Chemical and isotopic data have been used as geochemical tracers for a genetic characterization of hydrocarbon gases from a total of eleven manifestations located in Eastern and Central-Southern Sicily (Italy). The molecular analysis shows that almost all the samples are enriched in methane (up to 93.2% Vol.), with the exception of four gas samples collected around Mt. Etna showing high mantle-derived CO2 content. Methane isotope signatures suggest that these are thermogenic gases or a mixture between thermogenic gases and microbial gases. Although samples from some mud volcanoes in Southern Sicily (Macalube di Aragona) show isotope signatures consistent with a mixing model between thermogenic and microbial, by combining the molecular compositions (C1/(C2 + C3)) and the methane isotope ratios (δ13C1), such a process seems to be excluded. Therefore, the occurrence of secondary post-genetic processes should be invoked. Two main hypotheses have been considered: the first hypothesis includes that the gas is produced by microbial activity and altered post-genetically by microbial oxidation of methane, while according to the second hypothesis thermogenic gas have modified their molecular ratios due to vertical migration.