Vitellogenin, the egg yolk precursor, is a well-known biomarker of endocrine disruption in oviparous vertebrates. In invertebrates, such as bivalves, it has been used in the last 10 years for the same purpose, despite the limited knowledge of invertebrate endocrinology. In bivalves, vitellogenin levels are usually estimated using an indirect technique, alkali labile phosphate (ALP), that assumes that vitellogenin is the most abundant phosphorylated protein in the analyzed tissue. In this study, we applied shotgun proteomics for the identification and quantification of vitellogenin in marine mussel gonads and compared the results with those obtained with the ALP method. The proteomic analysis revealed that vitellogenin is only detected in female gonads with expression levels that are rather variable among female mussels at different stages of gonad development. ALP analysis, on the contrary, detected similar amounts of phosphorylated proteins regardless of sex or gonad development stage. These results show evidence that the ALP method is not providing reliable information about Vtg levels, at least in marine mussel gonads. ALP is not a good proxy to assess Vtg levels in marine mussels, and careful verification of the adequacy of the procedure should be done before ALP is further assumed as a proxy of Vtg in other bivalve mollusks.

The sea urchin (Paracentrotus lividus) was used to test the effects of one of the most abundant flame retardant additives for plastics, tris (1-chloro-2-propyl) phosphate (TCPP), and the synthetic hormone ethinylestradiol (EE2) on gametogenesis and gonad development of adults. With this aim, 403 individuals of both sexes were exposed to TCPP concentrations ranging from 0.2 to 10 µg/L, EE2 (0.01 µg/L), seawater and solvent controls for 7 and 28 days. EE2 and TCPP exposure did not cause histological damage in the gonads. Some evidence of estrogenic effects of TCPP within the µg/L range and EE2 within the ng/L range is reported. Females exposed to 1 µg/L TCPP for 7 days showed a significant increase in gonad development assessed as gametogenic stage, females exposed to 10 µg/L TCPP showed increased gonad development both in terms of weight (Gonad Index, GI, at both 7 and 28 days) and maturation (Pixelar Index, PI), and females exposed to 10 ng/L EE2 showed increased PI after 28 days exposure. Male sea urchins exposed to both TCPP and EE2 for 7 days showed increased frequencies of low development gonad stage. However, the patterns of response are affected by the high inter-individual variability, the differing initial stage of the gonad, as well as the dosage administered.

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