Immune responses to protein PPE44 (Rv2770c) in mice infected with Mycobacterium bovis BCG

Background The PPE protein family of Mycobacterium tuberculosis includes 69 glycine-rich proteins. Their role in tuberculosis is unknown, but it has been speculated that they may have an important immunological significance. In this investigation, the immunogenicity of the ppe44 (Rv2770c) gene product was evaluated in mice infected with Mycobacterium bovis BCG. Moreover, the protective efficacy of a DNA vaccine expressing ppe44 was evaluated in mice. Methods The gene ppe44 of M. tuberculosis H37Rv was cloned in E. coli and the purified recombinant protein (rPPE44) was used to evaluate antibody, cytokine and DTH responses in BALB/c mice infected with M. bovis BCG. The potential protective efficacy of PPE44 was evaluated in BALB/c and C57Bl/6 mice immunized intramuscularly with a DNA vaccine expressing ppe44 (ppe44-DNA) and challenged intravenously with M. bovis BCG. Results BALB/c mice infected subcutaneously developed high titers of anti-rPPE44 IgG1, but no IgG2a antibodies. Cultures of rPPE44-primed cells from draining lymph nodes and spleen produced low levels of IFN-; a moderate degree of DTH was observed following rPPE44 intracutaneous challenge. Similar results were obtained in mice infected intravenously. BALB/c mice immunized with ppe44-DNA developed anti-rPPE44 IgG1 predominant over IgG2a antibodies; rPPE44-primed spleen cell cultures did not produce significant levels of IFN-. This regime of immunization conferred a moderate protection against an intravenous challenge with M. bovis BCG, as shown by the reduction of CFUs in the lungs compared to control mice immunized with the empty vector. C57Bl/6 mice immunized with ppe44-DNA developed comparable titers of anti-rPPE44 IgG1 and IgG2c antibodies; rPPE44-primed spleen cell cultures produced IFN-but no IL-4. After an intravenous challenge with M. bovis BCG, a one-log10 reduction of CFUs was detected in the lungs of ppe44-DNA immunized animals. Conclusions PPE44 represents a novel mycobacterial antigen expressed during infection by M. bovis BCG in mice. Both subcutaneous and intravenous infections seem to polarize the immune response to PPE44 towards a Th2 phenotype. Immunization of mice with ppe44-DNA confers a partial protection against an intravenous challenge with M. bovis BCG. The protective efficacy of PPE44 needs to be further evaluated by other experimental models aimed at enhancing the Th1-type immune responsiveness.