Abstract The virulence of bacteria can be evaluated through both phenotypic and molecular assays. We applied these techniques to 114 strains of Salmonella enterica subsp. enterica collected from July 2010 to June 2012. Salmonella strains were of human origin (71/114) or isolated from food (43/114). The strain set included only the three predominant Salmonella serovars isolated in Italy from humans (S. Enteritidis, S. Typhimurium, S. 4,[5],12:i:-). These strains were screened via polymerase chain reaction for 12 virulence factors (gipA, gtgB, sopE, sspH1, sspH2, sodC1, gtgE, spvC, pefA, mig5, rck, srgA), while antimicrobial sensitivity was evaluated through the Kirby-Bauer assay. Fifty-nine different virulence profiles were highlighted; the genes showing the highest homology were those related to the presence of prophages (gipA, gtgB, sopE, sspH1, sspH2, sodC1, gtgE), while the genes related to the presence of plasmids were less frequently detected (spvC, pefA, mig5, rck, srgA). The Salmonella serovars Typhimurium and 4,[5],12:i:- were closely related in terms of both virulotyping and antibiotic resistance. S. Enteritidis showed higher antibiotic sensitivity and a higher prevalence of genes related to plasmids.