Phylogenetic and functional characterization of the MIB-MIP system of different Mollicutes using an engineered Mycoplasma feriruminatoris strain.

The Mycoplasma Immunoglobulin Binding/Protease (MIB-MIP) system is a candidate virulence factor present in multiple pathogenic species of the Mollicutes, including the fast-growing species M. feriruminatoris. The MIB-MIP system cleaves the heavy chain of host immunoglobulins, hence affecting antigen-antibody interactions and potentially facilitating immune evasion. In this work we analyzed the distribution and genetic relatedness between MIB-MIP systems of different species. Using -omics technologies, we showed that the four copies of the M. feriruminatoris MIB-MIP system have different expression levels, are transcribed as operons controlled by four different promotors. We introduced individual MIB-MIP gene pairs of M. feriruminatoris and other Mollicutes in an engineered M. feriruminatoris strain and tested their functionality using oriC-based plasmids. We were able to functionally express the two proteins at the surface of M. feriruminatoris, which confirms our ability to display large functional heterologous surface proteins in M. ferirumintoris. Interestingly, functionality of cloned heterologous MIB-MIP systems could not be achieved with phylogentic distant porcine Mollicutes like Mesomycoplasma hyorhinis or Mesomycoplasma hyopneumoniae. Finally, since M. feriruminatoris is a candidate for biomedical applications such as drug delivery, we confirmed its safety in vivo in domestic goats, which are the closest livestock relatives to its native host the Alpine ibex.