Neisseria Meningitidis Outer Membrane Vesicle Research Articles

Immunological Effects of Dimethyldioctadecylammonium Bromide and Saponin as Adjuvants for Outer Membrane Vesicles from Neisseria meningitidis.

The meningococcal disease is a global health threat, but is preventable through vaccination. Adjuvants improve meningococcal vaccines and are able to trigger different aspects of the immune response. The present work evaluated the immune response of mice against Neisseria meningitidis outer membrane vesicles (OMV) complexed with the adjuvants aluminium hydroxide (AH), via subcutaneous route; and dimethyldioctadecylammonium bromide (DDA) or Saponin (Sap), via intranasal/subcutaneous routes. ELISA demonstrated that all adjuvants increased IgG titers after the booster dose, remaining elevated for 18 months. Additionally, adjuvants increased the avidity of the antibodies and the bactericidal titer: OMVs alone were bactericidal until 1:4 dilution but, when adjuvanted by Alum, DDA or Sap, it increased to 1/32. DDA and Sap increased all IgG isotypes, while AH improved IgG1 and IgG2a levels. Thus, Sap led to the recognition of more proteins in Immunoblot, followed by DDA and AH. Sap and AH induced higher IL-4 and IL-17 release, respectively. The use of adjuvants improved both cellular and humoral immune response, however, each adjuvant contributed to particular parameters. This demonstrates the importance of studying different adjuvant options and their suitability to stimulate different immune mechanisms, modulating the immune response.

A COVID-19 vaccine candidate composed of the SARS-CoV-2 RBD dimer and Neisseria meningitidis outer membrane vesicles.

SARS-CoV-2 infection is mediated by the interaction of the spike glycoprotein trimer via its receptor-binding domain (RBD) with the host's cellular receptor. Vaccines seek to block this interaction by eliciting neutralizing antibodies, most of which are directed toward the RBD. Many protein subunit vaccines require powerful adjuvants to generate a potent antibody response. Here, we report on the use of a SARS-CoV-2 dimeric recombinant RBD combined with Neisseria meningitidis outer membrane vesicles (OMVs), adsorbed on alum, as a promising COVID-19 vaccine candidate. This formulation induces a potent and neutralizing immune response in laboratory animals, which is higher than that of the dimeric RBD alone adsorbed on alum. Sera of people vaccinated with this vaccine candidate, named Soberana01, show a high inhibition level of the RBD-ACE2 interaction using RBD mutants corresponding to SARS-CoV-2 variants of concern and wild-type expressed using the phage display technology. To our knowledge, this is the first time that the immunostimulation effect of N. meningitidis OMVs is evaluated in vaccine candidates against SARS-CoV-2.

Spontaneously released Neisseria meningitidis outer membrane vesicles as vaccine platform: production and purification

Outer membrane vesicles (OMVs) are nanoparticles produced by Gram-negative bacteria that can be used as vaccines. The application of OMVs as vaccine component can be expanded by expressing heterologous antigens on OMVs, creating an OMV-based antigen presenting platform. This study aims to develop a production process for such OMV-based vaccines and studies a production method based on meningococcal OMVs that express heterologous antigens on their surface. As a proof of concept, the Borrelia burgdorferi antigens OspA and OspC were expressed on Neisseria meningitidis OMVs to create a concept anti-Lyme disease vaccine. Production of OMVs released in the culture supernatant was induced by high dissolved oxygen concentrations and purification was based on scalable unit operations. A crude recovery of 90 mg OMV protein could be obtained per liter culture. Expressing heterologous antigens on the OMVs did result in minor reduction of bacterial growth, while OMV production remained constant. The antigen expression did not alter the OMV characteristics. This study shows that production of well characterized OMVs containing heterologous antigens is possible with high yields by combining high oxygen concentrations with an optimized purification process. It is concluded that heterologous OMVs show potential as a vaccine platform.

Advancing vaccine development for gonorrhoea and the Global STI Vaccine Roadmap.

