An amphibian high fat diet model confirms that endocrine disruptors can induce a metabolic syndrome in wild green frogs (Pelophylax spp. complex).

A pre-diabetes syndrome induced by endocrine disruptors (ED) was recently demonstrated in the model amphibian Silurana (Xenopus) tropicalis and was suggested to be a potential cause of amphibian population decline. However, such effects have not been found in wild type frogs exposed to ED and the capacity of amphibians to physiologically develop diabetes under natural conditions has not been confirmed. This study showed that a high fat diet (HFD) model displaying the important characteristics of mammal HFD models including glucose intolerance, insulin resistance and nonalcoholic fatty liver disease (NAFLD) can be developed with green frogs (Pelophylax spp.). Wild green frogs exposed to 10 µg L-1 benzo [a]pyrene (BaP) for 18 h also displayed several characteristics of the pre-diabetes phenotype previously observed in Xenopus including glucose intolerance, gluconeogenesis activation and insulin resistance. The study results confirmed that metabolic disorders induced by ED in wild green frogs are typical of the pre-diabetes phenotype and could serve as a starting point for field studies to determine the role of ED in the decline of amphibian populations. From an environmental perspective, the response of wild green frogs to different ED (10 µg L-1) suggests that a simple glucose-tolerance test could be used on wild anurans to identify bodies of water polluted with metabolic disruptors that could affect species fitness.

Exposure to a mixture of benzo[a]pyrene and triclosan induces multi-and transgenerational metabolic disorders associated with decreased female investment in reproduction in Silurana (Xenopus) tropicalis.

Animals must partition limited resources between their own growth and subsequent reproduction. Endocrine disruptors (ED) may cause maternal metabolic disorders that decrease successful reproduction and might be responsible for multi- and transgenerational effects in amphibians. We found that the frog Silurana (Xenopus) tropicalis, exposed to environmentally relevant concentrations of benzo[a]pyrene and triclosan throughout its life cycle, produced F1 females with delayed sexual maturity and decreased size and weight. These F1 females displayed a marked metabolic syndrome associated with decreased fasting plasma cholesterol and triglyceride concentrations and decreased gonadal development. F1 females from F0 exposed animals also had decreased reproductive investment highlighted by a decrease of oocyte lipid reserves associated with significant F2-tadpole mortality. F2 females from F0 exposed animals also displayed a marked metabolic syndrome but were able to correctly direct liver lipid metabolism to the constitution of fat bodies and oocyte yolk stores. In addition, the F2 females produced progeny that had normal mortality levels at 5 days post hatching compared to the controls suggesting a good reproductive investment. Our data confirmed that these ED, at concentrations often found in natural ponds, can induce multi- and transgenerational metabolic disorders in the progeny of amphibians that are not directly exposed. We present a hypothesis to explain the transmission of the metabolic syndrome across generations through modification of egg reserves. However, when high mortality occurred at the tadpole stage, surviving females were able to cope with metabolic costs and produce viable progeny through sufficient investment in the contents of oocyte reserves.

Transgenerational metabolic disorders and reproduction defects induced by benzo[a]pyrene in Xenopus tropicalis.

Metabolic disorders induced by endocrine disruptors (ED) may contribute to amphibian population declines but no transgenerational studies have evaluated this hypothesis. Here we show that Xenopus tropicalis, exposed from the tadpole stage, to the ED benzo[a]pyrene (BaP, 50 ng.L-1) produced F2 progeny with delayed metamorphosis and sexual maturity. At the adult stage, F2-BaP females displayed fatty liver with inflammation, tissue disorganization and metabolomic and transcriptomic signatures typical of nonalcoholic steato-hepatitis (NASH). This phenotype, similar to that observed in F0 and F1 females, was accompanied by a pancreatic insulin secretory defect. Metabolic disrupted F2-BaP females laid eggs with metabolite contents significantly different from the control and these eggs did not produce viable progeny. This study demonstrated that an ED can induce transgenerational disruption of metabolism and population collapse in amphibians under laboratory conditions. These results show that ED benzo[a]pyrene can impact metabolism over multiple generations and support epidemiological studies implicating environmental EDs in metabolic diseases in humans.

Responses of cytochrome P450, GST, and MXR in the mollusk Corbicula fluminea to the exposure to hospital wastewater effluents.

