RESUMEN
In June 2016, the Ministry of the Environment of Japan announced a program "EXTEND2016" on the implementation of testing and assessment for endocrine active chemicals, consisting of a two-tiered strategy. The aim of the Tier 1 screening and the Tier 2 testing is to identify the impacts on the endocrine system and to characterize the adverse effects to aquatic animals by endocrine disrupting chemicals detected in the aquatic environment in Japan. For the consistent assessment of the effects on reproduction associated with estrogenic, anti-estrogenic, androgenic, and/or anti-androgenic activities of chemicals throughout Tier 1 screening to Tier 2 testing, a unified test species, Japanese medaka (Oryzias latipes), has been used. For Tier 1 screening, the in vivo Fish Short-Term Reproduction Assay (OECD test guideline No. 229) was conducted for 17 chemicals that were nominated based on the results of environmental monitoring, existing knowledge obtained from a literature survey, and positive results in reporter gene assays using the estrogen receptor of Japanese medaka. In the 17 assays using Japanese medaka, adverse effects on reproduction (i.e., reduction in fecundity and/or fertility) were suggested for 10 chemicals, and a significant increase of hepatic vitellogenin in males, indicating estrogenic (estrogen receptor agonistic) potency, was found for eight chemicals at the concentrations in which no overt toxicity was observed. Based on these results, and the frequency and the concentrations detected in the Japanese environment, estrone, 4-nonylphenol (branched isomers), 4-tert-octylphenol, triphenyl phosphate, and bisphenol A were considered as high priority candidate substances for the Tier 2 testing.
Asunto(s)
Disruptores Endocrinos , Oryzias , Contaminantes Químicos del Agua , Animales , Disruptores Endocrinos/toxicidad , Masculino , Organización para la Cooperación y el Desarrollo Económico , Receptores de Estrógenos , Reproducción , Vitelogeninas/genética , Contaminantes Químicos del Agua/toxicidadRESUMEN
Under the Organisation for Economic Co-operation and Development (OECD), the Ministry of the Environment of Japan (MOE) added Japanese medaka (Oryzias latipes) to the test guideline fish short-term reproduction assay (FSTRA) developed by the United States Environmental Protection Agency (US EPA) using fathead minnow (Pimephales promelas). The FSTRA was designed to detect endocrine disrupting effects of chemicals interacting with the hypothalamic-pituitary-gonadal axis (HPG axis) such as agonists or antagonists on the estrogen receptor (Esr) and/or the androgen receptor (AR) and steroidogenesis inhibitors. We conducted the FSTRA with Japanese medaka, in accordance with OECD test guideline number 229 (TG229), for 16 chemicals including four Esr agonists, two Esr antagonists, three AR agonists, two AR antagonists, two steroidogenesis inhibitors, two progesterone receptor agonists, and a negative substance, and evaluated the usability and the validity of the FSTRA (TG229) protocol. In addition, in vitro reporter gene assays (RGAs) using Esr1 and ARß of Japanese medaka were performed for the 16 chemicals, to support the interpretation of the in vivo effects observed in the FSTRA. In the present study, all the test chemicals, except an antiandrogenic chemical and a weak Esr agonist, significantly reduced the reproductive status of the test fish, that is, fecundity or fertility, at concentrations where no overt toxicity was observed. Moreover, vitellogenin (VTG) induction in males and formation of secondary sex characteristics (SSC), papillary processes on the anal fin, in females was sensitive endpoints to Esr and AR agonistic effects, respectively, and might be indicators of the effect concentrations in long-term exposure. Overall, it is suggested that the in vivo FSTRA supported by in vitro RGA data can adequately detect effects on the test fish, O. latipes, and probably identify the mode of action (MOA) of the chemicals tested.
