The aim of our study was to describe the balance between health benefits and risks associated with the consumption of crawfish and nine fish species from lake Trasimeno. We thus determined both fatty acid profiles (particularly, eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids) and chemical pollutants (some polychlorinated biphenyls, pesticides, and heavy metals) in fish muscle tissues. The contents of all fatty acids varied significantly among species. Sand smelt, carp, and tench, which have a high fat content, contained considerable amounts of EPA and DHA; lean fish, like perch, pike, and largemouth bass, which have relatively high percentages of the predominant n-3 fatty acids EPA and DHA, showed lower amounts of these fatty acids because of their low lipid contents. Some species contributed strongly to the Dietary Reference Intake (RDI) of EPA and DHA. The contribution of lean fish to the RDI of EPA and DHA was more limited. The concentrations of all contaminants in fish muscle tissues were lower than the regulatory limits, demonstrating the safety of the environmental conditions of the lake. The contribution to health-based reference values and benefit–risk quotients indicated that the health benefits of consumption of fish from lake Trasimeno outweigh the potential risks.