EXPRESSION OF THE COPB OUTER MEMBRANE PROTEIN BY MORAXELLA CATARRHALIS IS REGULATED BY IRON AND AFFECTS IRON ACQUISITION FROM TRANSFERRIN AND LACTOFERRIN. • 974

Moraxella catarrhalis is a major etiologic agent of acute otitis media and chronic otitis media with effusion. The 81 kDa CopB outer membrane protein (OMP) of this pathogen is a promising vaccine candidate as it is surface-exposed, antigenically conserved and induces a bactericidal humoral immune response in mice. A monoclonal antibody directed against a surface-exposed CopB epitope of most strains of M. catarrhalis was shown to enhance pulmonary clearance in immunized mice. In this study, the role of CopB in iron acquisition by M. catarrhalis is described. The amino acid sequence is homologous to that of the FrpB protein of N. meningitidis and N. gonorrhoeae (overall identity 49% and 52%, respectively), whose function is unknown, and similar to those of TonB-dependent OMP. In several M. catarrhalis strains tested, the expression of CopB was upregulated by iron limitation in the growth medium. These strains were able to utilize iron from human transferrin (Tf) and lactoferrin (Lf), ferric citrate and heme, while being unable to grow with hemoglobin, hemoglobin-haptoglobin and heme-hemopexin as iron sources. An isogenic mutant of M. catarrhalis strain O35E lacking CopB expression was previously shown to be unable to survive in the mouse lung. This mutant was severely impaired in its ability to utilize Tf and Lf for growth in vitro. Similarly, the ability of this copB mutant to take up ⁵⁵Fe from Tf and Lf was 29% and 39%, respectively, when compared with the wild-type strain. Growth and iron uptake from ferric citrate and heme were unaffected. Binding of horseradish-peroxidase-coupled Tf and Lf to the cell surface was similar in both O35E wild-type strain and copB mutant. These results indicate that CopB, which is essential for survival ofM. catarrhalis in the mouse lung, plays a role in the iron acquisition from its main in vivo iron sources, Tf and Lf, without being directly involved in binding of these proteins to the bacterial surface. This finding reinforces the validity of CopB for further evaluation as a vaccine candidate.