Development of a recombinant antigen for antibody-based diagnosis of Mycoplasma bovis infection in cattle

Mycoplasma bovis induces various clinical manifestations in cattle, such as mastitis, arthritis, and pneumonia. We have evaluated the immunoreactivity of three variable surface proteins (Vsps) of M. bovis, namely VspA, VspB, and VspC, with sera collected from herds with mycoplasmosis or from cattle experimentally infected with M. bovis. Western blot analysis revealed that the Vsps are the predominant antigens recognized by the host humoral respon se during M. bovis infection. The immunoreactivity of VspA, VspB, and VspC with host antibodies was independent of the clinical manifestations, the ge ographical origin of the M. bovis isolates, the mode of infection, and the animal's history. Moreover, the results showed that Vsp-specific host antib odies can be detected about 10 days after experimental infection and for up to several months. The full-length or truncated versions of the VspA produ ct were overexpressed in Escherichia coli as fusion proteins (FP-VspA). Rec ombinant products showed strong immunoreactivity,vith the Vsp-specific mono clonal antibodies 1A1 and 1E5, with the corresponding epitopes localized at the VspA N-terminal and C-terminal ends, respectively. Anti-M. bovis sera of cattle naturally or experimentally infected also strongly recognized the full-length FP-VspA. The seroreactivity of sera collected from cattle betw een 6 and 10 days after experimental infection was weaker with truncated ve rsions of VspA lacking the 1E5 epitope than with the full-length VspA or th e truncated versions lacking the 1A1 epitope. Overall, the results indicate that the Vsps, despite their inter- and intraclonal variability, may be ap plied as target antigens in serodiagnostic assays for epidemiological studi es.