Efforts to develop vaccines against Neisseria gonorrhoeae have become increasingly important, given the rising threat of gonococcal antimicrobial resistance (AMR). Recent data suggest vaccines for gonorrhoea are biologically feasible; in particular, epidemiological evidence shows that vaccines against a closely related pathogen, serogroup B Neisseria meningitidis outer membrane vesicle (OMV) vaccines, may reduce gonorrhoea incidence. Vaccine candidates using several approaches are currently in preclinical development, including meningococcal and gonococcal OMV vaccines, a lipooligosaccharide epitope and purified protein subunit vaccines. The Global STI Vaccine Roadmap provides action steps to build on this technical momentum and advance gonococcal vaccine development. Better quantifying the magnitude of gonorrhoea-associated disease burden, for outcomes like infertility, and modelling the predicted role of gonococcal vaccines in addressing AMR will be essential for building a full public health value proposition, which can justify investment and help with decision making about future vaccine policy and programs. Efforts are underway to gain consensus on gonorrhoea vaccine target populations, implementation strategies and other preferred product characteristics that would make these vaccines suitable for use in low- and middle-income, as well as high-income, contexts. Addressing these epidemiological, programmatic and policy considerations in parallel to advancing research and development, including direct assessment of the ability of meningococcal B OMV vaccines to prevent gonorrhoea, can help bring about the development of viable gonococcal vaccines.

Outer Membrane Vesicles from Neisseria Meningitidis (Proteossome) Used for Nanostructured Zika Virus Vaccine Production

The increase of Zika virus (ZIKV) infections in Brazil in the last two years leaves a prophylactic measures on alert for this new and emerging pathogen. Concerning of our positive experience, we developed a new prototype using Neisseria meningitidis outer membrane vesicles (OMV) on ZIKV cell growth in a fusion of OMV in the envelope of virus particles. The fusion of nanoparticles resulting from outer membrane vesicles of N. meningitidis with infected C6/36 cells line were analyzed by Nano tracking analysis (NTA), zeta potential, differential light scattering (DLS), scan and scanning transmission eletronic microscopy (SEM and STEM) and high resolution mass spectometry (HRMS) for nanostructure characterization. Also, the vaccination effects were viewed by immune response in mice protocols immunization (ELISA and inflammatory chemokines) confirmed by Zika virus soroneutralization test. The results of immunizations in mice showed that antibody production had a titer greater than 1:160 as compared to unvaccinated mice. The immune response of the adjuvant and non-adjuvant formulation activated the cellular immune response TH1 and TH2. In addition, the serum neutralization was able to prevent infection of virus particles in the glial tumor cell model (M059J). This research shows efficient strategies without recombinant technology or DNA vaccines.

Impact of Reducing Complement Inhibitor Binding on the Immunogenicity of Native Neisseria meningitidis Outer Membrane Vesicles.

Neisseria meningitidis recruits host human complement inhibitors to its surface to down-regulate complement activation and enhance survival in blood. We have investigated whether such complement inhibitor binding occurs after vaccination with native outer membrane vesicles (nOMVs), and limits immunogenicity of such vaccines. To this end, nOMVs reactogenic lipopolysaccharide was detoxified by deletion of the lpxl1 gene (nOMVlpxl1). nOMVs unable to bind human complement factor H (hfH) were generated by additional deletions of the genes encoding factor H binding protein (fHbp) and neisserial surface protein A (NspA) (nOMVdis). Antibody responses elicited in mice with nOMVdis were compared to those elicited with nOMVlpxl1 in the presence of hfH. Results demonstrate that the administration of human fH to mice immunized with fHbp containing OMVlpxl1 decreased immunogenicity against fHbp (but not against the OMV as a whole). The majority of the OMV-induced bactericidal immune response (OMVlpxl1 or OMVdis) was versus PorA. Despite a considerable reduction of hfH binding to nOMVdis, and the absence of the vaccine antigen fHbp, immunogenicity in mice was not different from nOMVlpxl1, in the absence or presence of hfH (serum bactericidal titers of 1:64 vs 1:128 after one dose in the nOMVdis and nOMVlpxl1–immunized groups respectively). Therefore, partial inhibition of fH binding did not enhance immunity in this model.

Immuno-proteomic analysis of human immune responses to experimental Neisseria meningitidis outer membrane vesicle vaccines identifies potential cross-reactive antigens

