In Mediterranean coastal lagoons, the combination of human impact and wide variability of natural environmental factors can lead to upsets in ecosystem homeostasis resulting in biodiversity decline. Oxidative damage has been causally linked to various kinds of environmental stress, both natural and artificial, the result being impairment of cellular functions. DNA damage and the efficiency of antioxidant defences in Mytilus galloprovincialis from the highly eutrophicated Orbetello Lagoon (Tuscany, Italy) were investigated, respectively by the single cell gel electrophoresis (or Comet test) and the total oxyradical scavenging capacity assay. Results showed significantly higher levels of DNA damage in mussels collected from the inner parts of the lagoon compared to specimens from more external sites. Specimens with the lower genetic integrity also exhibited a reduced efficiency in neutralizing three potent cellular oxidizing species, namely peroxyl radicals (ROO*), hydroxyl radicals (*OH) and peroxynitrite (HOONO), suggesting the involvement of reactive oxygen species in mediating the genetic damage. The analyzed biological parameters also showed a seasonal variability with a minimum of both DNA integrity and antioxidant scavenging efficiency during the warm months and an opposite trend in winter. The potential of analyzed techniques is discussed for the assessment of both anthropogenic and natural disturbance.