Intranasal administration of recombinant Neisseria gonorrhoeae transferrin binding proteins A and B conjugated to the cholera toxin B subunit induces systemic and vaginal antibodies in mice

Abstract

The transferrin binding proteins (TbpA and TbpB) comprise the gonococcal transferrin receptor and are considered potential antigens for inclusion in a vaccine against Neisseria gonorrhoeae. Intranasal (IN) immunization has shown promise in development of immunity against sexually transmitted disease pathogens, in part due to the induction of antigen-specific genital tract immunoglobulin A (IgA) and IgG. Conjugation of antigens to the highly immunogenic cholera toxin B subunit (Ctb) enhances antibody responses in the serum and mucosal secretions following IN vaccination. In the current study, we characterized the anti-Tbp immune responses following immunization of mice IN with recombinant transferrin binding proteins (rTbpA and rTbpB) conjugated to rCtb. We found that both rTbpA-Ctb and rTbpB-Ctb conjugates administered IN induced antibody responses in the serum and genital tract. IN immunization resulted in both IgA and IgG in the genital tract; however, subcutaneous immunization mainly generated IgG. Surprisingly, rTbpA alone was immunogenic and induced serum and mucosal antibody responses similar to those elicited against the rTbpA-Ctb conjugate. Overall, rTbpB was much more immunogenic than rTbpA, generating serum IgG levels that were greater than those elicited against rTbpA. Bactericidal assays conducted with sera collected from mice immunized IN with TbpA and/or TbpB indicated that both antigens generated antibodies with bactericidal activity. Anti-TbpA antibodies were cross-bactericidal against heterologous gonococcal strains, whereas TbpB-specific antibodies were less cross-reactive. By contrast, antibodies elicited via subcutaneous immunization were not cross-bactericidal against heterologous strains, indicating that IN vaccination could be the preferred route for elicitation of biologically functional antibodies.

FIG. 1.

Serum IgG levels specific for TbpA, TbpB, and Ctb. (A) Serum IgG levels specific for TbpA detected at days 17, 28, 35, and 65. (B) Serum IgG levels specific for TbpB detected at the same time points. (C) Serum IgG levels specific for Ctb detected at the same time points. Results are expressed as the geometric mean of antibody titers ×/÷ standard deviation. For all immunization groups, n = 5.

FIG. 2.

Serum IgA levels specific for TbpB and Ctb. (A) Serum IgA levels specific for TbpA detected at days 17, 28, 35, and 65. (B) Serum IgA levels specific for TbpB detected at the same time points. (C) Serum IgA levels specific for Ctb detected at the same time points. Results are expressed as the geometric mean of antibody titers ×/÷ standard deviation. For all immunization groups, n = 5.

FIG. 3.

IgG1 and IgG2a subtype analysis. (A) IgG1 and IgG2a antibody levels specific for TbpA detected in sera collected at day 35. (B) IgG1 and IgG2a antibody levels specific for TbpB detected in sera collected at day 35. The bars represent the geometric mean ×/÷ standard deviation. The values above the bars represent the IgG1/IgG2a ratios of the corresponding immunization groups, indicated below each graph. For all groups, n = 5.