Phenolic endocrine-disrupting chemicals and intersex in wild crucian carp from Hun River, China
Introduction
Both natural and synthetic substances have been implicated as endocrine-disrupting chemicals (EDCs) in animal physiology (Pojana et al., 2007). EDCs are released into aquatic environments from domestic, industrial and agricultural discharge, and even very low concentrations of some EDCs may adversely affect animal physiology, including embryonic development, gonadal formation, and sex differentiation (Guillette et al., 1995, Jobling and Tyler, 2003, Hu et al., 2009).
Previous studies have shown that synthetic estrogenic substances such as some surfactants (e.g., nonylphenol (NP), 4-octylphenol (OP)) and plasticizers (e.g., bisphenol-A (BPA) are ubiquitous in aquatic environments and cause endocrine disruption in aquatic organisms (Jobling et al., 1996, Routledge and Sumpter, 1996, Oehlmann et al., 2000). Histopathological changes in animals have been measured to assess the endocrine-disrupting effects of pollutants. For example, NP, an intermediate degradation product of nonylphenol polyethoxylate surfactants (NPEOs), is known to inhibit testicular growth in rainbow trout (Jobling et al., 1996) and cause histopathological changes in germ and Sertoli cells of the male eelpout (Christiansen et al., 1998). Moreover, NP can cause intersex in medaka at the environmental concentrations (e.g., 10 μg L−1) (Balch and Metcalfe, 2006), which are comparable to some surface water concentrations (Blackburn and Waldock, 1995, Dachs et al., 1999, Shao et al., 2005). This suggests that current NP concentrations may be sufficient to adversely affect organisms in aquatic environments. Similarly, OP is an intermediate in the production of phenolic resins and octylphenol ethoxylates (OPEs), and is the most potent estrogenic alkylphenol known with a potency of approximately 10−3–10−7 relative to 17β-estradiol (E2) (Jobling and Sumpter, 1993, Arnold et al., 1996). The potency of OP was assessed using in vitro VTG induction assays using fish hepatocytes (Turker and Takemura, 2011). In addition, exposure to low concentrations of OP (10−9–10−7 M) can accelerate the onset of sexual differentiation in bullfrogs and disrupt sexually dimorphic expression of steroidogenic factor-1 (Mayer et al., 2003). Equivalent concentrations have been measured in aquatic environments (Bennie et al., 1997), implying that the present concentrations are sufficient to cause ecological harm. BPA is another non-steroidal xenoestrogen with ∼10−4 times the activity of estradiol that has received considerable attention from regulatory agencies and scientists (Witorsch, 2002) because of its extensive use. Previous studies have shown that BPA at 1 μg L−1 can adversely affect reproduction of prosobranch snails (Oehlmann et al., 2000, Sohoni et al., 2001), suggesting that it presents a serious risk to aquatic organisms at environmentally relevant levels. Recent studies have found BPA to be almost as effective as estradiol in triggering some receptor responses (Stahlhut et al., 2009), and to act as an androgen-receptor antagonist disrupting normal endocrine functions (Zoeller et al., 2005, Urbatzka et al., 2007). Moreover, low concentrations of BPA can activate the PKG and EGFR/ERK/c-fos pathways in male germ cells via cross-talk between GPR30 and ER-α (Sheng and Zhu, 2011). Thus, the ecological risks of these compounds in aquatic ecosystems are significant because of their ubiquity in the environment and their documented endocrine-disrupting effects on aquatic organisms (European Community, 2003, Servos et al., 2003, An et al., 2009). And some evidences had been found in the downstream of sewage treatment plants (STPs), including the increasing of vitellogenin (VTG) and the incidences of intersex of wild fish which caused by these EDCs (Minier et al., 2000, Jobling and Tyler, 2003).
China is implementing the Water Pollution Control and Management Project (WPCMP) in some key basins recently, including the Hun River as a key part of the Liao River basin. In accordance with the WPCMP guidelines, measurements of water and sediment contaminants have included estrogenic compounds (Wang et al., 2011, Meng, 2013); however, the potential risks posed by environmentally relevant mixtures of EDCs to aquatic organisms remain largely unknown. More scientific data is required to determine whether these chemicals affect molecular, cellular, biochemical, physiological, or histological changes in wild organisms. Therefore, we assessed the concentrations of synthetic estrogenic substances in the Hun River and the associated biological responses of wild fish. The crucian carp (Carassius carassius), an omnivore freshwater fish, was used as a bioindicator because it is distributed in lakes, rivers, and reservoirs throughout China. Moreover, it is tolerant to various conditions and easy to obtain for sampling (An et al., 2007).
Section snippets
Chemicals and materials
Technical grade NP (Product No.: 28640-96), a mixture of compounds with branched side chains, was purchased from Kanto Chemicals (Tokyo, Japan), 4-t-OP (Product No.: 442858) from Sigma–Aldrich (USA), BPA (Product No.: C10655500) from Dr. Ehrenstorfer GmbH (Germany), and the internal standard BPA-d16 from CDN (Canada). The derivatization reagent BSTFA + TMCS (99:1) was purchased from Supelco (USA). HPLC-grade dichloromethane, ethyl acetate, and cyclohexane were obtained from MREDA Corporation
Histopathology
The basic parameters including body length and body weight showed that the fish from November were larger than those from July, but they did not show significant difference among sites. The values of GSI in fish from S5 in July, however, were lower significantly than the fish from other sites (P < 0.05). The histopathological examination of gonad tissue revealed ovo-testis, the presence of oocytes in the testes of some presumed males, which was suggestive of intersex effects (Fig. 2). The
Conclusion
As we know, this was the first report that the occurrence of intersex morphology and the levels of phenolic compounds were determined in wild crucian carp from the Hun River. Especially, these synthetic estrogenic compounds appeared to originate from treated and untreated municipal sewage from Shenyang, and contribute, at least in part, to the occurrence of intersex morphology. Given the presence of other EDCs in aquatic environments, the actual risk of EDCs to wildlife and human health in Hun
Acknowledgements
The financial support of Water Pollution Control and Management (2013ZX07501-05 & 2009ZX07528-03) was gratefully acknowledged.
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