Abstract: Objective: To analysis the regularities of distribution of Vibrio cholera from food risk monitoring samples and evaluate the merits of VITEK-MS rapid analysis methods in identification of Vibrio cholera. Methods: V. cholera strains were isolated from freshwater products and patients. The VITEK-MS and VITEK-2 identification were operated respectively. The serotype was detected by serum agglutination and immunogold. The virulence genes ctxAB was detected by qPCR. All the virulence genes in different types of strains were analyzed by whole genome sequencing. Results: 17 of the 23 V. cholera strains were non-O1/O139 serotype. 6 strains were detected as O139 V. cholera and ctxAB gene carrier. All the V. cholera strain showed typical mass spectral characteristics based on the fingerprint spectrum. The particular and lacking peaks patterns of O139 V. cholera were representative for Serotype identification. ctxAB, ace and tcp islands genes were harbored in O139 Vibrio cholera, but only auxiliary virulence factor can be detected in non-O1/O139 Vibrio cholera except zot gene. Conclusion: There are many differences between the O139 and non-O1/O139 Vibrio cholera strains in whether virulence gene type or fingerprint spectrum. The thorough application for serotype identification by using microbial mass spectrometry may be a new assistance for cholera surveillance.