Asunto(s)
Bioensayo/métodos , Disruptores Endocrinos/toxicidad , Pruebas de Toxicidad/métodos , Antagonistas de Receptores Androgénicos/toxicidad , Andrógenos/toxicidad , Animales , Antagonistas del Receptor de Estrógeno/toxicidad , Estrógenos/agonistas , Femenino , Masculino , Oryzias/fisiología , Receptores de Progesterona/agonistas , Receptores de Progesterona/antagonistas & inhibidores , Reproducción/efectos de los fármacosRESUMEN
The medaka extended one-generation test (MEOGRT) was developed as a multigenerational toxicity test for chemicals, particularly endocrine-disrupting chemicals. Briefly, 3 generations of Japanese medaka (Oryzias latipes) are exposed to a chemical over a 20-wk period: 3 wk in the parental generation (F0), 15 wk in the first generation (F1), and 2 wk in the second generation (F2). The present study reports the first MEOGRT results concerning branched isomer mixtures of 4-nonylphenol (NP). Adult F0 medaka exposed to NP at 5 actual concentrations (1.27, 2.95, 9.81, 27.8, 89.4 µg/L) were unaffected in terms of reproduction, although vitellogenin in the male liver was increased dose-dependently at concentration of 2.95 µg/L and higher. In F1, in contrast, total egg (fecundity), fertile egg, and fertility decreased as NP increased; lowest-observed-effect concentrations (LOECs) for total egg, fertile egg, and fertility were 1.27, 1.27, 27.8 µg/L, respectively. In F1, but not in F0, secondary sex characteristics (i.e., anal fin papillae in males) were suppressed at 27.8 µg/L NP. Vitellogenin induction in adult male fish was slightly weaker in F1 than it was in F0, however. Gonadal sex abnormality and sex reversal occurred at 27.8 and 89.4 µg/L NP in F1 subadults. At 89.4 µg/L NP, all genotypic F1 males in breeding pairs had female phenotype, and some even demonstrated spawning. Concentrations of NP lower than 89.4 µg/L did not affect F2 survival or hatching. The highest detected NP level in environmental freshwater in Japan was approximately a half of the LOEC (1.27 µg/L for F1 fecundity); in other countries, however, environmental concentrations above the LOEC are reported, suggesting that NP may be affecting fish populations. Environ Toxicol Chem 2017;36:3254-3266. © 2017 SETAC.
Asunto(s)
Disruptores Endocrinos/toxicidad , Oryzias/fisiología , Fenoles/toxicidad , Reproducción/efectos de los fármacos , Animales , Femenino , Fertilidad/efectos de los fármacos , Agua Dulce , Japón , Hígado/química , Hígado/efectos de los fármacos , Masculino , Caracteres Sexuales , Pruebas de Toxicidad , Vitelogeninas/análisisRESUMEN
Environmental risk assessments show increased attention to the sublethal effects of chemicals on aquatic organisms. The Organization for Economic Cooperation and Development (OECD) established the "Fish, Short-term Toxicity Test on Embryo and Sac-fry Stages" (OECD test 212) to predict lethal effects. It is still unclear, however, whether this test can predict sublethal effects. Although their sublethal effects are still unknown, chlorinated anilines are widely used in various fields. The purpose of this study, therefore, is to investigate sublethal effects of chlorinated anilines using OECD test 212 with zebrafish, and to examine the correlation of several sublethal effects between embryo and larval stages. Embryos were exposed to aniline and nine chlorinated anilines until 8 days post-fertilization. A delayed lethal effect was observed from three of the 10 anilines tested. In the control group, the swim bladder inflated after hatching, but there was no swim-bladder inflation after exposure to the chlorinated anilines. Fertilized eggs exposed to lower concentrations of test chemicals showed effects during embryogenesis that did not affect mortality rates, such as changes in body curvature and edema. Our results show that chlorinated anilines induce not only lethal effects but also a variety of sublethal effects. Moreover, a detailed estimate of these effects requires study during both embryonic and larval stages. OECD test 212 may therefore prove useful as a method for screening chemicals for lethal and sublethal effects. Copyright © 2017 John Wiley & Sons, Ltd.
