Over the past decade, molecular, biochemical and cellular markers have been extensively used in pollution monitoring of aquatic environments. Biochemical markers have been selected among early molecular events occurring in the toxicological mechanisms of main contaminants. This paper assesses the marine environment quality along the Tunisian coasts using a statistical approach. Clams (Ruditapes decussatus) were collected during the four seasons of 2003 on seven different sites from the Tunisian coasts. Oxidative stress was evaluated in gills using catalase activity (Cat), neutral lipids and malonedialdehyde accumulation. Glutathione S-transferase activity is related to the conjugation of organic compounds and was evaluated in both, gills and digestive glands. Acetylcholinesterase activity was evaluated as the biomarker of exposure to organophosphorous, carbamate pesticides and heavy metals. For each biomarker, a discriminatory factor was calculated and a response index allocated. For each site, a global response index was calculated as the sum of the response index of each biomarker. Discriminant analysis shows significant differences between sites and seasons compared with control sample. Faroua (site 1) and Menzel Jemile (site 2) seem to be the less polluted with respect to the other sites for all seasons. Gargour (site 6) shows the highest Multimarker Pollution Index during the four seasons, indicating higher contamination level.

In mussel Mytilus galloprovincialis tissues, metallothionein belongs to two different gene classes, mt10 and mt20, showing differential expression at both basal conditions and under heavy metal challenge. In this study, a new more highly sensitive technique, expression analysis of mt10 and mt20 mRNA levels by quantitative reverse transcription polymerase chain reaction, was used to assess the effects of heavy metal contamination in the digestive glands of mussels caged along the Tunisian coast. To validate the new assay, total metallothionein protein, amount of heavy metals (zinc, copper, cadmium), and a biomarker of oxidative stress such as malondialdehyde content, were assessed in the same tissues. At the investigated sites, the molecular assay showed variations of mt20 relative gene expression levels within one or two orders of magnitude, with maximum values at two sites severely polluted with cadmium, Mahres (100-fold) and Menzel Jemile (165-fold). Changes in mt10 expression were recorded at all sites where copper had significantly accumulated, although fold induction levels were less pronounced than those of mt20. In this paper, gene expression data are discussed in relation to the studied biomarkers, demonstrating that the molecular technique based on the differential expression of mt10 and mt20 genes represents (i) a useful and robust tool for studying and monitoring heavy metal pollution under field conditions, and (ii) an improvement in the application of metallothionein as a biomarker of response to exposure to heavy metals in marine mussels.

The potential use of acetylcholinesterase (AChE) and metallothionein (MT) responses as biomarker of organophosphorous (OPs) and trace metal were assessed in fish Seriola dumerilli exposed to 0, 4, 6 mg/kg of malathion for 2, 7 and 13 days, and to 0, 50, 100, 250 mug/kg of Cd for 2 days. Brain AChE was significantly inhibited after 2 and 7 days of malathion exposure, in a dose-response manner, but no inhibition was observed after 13 days of exposure. When exposed to Cd for 2 days, S. dumerelli presented an increase in AChE activity at a concentration of 50 mug/kg, but a strong and dose-dependent AChE inhibition at 100 and 250 mug/kg. Cd treatment also caused a rapid increase in MTs concentration in liver, even at the lower concentration. Our experiments indicate that the measurement of hepatic MT concentration and brain AChE activity in S. dumerilli would be useful biomarkers of OP and Cd exposure and/or effects.

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The levels of metallothionein (MT), a biomarker of metal exposure, and of Cd and Cu, known as MT inducers, were investigated in Sparus aurata intraperitoneally injected with 500 microg/kg of Cu and Cd for 2 days. MT and metal concentrations (Cd and Cu) were determined in liver, gills and kidney. MT levels were significantly increased in all investigated tissues, with the highest value in liver of Cu as Cd-treated fishes (3.56-fold and 3.3- fold, respectively). Metal concentrations were statistically different between all tissues. Highest metal concentrations were in the liver. The higher metal concentrations and MT induction levels support the main role of MT in metal homeostasis and detoxification.