Human volunteers were vaccinated with experimental Neisseria meningitidis serogroup B vaccines based on strain H44/76 detoxified L3 lipooligosaccharide (LOS)-derived outer membrane vesicles (OMV) or the licensed Cuban vaccine, VA-MENGOC-BC. Some volunteers were able to elicit cross-bactericidal antibodies against heterologous L2-LOS strain (760676). An immuno-proteomic approach was used to identify potential targets of these cross-bactericidal antibodies using an L2-LOS derived OMV preparation. A total of nine immuno-reactive spots were detected in this proteome: individuals vaccinated with the detoxified OMVs showed an increase in post-vaccination serum reactivity with Spots 2–8, but not with Spots 1 and 9. Vaccination with VA-MENGOC-BC induced sera that showed increased reactivity with all of the protein spots. Vaccinees showed increases in serum bactericidal activity (SBA) against the heterologous L2-LOS expressing strain 760676, which correlated, in general, with immunoblot reactivity. The identities of proteins within the immuno-reactive spots were determined. These included not only well-studied antigens such as Rmp, Opa, PorB and FbpA (NMB0634), but also identified novel antigens such as exopolyphosphatase (NMB1467) and γ-glutamyltranspeptidase (NMB1057) enzymes and a putative cell binding factor (NMB0345) protein. Investigating the biological properties of such novel antigens may provide candidates for the development of second generation meningococcal vaccines.

Heterologous prime-boost strategy to overcome weak immunogenicity of two serosubtypes in hexavalent Neisseria meningitidis outer membrane vesicle vaccine

In the hexavalent meningococcal B OMV vaccine (HexaMen), two of the six Porin A proteins present are weakly immunogenic in mice and humans. We investigated the possibility that the lower immunogenicity of these serosubtypes (P1.7-2,4 and P1.19,15-1) could be overcome by using HexaMen and monovalent OMVs in heterologous immunisation protocols. Whereas HexaMen priming on day 0 followed by a monovalent P1.7-2,4 OMV boosting on day 28 (specific boost) did not result in higher titres against P1.7-2,4 (on day 42), the reverse order of immunisations (specific priming) resulted in significantly higher ELISA and SBA titres, but with lower avidity. For the strongly immunogenic PorA P1.5-1,2-2, all strategies gave high antibody responses, while avidity was highest after two monovalent P1.5-1,2-2 OMV immunisations. Based on the improved antibody titres obtained by specific priming with the weakly immunogenic PorA, we extended our study with combined P1.7-2,4 and P1.19,15-1 priming followed by two HexaMen booster immunisations. This resulted in higher ELISA and SBA titres against these weakly immunogenic PorAs, while the response against the other four PorAs was unaffected. Also, we observed an increase in antibody avidity using this schedule, indicating that affinity maturation has occurred. In conclusion, we found that specific priming, rather than specific boosting with monovalent OMVs, gave a significant rise in the serosubtype-specific immune response against a weakly immunogenic PorA, with high avidity antibodies in an extended immunisation schedule.

Immunization with Neisseriameningitidis outer membrane vesicles prevents bacteremia in neonatal mice

Although vaccines based on outer membrane vesicles (OMV) of Neisseria meningitidis have been developed and administered to children, little is known about the magnitude and quality of the immune response in animal models of early life immunization. We investigated the immunogenicity of meningococcal OMV, and the influence of route and immunization schedule, in neonatal mice. The administration of two intraperitoneal doses of OMV, given at 7 and 14 days after birth, induced a significant antibody response and was highly effective in conferring protection against bacteremia in 21-day-old mice challenged with meningococci. Intranasal immunization was less effective and did not generate a protective immune response. The antibodies elicited by intraperitoneal immunization were cross-reactive with several meningococcal strains and a memory response was demonstrated when mice immunized as neonates were given a booster immunization at 6 weeks of age.

Direct isolation of recombinant human antibodies against group B Neisseria meningitidis from scFv expression libraries

The successful generation of human antibodies from large naı̈ve antibody libraries requires iterative selection steps. Here, we describe a new and fast method for the isolation of high affinity antibodies directly from human single chain Fv antibody (scFv) expression libraries. Escherichia coli scFv expression libraries were made from peripheral blood lymphocytes from four individuals vaccinated with group B Neisseria meningitidis outer membrane vesicle (OMV) vaccine. Forty thousand clones were directly screened for antibodies binding N. meningitidis strain 44/76 (B:15:P1.7,16). Of 430 specific clones detected, 225 candidates were isolated and re-screened against the N. meningitidis strains NZ-98/254 (B:4:P1.7b,4) giving 4% cross-reactive clones. Antibodies were further characterized by DNA sequencing, ELISA and surface plasmon resonance (SPR) analysis, showing broad V-gene diversity and nanomolar scFv affinities. Antibodies derived by this method may assist in the discovery and development of new vaccine antigens as well as therapeutic antibody agents for the treatment of meningococcal diseases.

