Objective To determine the main serogroups, virulence genes and molecular typing of Listeria monocytogenes isolates in Wenzhou, and investigate clustering patterns and pathogenicity of the local strains. Methods Polymerase chain reaction(PCR) was applied to determine the serogroups and virulence genes of the isolates. Multilocus sequence typing (MLST) was used to identify molecular types of the strains. Results Ninety-seven strains were divided into four serogroups. The major serogroups were 1/2b(48.45%,47/97) and 1/2a(35.05%,34/97). The positive rate of pathogenicity gene iap and prfA were both 100.00%(97/97). All strains from patients carried five virulence genes,the overall hemolysin gene hlyA and inlA were 97.94%(95/97) and plcB was 96.91%(94/97). These ninety-seven stains were divided into twenty MLST types including the prevalent ST87, ST121 and ST9, patient-specific ST1 and ST779 as well as ST2, ST3 and ST5 that were found both in food and in patients. Conclusion This result demonstrated a high prevalence and genetic polymorphism of Listeria monocytogenes from diverse sources in Wenzhou. The majority of isolates carried pathogenicity-related genes, which posed a potential threat to the public health. The surveillance and management should be strengthened to prevent the infectious listeriosis.