Several environmental chemicals are suspected to be responsible for adverse health effects on the reproductive system in various organisms. During this work, environmentally relevant concentrations of North Sea oil were used alone or in combination with alkylphenols and additional PAH to study the effect on vitellogenin-like protein expression and gonadal development in mussels. North Sea oil (0.5 ppm) induced the expression of phospho-proteins in both sexes indicating that some compounds are oestrogen-mimics. This induction was not seen in samples dosed with the mixture but signs of toxic effects were observed in the gonads. Indeed, numerous degenerating ovarian follicles in females and foci, similar to vertebrate melanomacrophage centres, were observed in testes.
"Several characteristics such as filter feeding, sessile life form and ability to accumulate pollutants in addition to their wide distribution, makes them favored organisms for estimating environmental pollution levels  . With a range of physiological, histological and molecular responses, including abnormal morphology , alterations of anti-oxidative status, induction of DNA strand-breaks, etc., they have been shown to be applicable for in situ and ex situ assessment of the effects of pollutants present in environment   . Most importantly, they are widely employed for assessing genotoxicity    . "
[Show abstract][Hide abstract] ABSTRACT: Specimens of the mussel Mytilus galloprovincialis were collected from five sites in the Boka Kotorska Bay (Adriatic Sea, Montenegro) during the period summer 2011-autumn 2012. Three types of tissue, haemolymph, digestive gland were used for assessment of DNA damage. Images of randomly selected cells were analyzed with a fluorescence microscope and image analysis by the Comet Assay IV Image-analysis system. Three parameters, viz, tail length, tail intensity and Olive tail moment were analyzed on 4200 nuclei per cell type. We observed variations in the level of DNA damage in mussels collected at different sites, as well as seasonal variations in response. Sum of ranking differences (SRD) was implemented to compare use of different types of cell and different measure of comet tail per nucleus. Numerical scales were transferred into ranks, range scaling between 0 and 1; standardization and normalization were carried out. SRD selected the best (and worst) combinations: tail moment is the best for all data treatment and for all organs; second best is tail length, and intensity ranks third (except for digestive gland). The differences were significant at the 5% level. Whereas gills and haemolymph cells do not differ significantly, cells of the digestive gland are much more suitable to estimate genotoxicity. Variance analysis decomposed the effect of different factors on the SRD values. This unique combination has provided not only the relative importance of factors, but also an overall evaluation: the best evaluation method, the best data pre-treatment, etc., were chosen even for partially contradictory data. The rank transformation is superior to any other way of scaling, which is proven by ordering the SRD values by SRD again, and by cross validation.
"An heterosynthesis merges also from the assay of alkali-labile phosphate (ALP), an indirect technique utilized to determine the presence of VTG-like proteins (Blaise et al., 1999; Pampanin et al., 2005; Ricciardi et al., 2008). Indeed after ALP, the presence of vitelline-like proteins was also reported in the hemolymph of several molluscs treated with estrogen (Blaise et al., 1999; Gagné et al., 2001) or exposed to environmental contaminants (endocrine-disrupting chemicals) known to induce changes and alterations of the endocrine functions (Blaise et al., 1999; Gagné et al., 2002; Verslycke et al., 2002; Aarab et al., 2004, 2006; Marin and Matozzo, 2004; Quinn et al., 2004; Versonnen et al., 2004; Matozzo and Marin, 2005; Pampanin et al., 2005; Ortiz-Zarragoitia and Cajaraville, 2006; Zorita et al., 2006; Matozzo et al., 2008). "
"Regarding the use of VTG as a biomarker in invertebrates, studies on the effect of hydrocarbons on protein-like-vitellogenin are restricted to the bivalve mollusk Mytilus edulis exposed to crude oil (Ortiz-Zarragoitia and Cajaraville, 2006; Aarab et al., 2004) where the amount of vitellogenin was determined using alkali-labile phosphate, an indirect marker of vitellin. Though this assay is considered as an alternative assessment of ELISA, this method should be applied with caution as alkali-labile phosphorus is a minor part of the total phosphorus pool in crustacean vitellogenin (Volz et al., 2002), and the presence of other phosphorus sources can substantially affect the results (Stanton, 1968). "
[Show abstract][Hide abstract] ABSTRACT: During reproduction vitellogenin (VTG) is transported to vitellogenic oocytes as a precursor of egg yolk lipovitellin (LV). As VTG synthesis is affected by environmental stressors, it is widely used as biomarker in endocrine disruption studies. However, it has seldom been employed to evaluate invertebrate hydrocarbon pollution. An ELISA with anti-LV antibody was developed to evaluate the impact of water-soluble fraction of crude oil (WSF) on Macrobrachium borellii vitellogenesis. Prawn VTG concentration was within the range reported for other crustaceans; LV values were positively correlated with gonadosomatic index (GSI). Females at different vitellogenic stages were exposed to a sub-lethal concentration of WSF for 7 days. Exposed animals with GSI>7 increased their VTG and LV titer as compared to control organisms (190% and 140%, respectively). VTG levels in M. borellii were upregulated and highly sensitive to WSF exposure. This assay could be employed as a biomarker for freshwater hydrocarbon pollution.
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