Pharmaceutical products are a major group of chemical compounds that are continuously released into the environment. The primary pathway of pharmaceuticals to the aquatic environment is the discharge of wastewater effluents. The Psychiatric hospital of Montpon (Dordogne, France) operates with its wastewater treatment plant. We first evaluated the presence and concentrations of 27 pharmaceuticals compounds in these effluents. All of the 27 compounds were detected in these wastewater effluents at concentrations ranging between 37,500 ng L(-1) (paracetamol) and 150 ng L(-1) (citalopram). The aim of the study was then to evaluate the exposure effects of the effluents on cytochrome P450, GST, and MXR responses in Corbicula fluminea gills and digestive glands. Experiments on clams exposed during 1, 3, 7 14, and 21 days revealed a strong and continuous overexpression of mdr1 (multidrug resistant 1) gene expression in gills and transitory variations in pi-gst expression and GST activity. EROD activity increased also transitory after 1 day in the digestive gland of exposed clams. These results indicated that in the effluent, some molecules have undergone metabolism of phase 1 and/or phase 2.

Cross-effects of nickel contamination and parasitism on zebra mussel physiology.

Aquatic organisms are exposed to pollution which may make them more susceptible to infections and diseases. The present investigation evaluated effects of nickel contamination and parasitism (ciliates Ophryoglena spp. and intracellular bacteria Rickettsiales-like organisms), alone and in combination, on biological responses of the zebra mussel Dreissena polymorpha, and also the infestation abilities of parasites, under laboratory controlled conditions. Results showed that after 48 h, more organisms were infected in nickel-exposed groups, which could be related to weakening of their immune system. Acting separately, nickel contamination and infections were already stressful conditions; however, their combined action caused stronger biological responses in zebra mussels. Our data, therefore, confirm that the parasitism in D. polymorpha represents a potential confounding factor in ecotoxicological studies that involve this bivalve.

Seasonal variability of antioxidant biomarkers and energy reserves in the freshwater gammarid Gammarus roeseli.

In gammarids, behavioural and biochemical biomarkers are commonly used in ecotoxicological studies. In our study, we have investigated seasonal variations of several biochemical biomarkers in Gammarus roeseli, a freshwater species. Organisms were sampled monthly over a 1-year period. Gender was distinguished to measure antioxidant enzyme activities like total glutathione peroxidase (GPxtot), selenium-dependent glutathione peroxidase (SeGPx) and catalase enzymes, lipoperoxidation end-product (malondialdehyde, MDA), and energy reserves with protein and lipid contents. In the same time, usual water physico-chemical parameters were measured at the sampling site. A based-gender difference was observed for parameters related to oxidative stress. Females showed higher antioxidant enzyme activities and lower MDA level than males. Parameters related to oxidative stress and energy reserves appeared correlated with temperature and physiological status of organisms. Females GPx activities were lower in autumn and winter when no breeding occurred. In both gender, MDA levels were correlated with temperature with an increase of lipoperoxidation in summer. Lipid contents were the lowest in summer and the highest in winter, probably due to the reproductive status of organisms and their feeding behaviour. Gender-based differences of biochemical parameters suggest a specific sensitivity of males and females in ecotoxicological experiments. Moreover, organisms could be more vulnerable in summer when MDA levels are high and energy reserves low. Deleterious effect of xenobiotics would be different with gender and season.

Effects of sublethal copper exposure on two gammarid species: which is the best competitor?

Biomarker responses in organisms exposed to copper were examined by comparing two gammarid species, Gammarus roeseli and Dikerogammarus villosus, based on gender. G. roeseli specimens were exposed to 20 µg/L of copper for 6, 12, 24 and 48-h periods, while D. villosus were exposed to 20 and 30 µg/L of copper for 12, 48 and 72 h. Males and females of each species were exposed separately and biomarker measurements were performed for each species and gender. The selected biomarkers were antioxidant enzymes as total glutathione peroxidase (GPxtot), selenium-dependent glutathione peroxidase (SeGPx), and catalase activities. Malondialdehyde level (MDA) was measured as a biomarker of toxic effect. Energy reserves were evaluated by means of lipid, glycogen and protein levels. For both species and gender, antioxidant enzyme activities were weakly modified by copper exposure and differences were transient. MDA levels were increased in both species and genders in exposed animals compared to controls, when energy reserves were decreased. G. roeseli was more rapidly overwhelmed by copper toxicity while the first response of D. villosus was the mobilization of its energetic content. D. Villosus probably has specific physiological properties that enable it to cope with copper toxicity and thus become the best competitor.

Effect of gender on physiological and behavioural responses of Gammarus roeseli (Crustacea Amphipoda) to salinity and temperature.

The importance of potentially interacting factors in organisms responses to a stress are often ignored or underestimated in ecotoxicology. In laboratory experiments we investigated how gender, temperature and age influence the behaviour and the physiology of the freshwater amphipod Gammarus roeseli under salinity stress. Our results revealed a significant higher sensitivity of females in survival, ventilation and ionoregulation whereas no inter-age differences were reported. Water temperature also exerted a significant effect in survival and ventilation of G. roeseli. Some of those factors appeared to interact significantly. This study provides evidence that gender can affect organisms responses to a stressor and consequently has to be considered while assessing a stress impact. We discussed the potential relationships between biological and behavioural responses.