Asunto(s)
Compuestos de Anilina/toxicidad , Carcinógenos/toxicidad , Embrión no Mamífero/efectos de los fármacos , Desarrollo Embrionario/efectos de los fármacos , Larva/fisiología , Contaminantes Químicos del Agua/toxicidad , Pez Cebra/embriología , Animales , Monitoreo del Ambiente/métodos , Dosificación Letal MedianaRESUMEN
Butylparaben and benzylparaben, used as preservatives mainly in cosmetic products, have recently been found to be weakly estrogenic. Batch activated-sludge treatment and batch chlorination were carried out to roughly determine the removal efficiency of a wastewater treatment plant. Combining the removal efficiency with the estimated annual consumption and the unaltered excretion ratio, the maximum predicted environmental concentration (PEC) was estimated. Conventional acute/chronic toxicity tests were conducted using Japanese medaka (Oryzias latipes), daphnia (Daphnia magna), and green algae (Pseudokirchneriella subcapitata) for n-butylparaben, i-butylparaben, and benzylparaben. Medaka vitellogenin assays were also conducted for the three compounds and DNA microarray analysis was carried out to examine the effects of benzylparaben on gene expression. The plasma vitellogenin concentration of male medaka increased for concentrations of 200, 100, and 100 microg L(-1) n-butylparaben, i-butylparaben, and benzylparaben for 14 days, respectively, while the expression levels of genes encoding proteins such as p53, cytochrome P450 3A40, and choriogenin-L increased for concentrations higher than 4 microg L(-1) of benzylparaben. Furthermore, the predicted no-effect concentration (PNEC) was calculated using the lethal or effect concentration 50 (LC50 or EC50) values and no-effect concentrations (NOECs) obtained in the toxicity tests for these compounds. The maximum concentrations found in the aquatic environment or sewage effluent (MEC eff) were used to carry out preliminary environmental risk assessment. The calculated MEC/PNEC ratio suggests the necessity of further study such as a more detailed large-scale monitoring and chronic toxicity tests including reproduction inhibition and endocrine disruption.
Asunto(s)
Expresión Génica/efectos de los fármacos , Residuos Industriales , Oryzias/genética , Parabenos/toxicidad , Contaminantes Químicos del Agua/toxicidad , Animales , Ecología , Nivel sin Efectos Adversos Observados , Análisis de Secuencia por Matrices de Oligonucleótidos , Parabenos/aislamiento & purificación , Medición de Riesgo , Aguas del Alcantarillado , Administración de Residuos , Contaminantes Químicos del Agua/aislamiento & purificaciónRESUMEN
Two groups of Japanese medaka (Oryzias latipes) were exposed to 17beta-estradiol (E2: 150ng/L, nominal concentration) for either a short-term exposure (STE: 0-31 days after fertilization (daf); egg-larval period) or a long-term exposure period (LTE: 0-81 daf; egg-adult period) and their subsequent spawning performance was compared in terms of fecundity, spawning time, and fertility. Most genetic males were transformed to phenotypic females by E2 following both short-term and long-term exposure, but spawning performance and gonad somatic index (GSI) of sex-transformed females (XY females) following long-term exposure were lower than those of sex-transformed females following short-term exposure and those of normal females (XX) in the control group. Sex-transformed females in the STE group and normal females possessed mature ovary, whereas most of the sex-transformed females in the LTE group possessed immature ovary, with most oocytes being in the pre-vitellogenic phase. Moreover, the chromosome types of first filial generation delivered from sex-transformed female in STE group composed with 51.9% as XY, 18.5% as YY, and 29.6% as XX. From these results, it seems that exposure to E2 until the end of the larval period produces sex-transformed medaka with high reproductive ability, similar to normal females, but longer exposure to E2 may inhibit sexual maturation in the sex-transformed female.
Asunto(s)
Estradiol/toxicidad , Feminización/veterinaria , Enfermedades de los Peces/inducido químicamente , Oryzias/fisiología , Reproducción/efectos de los fármacos , Contaminantes Químicos del Agua/toxicidad , Animales , Estradiol/análisis , Femenino , Feminización/inducido químicamente , Larva/efectos de los fármacos , Masculino , Oocitos/efectos de los fármacos , Ovario/efectos de los fármacos , Cromosomas Sexuales/genética , Estadística como Asunto , Factores de Tiempo , Vitelogeninas/análisis , Contaminantes Químicos del Agua/análisisRESUMEN
Gonad histological changes were examined in Japanese medaka exposed to 17beta-estradiol (E2) during early-life stages. Two experiments were conducted at different concentrations of E2 (33.5 and 140.6 ng/L, mean value of measurement) and larvae and juveniles were observed for histological changes in the gonad. Differentiation of ovary and testis in control fish was apparent 12 days post-hatch (dph). At 12 dph, normal testes were observed in male fish that had been exposed to 33.5 ng/L E2, but at 14 and 20 dph, testis-ova was recognized in male fish. Male fish exposed to 140.6 ng/L E2 had testis-ova at 12 dph and gradual transformation to ovary was observed in male fish until 20 dph. In both experiments, the ovarian tissue in testis of male fish exposed to E2 was frequently distributed along the central transverse axis of the gonad, expanding into the transverse axis. The results indicated that 17beta-estradiol can induce testis-ova in male medaka during the larval period and sex-transformation is more frequent at higher (140.6 ng/L) than lower concentrations (33.5 ng/L) of estradiol. The results also demonstrated that testis-ova first appear in the central area of the transverse axis of testis.
