Biomarkers in Solea Senegalensis Kaup, 1858 exposed to contaminated estuarine sediments: a multi-level approach

Thesis submitted to the Universidade Nova de Lisboa, Faculdade de Ciências e Tecnologia for the degree of Doctor of Philosophy in Environmental Sciences

Estuarine sediments can be a reservoir of contaminants from several sources, among which can be found pollutants of anthropogenic origin that, under certain circumstances, may be returned available to the biota. Nevertheless, assessment of the toxicological potential of sediment-bound xenobiotics has many constraints, especially related to the complex geochemical nature of the sediment matrix and to the potential existence of multiple classes of contaminants. In order to contribute to a weight-of-evidence approach to assess the ecological risk of estuarine sediments, an array of 28-day bioassays was enforced, in situ and in the laboratory, using juvenile Solea senegalensis as test subjects. The battery of assays aimed at contributing with two lines-of-evidence for an ecological risk assessment approach to the Sado Estuary, namely sediment chemistry and toxicity. For the purpose, three sites were selected (a reference plus two contaminated), from which sediments were collected and analysed for physical parameters and contaminant concentrations (both metallic and organic). Having the liver of tested fish been surveyed as the main target organ, several biomarkers were analysed, corresponding to different levels of biological organization: molecular, cellular, histological and organ itself. Several methods were employed, from the classic immune- and electrochemical methods to determine CYP1A and metallothionein induction, respectively; to more state-of-the--art approaches that are now being employed to find and validate new sets of biomarkers, namely “omics” - toxicogenomics and proteomics, besides semi-quantitative histopathological methods, genotoxicity assessment and others. The integration of biological responses with sediment parameters revealed that, although differences between in situ and laboratory assays exist, the biomarkers that in essence reflect some measure of lesion allow a much more consistent distinction between contaminated and uncontaminated sediments. In addition, molecular biomarkers permitted inferring patterns of metabolic change and assess how these changes contribute to the impairment of the response machinery to chemical insult, from apoptosis to anti-oxidative defence, among others. The multi-level approach was complemented by a series of laboratory assays with model xenobiotics, isolated or combined (cadmium + benzo[a]pyrene), aiming at technique testing and set-up and, most importantly, at surveying the complex mechanisms underlying contaminant interactions. The results demonstrate that even moderately contaminated sediments can cause adverse effects to organisms and trigger responses that reflect the intricate machinery beneath exposure to complex mixtures of xenobiotics, either for monitoring or for more mechanistic studies, especially when the existence of multiple contaminants tends to dilute biomarker specificity.

Fundação para a Ciência e Tecnologia fellowship, reference number SFRH/BD/28465/2006