Tetrameric Neuraminidase of Influenza A Virus Is Required to Induce Protective Antibody Responses in Mice

Influenza neuraminidase (NA) is able to induce cross-subtype immunity and is considered as a promising target for the development of universal influenza vaccines. However, commercial influenza vaccines only induced low NA-specific immune responses due to the low amounts and the denatured conformation of NA proteins in current inactivated or split influenza vaccines. Here we investigated the protective efficacy of recombinant tetrameric and monomeric NA proteins to determine whether the conformation contributed to induce protective immunity. We found that H1N1(P)(R)(8)NA tetramer (NA(tet)) could provide complete homologous protection against A/PR8 (H1N1) virus infection in mice, while the protection of H1N1(P)(R)(8)NA monomer (NA(mono)) was moderate. Higher levels of NA-reactive binding and inhibition antibodies and less weight loss were observed in the H1N1(P)(R)(8)NA(tet)-vaccinated group. Similarly, H5N1(V)(N)NA(tet) immunization exhibited a preferable heterologous protection than H5N1(V)(N)NA(mono), but neither H7N9(S)(H)NA(tet) nor H7N9(S)(H)NA(mono) vaccination showed heterosubtypic protection. We also compared the effect of three adjuvants, aluminum, 3′3′-cGAMP (cGAMP), and Poly(I:C), on the humoral response and protective efficacy induced by H1N1(P)(R)(8)NA(tet). H1N1(P)(R)(8)NA(tet) protein adjuvanted with aluminum was observed to exhibited better capacity in inducing NA-specific humoral immunity and preventing weight loss than with cGAMP or Poly(I:C). In conclusion, our data demonstrate that tetrameric NA with natural conformation is required to induce protective anti-NA immunity. The NA tetramer could provide homologous protection and subtype-specific cross-protection. In addition, the aluminum adjuvant is preferable in recombinant NA protein vaccination.