A FAMILY OF PHASE-VARIANT AND SIZE-VARIANT MEMBRANE-SURFACE LIPOPROTEIN ANTIGENS (VSPS) OF MYCOPLASMA-BOVIS

A set of strain- and size-variant highly immunogenic membrane surface protein antigens of Mycoplasma bovis, which has been identified by a m onoclonal antibody, is shown in this report to make up a family of ant igenically and structurally related lipid-modified proteins, designate d Vsps (variable surface proteins). By systematic analysis of several isogenic clonal lineages of the type strain PG45, three members of thi s family have been identified, VspA, VspB, and VspC, each of which was shown to undergo independent high-frequency changes in size as well a s noncoordinate phase variation between ON and OFF expression states. The monoclonal antibody-defined epitope common to VspA, VspB, and VspC was accessible on the tell surface in most, but not all, of the clona l populations analyzed and was present on a C-terminal limit tryptic f ragment of each Vsp variant that was released from the membrane surfac e. VspA and VspC were distinguished from VspB by their selective detec tion with colloidal gold and by their distinctive reaction with a poly clonal antibody against M. bovis D490. VspA, VspB, and VspC were furth er distinguishable from one another by their characteristic patterns o f degradation at carboxypeptidase Y pause sites. While these Vsp-speci fic structural fingerprints with an irregular periodic spacing were co nstant for similarly sized variants of a defined Vsp product, they sho wed distinct differences among variants differing in size. This variab ility included gain or loss of individual bands within distinct subset s of bands, as well as shifts of the entire banding patterns up- or do wnwards, indicating that insertions or deletions underlying Vsp size v ariation can occur at various locations either within the C-terminal d omain or within other regions of these proteins. This was similarly co nfirmed by comparative epitope mapping analysis of tryptic cleavage pr oducts generated from different Vsp size variants. The Vsp family of M . bovis described in this study represents a newly discovered system o f surface antigenic variation in mycoplasmas displaying features which closely resemble but are also different from the characteristics repo rted for the Vip (variable lipoprotein) system of M. hyorhinis. The is ogenic lineages established here provide key populations for subsequen t analysis of corresponding genes to further elucidate Vsp structure a nd variation, which may have important relevance for a better understa nding of the pathogenicity of this agent.