaine, and 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrolidine, a metabolite of methadone--were investigated in an urban watershed that is heavily impacted by discharges of municipal wastewater. The artificial sweetener sucralose was also monitored as a persistent tracer of contamination from municipal wastewater. Monitoring was conducted in a municipal wastewater treatment plant (WWTP) and at sites upstream and downstream of the WWTP discharge, as well as in a drinking water treatment plant (DWTP) located 19 km downstream of the WWTP discharge that withdraws raw water from the river. Drug concentrations were monitored with polar organic chemical integrative samplers deployed for 2 wk in the river and in the WWTP and DWTP. Several of the investigated compounds exhibited a decrease in concentration with distance downstream from the wastewater discharge into the river, but there was little attenuation of sucralose, cocaine, benzoylecgonine, morphine, acetylmorphine, acetylcodeine, and oxycodone. Heroin and methadone were not detected at any sample locations. Amphetamine, methamphetamine, 3,4-methylenedioxy-methamphetamine, and 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrolidine were not detected in the samples collected at the drinking water intake. Many of the drugs of abuse were not removed effectively in the DWTP, including cocaine, benzoylecgonine, methylenedioxyamphetamine, ephedrine, and several prescription opioids, most probably because the DWTP was operating at or above its rated treatment capacity. These data indicate that there can be transport of drugs of abuse from wastewater sources into drinking water in urban watersheds.

Treatment of municipal wastewater reduces the concentrations of some pharmaceuticals and personal care products (PPCPs), hormones, and drugs of abuse. However, reduced concentrations of these micropollutants in wastewater may not correlate with reduced toxicity because transformations of micropollutants and/or the formation of disinfection by-products may generate toxic compounds. In the present study, we prepared extracts by solid phase extraction of samples collected from wastewater treatment plants (WWTPs) at various stages of treatment and tested these extracts for toxicity to early life stages of Japanese medaka (Oryzias latipes). Toxicity data for extracts prepared from a WWTP with secondary treatment showed that the numbers of exposed embryos (n = 12 per treatment) that did not hatch increased from 1 of 12 for the treatment with untreated effluent to 5 of 12 for the treatment with final treated effluent. For extracts prepared from a WWTP with tertiary treatment, toxicity among exposed embryos (n = 12 per treatment) also increased with each step of wastewater treatment, as shown by mortalities of 2 of 12 and 8 of 12 in treatments with extracts from untreated and final treated effluent, respectively, as well as an increase in the numbers of embryos that did not hatch from 2 of 12 to 9 of 12 in treatments with untreated and final treated effluent, respectively. Ozonation of treated wastewater collected from a third WWTP caused a high incidence of delayed hatch in exposed embryos (n = 24 per treatment). However, hatching success and the numbers of developmental abnormalities in embryos from this ozonation treatment were not different from controls. The present study shows the value of including toxicity testing to assess the effectiveness of technologies for treatment of municipal wastewater. Environ Toxicol Chem 2018;37:136-144. © 2017 SETAC.

Although conventional wastewater treatment technologies are effective at removing many contaminants of emerging concern (CECs) from municipal wastewater, some contaminants are not removed efficiently. Ozonation may be a treatment option for reducing the concentrations of recalcitrant CECs in wastewater, but this process may generate toxic transformation products. In the present study, we conducted semibatch experiments to ozonate municipal wastewater effluent spiked with 5 commonly detected CECs. The purpose of the present study was to evaluate whether ozonation increased or decreased biological responses indicative of sublethal toxicity in juvenile rainbow trout (Oncorhynchus mykiss) injected intraperitoneally (i.p.) with extracts prepared from ozonated and nonozonated wastewater effluent. Blood, liver, and brain tissues were collected from the fish at 72 h post injection for analysis of a battery of biomarkers. In fish i.p. injected with the extracts from nonozonated wastewater effluent, significant induction of plasma vitellogenin (VTG) was observed, but ozonation of the municipal wastewater effluent spiked with CECs significantly reduced this estrogenic response. However, in fish injected with extracts from spiked municipal wastewater effluent after ozonation, the balance of hepatic glutathione in its oxidized (glutathione disulfide [GSSG]) form was altered, indicating oxidative stress. Levels of the neurotransmitter serotonin were significantly elevated in brain tissue from trout injected with the extracts from ozonated spiked municipal wastewater effluent, a biological response that has not been previously reported in fish. Other in vivo biomarkers showed no significant changes across treatments. These results indicate that ozonation reduces the estrogenicity of wastewater, but may increase other sublethal responses. The increase in biomarker responses after ozonation may be because of the formation of biologically active products of transformation of CECs, but further work is needed to confirm this conclusion. Environ Toxicol Chem 2018;37:274-284. © 2017 SETAC.

