Linking embryo toxicity with genotoxic responses in the freshwater snail Physa acuta: Single exposure to benzo(a)pyrene, fluoxetine, bisphenol A, vinclozolin and exposure to binary mixtures with benzo(a)pyrene

Genotoxic effects on fauna after waterborne pollutant exposure have been demonstrated by numerous research programmes. Less effort has been focused on establishing relationship between genotoxicity and long-term responses at higher levels of biological organization. Taking into account that embryos may be more sensitive indicators of reproductive impairment than alterations in fertility, we have developed two assays in multiwell plates to address correlations between embryo toxicity and genotoxicity. The potential teratogenicity was assessed by analyzing abnormal development and mortality of Physa acuta at embryonic stage. Genotoxicity was measured by the micronucleus (MN) test using embryonic cells. Our results showed that linkage between genotoxicity and embryo toxicity depends on mechanisms of action of compounds under study. Embryo toxic responses showed a clear dose-related tendency whereas no clear dose-dependent effect was observed in micronucleus induction. The higher embryo toxicity was produced by benzo(a)pyrene exposure followed by fluoxetine and bisphenol A. Vinclozolin was the lower embryo toxic compound. Binary mixtures with BaP always resulted in higher embryo toxicity than single exposures but antagonistic effects were observed for MN induction. Benzo(a)pyrene produced the higher MN induction at 0.04 mg/L, which also produced clear embryo toxic effects. Fluoxetine did not induce cytogenetic effects but 0.25 mg/L altered embryonic development. Bisphenol A significantly reduced hatchability at 0.5 mg/L while MN induction appeared with higher treatments than those that start causing teratogenicity. Much higher concentration of vinclozolin (5 mg/L) reduced hatchability and induced maximum MN formation. In conclusion, while validating one biomarker of genotoxicity and employing one ecologically relevant effect, we have evaluated the relative sensitivity of a freshwater mollusc for a range of chemicals. The embryo toxicity test is a starting point for the development of a life cycle test with freshwater snails even for undertaking multigeneration studies focused on transgenerational effects.

► Linkage between genotoxicity and embryo toxicity depends on mechanism of action.
► Micronucleus test could be used as a short-term assay for warning teratogenic effects.
► Benzo(a)pyrene>fluoxetine>bisphenol A>vinclozolin was the embryo toxicological order.
► Co-exposure with benzo(a)pyrene enhanced embryo toxicity.
► Mixtures of benzo(a)pyrene and bisphenol A produced antagonist effects in micronuclei.