Asunto(s)
Estradiol/toxicidad , Feminización/veterinaria , Enfermedades de los Peces/inducido químicamente , Gónadas/efectos de los fármacos , Oryzias/fisiología , Contaminantes Químicos del Agua/toxicidad , Animales , Cartilla de ADN/química , Feminización/inducido químicamente , Gónadas/anomalías , Gónadas/patología , Masculino , Oryzias/embriología , Reacción en Cadena de la Polimerasa/veterinariaRESUMEN
Bioconcentration factors (BCF) for pentachlorophenol (PCP) and 2,4-dichlorophenol (2,4-DCP) in Japanese medaka (Oryzias latipes) were determined at five different concentrations of the chemicals, between 0.1 and 10 microg/l (PCP), 0.3 and 30 microg/l (2,4-DCP), in the ambient water. Medaka were exposed to each chemicals in a continuous-flow system during the embryonic development period and 60 days after hatching from eggs collected in the laboratory. Both the exposure time and the aqueous concentrations are much more realistic and closer to natural aquatic environments than those used in conventional BCF studies. The BCF values of PCP were from (4.9+/-2.8)x10(3) at the aqueous concentration of 0.074+/-0.028 microg/l to (2.1+/-1.4)x10(3) at 9.70+/-0.56 microg/l. The BCF value of 2,4-DCP were from (3.4+/-3.0)x10(2) at 0.235+/-0.060 microg/l to 92+/-27 at 27.3+/-1.6 microg/l. Generally, BCF values increased as the aqueous concentrations of PCP or 2,4-DCP decreased. This finding suggests that a relatively low and realistic aqueous concentration of these compounds is necessary to more accurately determine their BCF values in natural aquatic environments. Conventional BCF experiments at higher aqueous concentrations may underestimate the BCF values.
Asunto(s)
Clorofenoles/farmacocinética , Oryzias , Pentaclorofenol/farmacocinética , Contaminantes Químicos del Agua/farmacocinética , Animales , Clorofenoles/análisis , Monitoreo del Ambiente , Japón , Pentaclorofenol/análisis , Distribución Tisular , Contaminantes Químicos del Agua/análisisRESUMEN
Sinotaia quadrata histrica is a fresh water viviparous snail distributing from the Kanto region to Kyushu Island, Japan. About 7000 snails were collected in Lake Kasumigaura (L. Nishiura and L. Kitaura) in 2001 and 2002, and the sex ratio, maturity in terms of the gonad-somatic index (GSI) and operculum diameter were determined. The total female proportion was 55.2% in 2001, 53.0% in 2002 in L. Nishiura, and that of L. Kitaura was 60.4% in 2002. Comparing the season, the female proportion was the highest during early summer in both 2001 (59.6%, July in L. Nishiura) and 2002 (61.6%, June in L. Nishiura, 65.8%, July in L. Kitaura). The GSI of females in L. Nishiura significantly increased from April to May and significantly decreased from June to August. The GSI of males was higher in spring, but significantly lower from June to August. The mean female operculum diameter was consistently larger than that of males for each month and year, and a particularly significant difference was found between females and males from April to August 2001, and from April to September 2002. The number of resting zones on the operculum correlates with the operculum diameter and the female proportion was larger in the snails, which have a high number of resting zones, suggesting a sex-dependent difference in age composition. This study estimated that the sex ratio and seasonal maturation of S. quadrata histrica and the sex-dependent difference in age composition might contribute to the population structure in L. Kasumigaura.