The aim of the present work was to evaluate the expression of 8-OHdG (8-hydroxydeoxyguanosine) in the benthic fish Zosterisessor ophiocephalus collected in two differently polluted sites of the Venetian lagoon (Porto Marghera and Caroman). We compared our data on 8-OHdG with those of CYP1A (Cytochrome P450, family 1, subfamily A, polypeptide 1), which is a well known biomarker for detoxification of contaminants. Immunohistochemistry with an antibody to 8-OHdG showed immunopositivity in nuclei of hepatocytes as well as in melanomacrophage centres of spleen and kidney, whereas an anti-CYP1A antibody exhibited positive immunostaining in the liver, kidney and ovary. The liver of males showed higher expression of both proteins than females. In animals from Porto Marghera site, the enzymatic assay for 8-OHdG exhibited higher levels in liver of males than in females. Western Blot analysis using the antibody anti-CYP1A recognized the presence of a band of about 60 kDa in the liver of males and females. Males exhibited a strong band, whereas in females the band showed a lower intensity. By using Real-Time PCR, the mRNA expression of CYP1A did not show any differences between males and females from each site, but it was at borderline significance level. Comparing the two sites, mRNA expression of CYP1A was significantly higher in the liver of both males and females from Porto Marghera than that of Caroman. The present data suggest that pollutants are bio-available as demonstrated by our biomarker analyses and may have a harmful effect on aquatic organisms such as Z. ophiocephalus. We report that the highest levels of hepatic 8-OHdG and CYP1A expression were detected in males, showing clear gender specificity.