Mollicutes are minute-bacteria with a reduced genome that colonize various hosts, including plants, animals and human. Because they lack the cell-wall found in other bacteria, the surface of their single membrane has emerged as a key interface in mediating interactions with their host. While genomic data are piling up, not much is known regarding the expression and the conservation of surface proteins across Mollicutes. In order to fill this gap, the membrane proteome of three species was compared. These species were (i) Mycoplasma mycoides subsp. mycoides SC (MmmSC) and Mycoplasma agalactiae that belong to different phylogenetic groups but share the ruminant host and (ii) Spiroplasma citri that clusters with MmmSC in the same phylogenetic group but is an insect-transmitted plant pathogen. Mass spectrometry detection was applied to lipoprotein-enriched Triton X-114 fractions and on membrane-enriched fractions. No lipoprotein and only 24 transmembrane proteins were detected in the predicted common set of 285 proteins. These transmembrane proteins were part of F1F0 ATP synthase, V-type ATPase, secY secretion pathway and several ABC transporters. About 60-70% of the predicted lipoproteins were detected in all three species. Surface subproteome shared by the two ruminant mycoplasmas contains 12 lipoproteins and 16 transmembrane proteins encoded by genes potentially exchanged by horizontal gene transfers (HGT). In S. citri a large number of lipoproteins encoded on plasmids and/or of viral origin were expressed. Taken altogether, these data suggest that HGT, as well as plasmid and phage genome integration events may have important impact on host-pathogen interactions and adaptation by (re)-shaping part of bacterial surface architecture. This proteomic study provides clues regarding surface proteins that are involved in common or specific functions across Mollicutes depending on their evolutionary pathways, which include vertical and horizontal inheritance as well as host-adaptation. NB: LB, CC and PSP contributed equally to the work.