Immunization with meningococcal outer-membrane protein vesicles containing lipooligosaccharide protects mice against lethal experimental group B Neisseria meningitidis infection and septic shock.

Detergent-treated group B Neisseria meningitidis outer membrane vesicles (D-OMVs) from wild-type M986 and from nonencapsulated mutant M986-non-capsule variant (NCV) were compared as immunogens. Eight weeks after 3 consecutive immunizations with the immunogens, mice were challenged with a lethal dose of purified endotoxin or heat-killed or living N. meningitidis, plus d-galactosamine (400 mg/kg). D-OMVs from M986 induced bactericidal antibodies to both M986 (B : 2a : P1.5,2 : L3,7) and 6275 (B : 2a : P1.2,5 : L3) and protected the animals against both strains, whereas D-OMVs from M986-NCV did not protect the animals against infection with 6275 even when high serum bactericidal activity was induced. Tumor necrosis factor-alpha detected after bacterial infection was high in both protected and unprotected mice; interleukin (IL)-6 was high in mice that died but low in animals that survived. Exogenous administration of recombinant mouse IL-6 reversed the immunogens' protective effects. Protection against infection in mice does not necessarily correlate with the measured levels of serum bactericidal antibody alone, opsonic antibody alone, or cytokine profile alone. A comprehensive assessment of the preclinical efficacy of group B outer-membrane protein vaccines should include monitoring humoral antibodies, cytokine response, and protective effects against lethal infection.

Human T-cell responses after vaccination with the Norwegian group B meningococcal outer membrane vesicle vaccine.

We have analyzed human T-cell responses in parallel with serum immunoglobulin G (IgG) antibody levels after systemic vaccination with the Norwegian group B Neisseria meningitidis outer membrane vesicle (OMV) vaccine. Ten adult volunteers, with no or very low levels of serum IgG antibodies against meningococci, received three doses intramuscularly of the OMV vaccine (at weeks 0, 6, and 46). T-cell proliferation against the OMV vaccine, purified outer membrane proteins (PorA and PorB), and control antigens (Mycobacterium bovis BCG vaccine and tetanus toxoid) was measured by thymidine incorporation of peripheral blood mononuclear cells before and after vaccination. The results showed that vaccination with OMV elicits strong primary and booster T-cell responses specific to OMV as well as the PorA (class 1) protein and significant, but markedly lower, responses against the PorB (class 3) protein. The median responses to OMV and PorA were 26 and 16 times the prevaccination levels, respectively. Most of the vaccinees showed low T-cell responses against OMV and PorA before vaccination, and the maximum T-cell responses to all vaccine antigens were usually obtained after the second vaccine dose. We found a positive correlation between T-cell responses and anti-OMV IgG antibody levels (r = 0.50, P < 0.0001, for OMV and PorA). In addition, we observed a progressive increase in the percentage of CD45R0+ (memory) CD4-positive T cells (P = 0.002). In conclusion, we have shown that the Norwegian OMV vaccine against meningococcal B disease induced antigen-specific T-cell responses, kinetically accompanied by serum IgG responses, and that vaccination increased the proportion of memory T-helper cells.

Combination of DTP and Haemophilus influenzae Type b Conjugate Vaccines can Affect Laboratory Evaluation of Potency and Immunogenicity

Abstract. A commonly used Diphtheria-Tetanus-Pertussis (whole-cell) vaccine was combined with each of three different Haemophilus influenzae type b (Hib) capsular polysaccharide vaccines. Each Hib vaccine incorporated one of three different protein conjugates: tetanus toxoid, diphtheria CRM 197 toxoid or group B Neisseria meningitidis outer membrane vesicles. The effects of these combinations on the subsequent laboratory control testing were examined. The addition of the Hib vaccines had no significant effect on the reactogenicity or the potency of the whole-cell pertussis component. The potency of, and antibody responses to, the diphtheria component were also unaffected in all three combinations. However, combination with the Hib vaccine comprising polysaccharide conjugated to tetanus toxoid had dramatic effects on tetanus potency and immunogenicity when assayed in mice. This combination resulted in a five-fold potentiation of the tetanus potency and a similarly large increase in the antibody responses to tetanus toxin and toxoid. The level of the antibody response to the Hib polysaccharide in this vaccine was also elevated, more than 20-fold, as a result of the combination. Such phenomena were not evident with combinations involving the other two Hib vaccines. These results have implications for the control testing of combined vaccines containing a whole-cell pertussis component and Hib polysaccharide-tetanus protein conjugate vaccine.