Mouse vaginal lavage IgG against F62 induced by the combined antigens and Alum were higher than when using Alum+MPLA (Figure 4C, black bars), while similar vaginal lavage IgG were detected against MS11, regardless of the adjuvant used (Figure 4D, black bars)
Mouse vaginal lavage IgG against F62 induced by the combined antigens and Alum were higher than when using Alum+MPLA (Figure 4C, black bars), while similar vaginal lavage IgG were detected against MS11, regardless of the adjuvant used (Figure 4D, black bars). Recurring gonococcal exposure may result in some strain-specific immunity, but protective memory responses are scarce [7,8]. Disseminated gonococcal infections (DGI) and co-infections with in humans [34]. SBA is regarded as an in vitro surrogate of protection to guide the evaluation of vaccine candidates efficacy in vitro [35,36]. Other mechanisms of protection may include antibody-dependent opsonophagocytosis, the blocking of bacteria adhesion/invasion at the site of colonization, and T cell responses [37], although these are not all confirmed in human studies or unequivocally in experimental mouse models of gonococcal vaginal colonization. Interest in the use of outer membrane vesicles (OMVs) as a multi-antigen vaccine has been revitalized by the recent observation of the decreased risk of gonococcal infection in individuals vaccinated with meningococcal OMV-based vaccines. The MeNZB vaccine was reported to (+)-DHMEQ have 31% efficacy against gonococcal infection in a retrospective epidemiologic study of people immunized with the MenZB vaccine BTD [38,39]. VA-MENGOC-BC- and 4CMenB (Bexsero)-vaccinated cohorts (+)-DHMEQ were also reported to incur in lower rates of gonorrhea [40,41,42,43,44]. Currently, there are multiple randomized clinical trials underway to evaluate the protective efficacy of existing OMV vaccines against gonococcal infection [45]. In mice, 4CMenB accelerated the clearance (+)-DHMEQ of gonococcal infection and induced antibodies with bactericidal activity [46,47,48]. Cross-protection is attributed to shared meningococcal and gonococcal antigens [49,50,51]. The administration of OMVs with IL-12 as an adjuvant accelerated bacterial clearance in a mouse model of gonococcal colonization, likely due to the stimulation of protective Th1 responses and the concomitant reduction in deleterious Th17 responses [52]. Protection in mice afforded by this vaccine has also relied on antibodies, evidenced by a lack of efficacy in B cell-deficient mice [53]. Using a collection of antigens, such as in an OMV, may result in a more diversified functional response against multiple epitopes compared to mono-antigen vaccines [54,55,56]. Our previous studies using the gonococcal transcriptome expressed during natural human mucosal infection in men and women highlighted the following: (1) gene expression varies in the male and female reproductive tract environments, (2) gonococcal gene expression and regulation are different in vivo and in vitro, and (3) a large number of gonococcal genes expressed during human infection encode (+)-DHMEQ hypothetical proteins. Focusing on the latter, we designed a novel bioinformatics-based Candidate Antigen Selection Strategy (CASS) and identified several new potential vaccine antigens that are expressed in vivo during natural human gonococcal infection [24] (in contrast to antigens expressed in bacteria grown in vitro). Our initial studies in mice with three CASS antigens, NGO0690, NGO0948 and NGO1701, showed the robust induction of antibodies with serum bactericidal activity (SBA) against several strains in mice immunized with Alum as the adjuvant. SBA titers were increased by combining individual mouse sera, showing that the presence of bactericidal antibodies against more than one antigen enhanced the killing of [24]. Here, we expand the characterization of these antigens by verifying recognition by human serum antibodies from = 5) were immunized subcutaneously three times following a three-week schedule with recombinant NGO0690, NGO0948 and NGO1701 combined (10 g each), adsorbed with Alum (Imject, 40 mg/mL aluminum hydroxide, 40 mg/mL magnesium hydroxide) (Thermo Fisher Scientific, Waltham, MA, USA, #77161) at a 1:1 ratio, as specified by the manufacturer, in a final volume of 100 L of antigen/adjuvant mixture. For adsorption, Alum was added dropwise to the antigens and mixed for 30 min at room temperature (R.T.) prior to use, as specified by the manufacturer. Additional mice (= 10) were immunized with antigens adsorbed with Alum as above and with MPLA (10 g/mouse/dose) (Avanti Lipids, Alabaster, AL, USA) as an adjuvant. Control groups were immunized with adjuvants alone in PBS. Before the first immunization, preimmune sera were collected, and immune sera two weeks after each immunization (weeks 2,.