Phthalates are compounds used in polymers to increase their flexibility and are now ubiquitous in the environment as a result of widespread use. Because few studies have focused on the adverse effects of these chemicals in aquatic species, the present study aimed to determine the effects of phthalate diesters in amphibians. Western clawed frog (Silurana tropicalis) tadpoles were acutely exposed to water spiked with monomethyl phthalate (MMP; 1.3-1595.5 mg/L), dimethyl phthalate (DMP; 0.03-924.0 mg/L), or dicyclohexyl phthalate (DCHP; 0.3-99.3 mg/L). Because few studies have addressed the toxicity of these specific phthalates in most organisms, the present study used higher concentrations of these chemicals to determine their toxicity pathways in amphibians and at the same time investigate a suite of genes known to be altered by the well-studied phthalates. Both DMP and DCHP increased larval mortality (9.1-924.0 mg/L DMP and 4.1-99.3 mg/L DCHP), increased frequency of malformations in tadpoles (0.1-34.1 mg/L DMP and 4.1-19.0 mg/L DCHP), and up-regulated cellular stress-related messenger-RNA (mRNA) levels (4.1 mg/L DCHP). To characterize the molecular toxicity pathway of these phthalates in tadpoles, transcriptome analysis was conducted using a custom microarray. Parametric analysis of gene set enrichment revealed important changes in the expression of genes related to drug metabolism and transport, liver metabolism, xenobiotic clearance, and xenobiotic metabolism after DMP and DCHP treatments, although these responses were less pronounced with MMP (the metabolite of DMP). The present study is one of the few studies that demonstrated complementarity between gene expression analysis and organismal effects. Environ Toxicol Chem 2016;35:2511-2522. © 2016 SETAC.

The concentrations of 17 drugs of abuse, including cocaine, several amphetamines, opioid drugs, and 2 metabolites--benzoylecgonine, a metabolite of cocaine, and 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrolidine, a metabolite of methadone--were investigated in an urban watershed that is heavily impacted by discharges of municipal wastewater. The artificial sweetener sucralose was also monitored as a persistent tracer of contamination from municipal wastewater. Monitoring was conducted in a municipal wastewater treatment plant (WWTP) and at sites upstream and downstream of the WWTP discharge, as well as in a drinking water treatment plant (DWTP) located 19 km downstream of the WWTP discharge that withdraws raw water from the river. Drug concentrations were monitored with polar organic chemical integrative samplers deployed for 2 wk in the river and in the WWTP and DWTP. Several of the investigated compounds exhibited a decrease in concentration with distance downstream from the wastewater discharge into the river, but there was little attenuation of sucralose, cocaine, benzoylecgonine, morphine, acetylmorphine, acetylcodeine, and oxycodone. Heroin and methadone were not detected at any sample locations. Amphetamine, methamphetamine, 3,4-methylenedioxy-methamphetamine, and 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrolidine were not detected in the samples collected at the drinking water intake. Many of the drugs of abuse were not removed effectively in the DWTP, including cocaine, benzoylecgonine, methylenedioxyamphetamine, ephedrine, and several prescription opioids, most probably because the DWTP was operating at or above its rated treatment capacity. These data indicate that there can be transport of drugs of abuse from wastewater sources into drinking water in urban watersheds.