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Mycoplasma mycoides subsp mycoides (Mmm) is a major pathogen of cattle, causing contagious bovine pleuropneumonia (CBPP), a severe–frequently fatal-disease that is widespread in sub-tropical countries. Despite recognition as a pathogen in the 19th century and advances made through genome sequencing and synthetic biology, many of the molecular characteristics of Mmm remain poorly characterized. Using a proteomics approach, this study aimed to define Mmm major protein determinants, including surface membrane proteins and those involved in host colonization and pathogenicity. For this purpose, both whole cells and a Triton X-114 enriched membrane fraction of a highly pathogenic African field strain (N6) and a vaccine strain (KH3J) were analysed and compared using gelbased methodologies i.e. 2-DGE and SDS-PAGE in combination with MALDI-MS/MS and LC-ESI-MS/MS.