Immunization with pneumococcal neuraminidases NanA, NanB and NanC to generate neutralizing antibodies and to increase survival in mice.

Pneumococcal virulence protein-based vaccines can provide serotype-independent protection against pneumococcal infections. Many studies, including clinical observational studies on Thomsen-Friedenrich antigen exposure and haemolytic uremic syndrome, defined the role of neuraminidases NanA, NanB and NanC in host-pneumococcus interaction. Since neuraminidases are major virulence proteins, they are potential targets for both vaccines and small molecule inhibitors. Here we explored the utility of three neuraminidases as protein vaccine antigens to generate neutralizing antibodies and to increase survival following pneumococcal infections. Rabbits and mice were immunized subcutaneously with enzymatically active recombinant NanA, NanB and NanC as individual or a combination of the three neuraminidases. Antisera titres were determined by ELISA. Neuraminidase activity inhibition by antiserum was tested by peanut lectin and flow cytometry. Clinical isolates with serotype 3, 6B, 14, 15B, 19A and 23F were used to infect immunized mice by tail vein injection. Presence of high levels of IgG antibodies in antisera against NanA, NanB and NanC indicates that all of the three neuraminidases are immunogenic vaccine antigens. To generate potent NanA neutralizing antibodies, both lectin and catalytic domains are essential, whereas for NanB and NanC a single lectin domain is sufficient. Immunization with triple neuraminidases increased the survival of mice when intravenously challenged with clinical isolates of serotype 3 (40 %), 6B (60 %), 15B (60 %), 19A (40 %) and 23F (30 %). We recommend the inclusion of three pneumococcal neuraminidases in future protein vaccine formulations to prevent invasive pneumococcal infection caused by various serotypes.