Norovirus (NoV) specific protective immune responses induced by recombinant P dimer vaccine are enhanced by the mucosal adjuvant FlaB.

Vivek Verma,Sao Puth,Kyoung-Oh Cho,Wenzhi Tan,Shee Eun Lee,Joon Haeng Rhee

doi:10.1186/s12967-016-0899-4

Abstract

BackgroundNoroviruses (NoVs) are a major cause of childhood gastroenteritis and foodborne diseases worldwide. Lack of appropriate animal models or cell-based culture systems makes the development and evaluation of NoV-specific vaccines a daunting task. VP1 is the major capsid protein of the NoVs that acts as a binding motif to human histo-blood group antigens (HBGAs) through its protruding 2 (P2) domain and can serve as a protective antigen candidate for vaccine development.MethodsRecombinantly produced NoV specific P domain (Pd) vaccine was inoculated into groups of mice either alone or in conjugation with mucosal adjuvant FlaB, the flagellar protein from Vibrio vulnificus. Antigen specific humoral and cell mediated immune responses were assessed by enzyme linked immunosorbent assay (ELISA) or fluorescent activated cell sorting (FACS). A comparative analysis of various routes of vaccination viz. intranasal, sublingual and subcutaneous, was also done.ResultsIn this study, we show that a recombinant Pd-vaccine administered through intranasal route induced a robust TH2-dependent humoral immune response and that the combination of vaccine with FlaB significantly enhanced the antibody response. Interestingly, FlaB induced a mixed TH1/TH2 type of immune response with a significant induction of IgG1 as well as IgG2a antibodies. FlaB also induced strong IgA responses in serum and feces. FlaB mediated antibody responses were toll like receptor 5 (TLR5) dependent, since the FlaB adjuvanticity was lost in TLR5−/− mice. Further, though the Pd-vaccine by itself failed to induce a cell mediated immune response, the Pd-FlaB combination stimulated a robust CD4+IFNγ+ and CD8+IFNγ+ T cell response in spleen and mesenteric lymph nodes. We also compared the adjuvant effects of FlaB with that of alum and complete Freund’s adjuvant (CFA). We found that subcutaneously inoculated FlaB induced more significant levels of IgG and IgA in both serum and feces compared to alum or CFA in respective samples.ConclusionWe validate the use of TLR5 agonist as a strong mucosal adjuvant that would facilitate the development of NoV specific vaccines for humans and veterinary use. This study also highlights the importance of route of immunization in inducing the appropriate immune responses in mucosal compartments.

Highlights

Noroviruses (NoVs) are a major cause of childhood gastroenteritis and foodborne diseases worldwide
Homology modelling [20] suggested that though the complete viral protein had slightly dispersed structure (Fig. 1b), the P domain from ORF2 encompassing amino acid 222–539 had globular structure that would tend to have a compact three dimensional (3D) organization (Fig. 1b). This observation was substantiated by the non-denaturing native polyacrylamide gel electrophoresis (PAGE) of the cloned protein that showed that most of P dimer (Pd) protein remained in the stacking gel (Fig. 1c)
FlaB enhances Pd‐specific antibody responses in a toll like receptor 5 (TLR5)‐dependent manner In order to estimate the immunogenicity of P dimers expressed in bacterial system, mice were immunized through i.n. or s.l. route using Pd antigen alone or in combination with FlaB (Pd + FlaB)

Summary

Introduction

Noroviruses (NoVs) are a major cause of childhood gastroenteritis and foodborne diseases worldwide. Subunit protein vaccines (e.g. Pd-vaccine) are poor inducers of humoral immune responses, in particular secretory antibody responses [14] In this context, the use of adjuvants potentiating immune responses in the mucosal immune compartments will significantly boost the vaccine efficacy [15]. In our previous studies we have shown that FlaB, a flagellin protein from Vibrio vulnificus and an agonist of TLR5, is a potent adjuvant for mucosally administered vaccines [18]. We evaluated the potential of FlaB coadministration along with the recombinant Pd antigen in enhancing the antigen-specific protective immune responses. We report that the intranasal immunization of mice with Pd+FlaB mixture vaccine induced a potent antibody and cell mediated immune response in both systemic and mucosal compartments. We evaluated the advantages of FlaB over known vaccine adjuvants such as alum and complete Freund’s adjuvant (CFA)

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Discussion

Conclusion