Gonorrhea, an obligate human infection, is on the rise worldwide and gonococcal strains resistant to many antibiotics are emerging. Appropriate antimicrobial treatment and prevention, including ...effective vaccines, are urgently needed. To guide investigation, an experimental model of genital tract infection has been developed in female mice to study mechanisms by which
Neisseria gonorrhoeae
evades host-derived antimicrobial factors and to identify protective and immunosuppressive pathways. Refinements of the animal model have also improved its use as a surrogate host of human infection and accelerated the testing of novel therapeutic and prophylactic compounds against gonococcal infection. Reviewed herein are the (
a
) history of antibiotic usage and resistance against gonorrhea and the consequences of resistance mechanisms that may increase gonococcal fitness and therefore the potential for spread, (
b
) use of gonococcal infection in the animal model system to study mechanisms of pathogenesis and host defenses, and (
c
) current status of vaccine development.
The six-component maintenance of lipid asymmetry (Mla) system is responsible for retrograde transport of phospholipids, ensuring the barrier function of the Gram-negative cell envelope. Located ...within the outer membrane, MlaA (VacJ) acts as a channel to shuttle phospholipids from the outer leaflet. We identified Neisseria gonorrhoeae MlaA (ngo2121) during high-throughput proteomic mining for potential therapeutic targets against this medically important human pathogen. Our follow-up phenotypic microarrays revealed that lack of MlaA results in a complex sensitivity phenome. Herein we focused on MlaA function in cell envelope biogenesis and pathogenesis. We demonstrate the existence of two MlaA classes among 21 bacterial species, characterized by the presence or lack of a lipoprotein signal peptide. Purified truncated N. gonorrhoeae MlaA elicited antibodies that cross-reacted with a panel of different Neisseria. Little is known about MlaA expression; we provide the first evidence that MlaA levels increase in stationary phase and under anaerobiosis but decrease during iron starvation. Lack of MlaA resulted in higher cell counts during conditions mimicking different host niches; however, it also significantly decreased colony size. Antimicrobial peptides such as polymyxin B exacerbated the size difference while human defensin was detrimental to mutant viability. Consistent with the proposed role of MlaA in vesicle biogenesis, the ΔmlaA mutant released 1.7-fold more membrane vesicles. Comparative proteomics of cell envelopes and native membrane vesicles derived from ΔmlaA and wild type bacteria revealed enrichment of TadA-which recodes proteins through mRNA editing-as well as increased levels of adhesins and virulence factors. MlaA-deficient gonococci significantly outcompeted (up to 16-fold) wild-type bacteria in the murine lower genital tract, suggesting the growth advantage or increased expression of virulence factors afforded by inactivation of mlaA is advantageous in vivo. Based on these results, we propose N. gonorrhoeae restricts MlaA levels to modulate cell envelope homeostasis and fine-tune virulence.
•78 million new infections annually; greatest impact on women and neonates in LMIC.•Current control measures are inadequate and challenged by antibiotic resistance.•Conserved candidate vaccine ...antigens and adjuvant strategies are being developed.•There is a need for human studies to investigate correlates of immunity.•A meningococcal outer membrane vesicle vaccine may protect against gonorrhea.
There is a growing public health interest in controlling sexually transmitted infections (STIs) through vaccination due to increasing recognition of the global disease burden of STIs and the role of STIs in women’s reproductive health, adverse pregnancy outcomes, and the health and well-being of neonates. Neisseria gonorrhoeae has historically challenged vaccine development through the expression of phase and antigenically variable surface molecules and its capacity to cause repeated infections without inducing protective immunity. An estimated 78 million new N. gonorrhoeae infections occur annually and the greatest disease burden is carried by low- and middle-income countries (LMIC). Current control measures are clearly inadequate and threatened by the rapid emergence of antibiotic resistance. The gonococcus now holds the status of “super-bug” as there is currently no single reliable monotherapy for empirical treatment of gonorrhea. The problem of antibiotic resistance has elevated treatment costs and necessitated the establishment of large surveillance programs to track the spread of resistant strains. Here we review the need for a gonorrhea vaccine with respect to global disease burden and related socioeconomic and treatment costs, with an emphasis on the impact of gonorrhea on women and newborns. We also highlight the challenge of estimating the impact of a gonorrhea vaccine due to the need for more data on the burden of gonococcal pelvic inflammatory disease and related sequelae and of gonorrhea-associated adverse pregnancy outcomes and the problem of empirical diagnosis and treatment of STIs in LMIC. There is also a lack of clinical and basic science research in the area of gonococcal/chlamydia coinfection, which occurs in a high percentage of individuals with gonorrhea and should be considered when testing the efficacy of gonorrhea vaccines. Finally, we review recent research that suggests a gonorrhea vaccine is feasible and discuss challenges and research gaps in gonorrhea vaccine development.
There is a pressing need for drug development for gonorrhea. Here we describe a pharmacokinetic (PK)/pharmacodynamic (PD) analysis of extended-spectrum cephalosporins (ESC) against drug-susceptible ...and drug-resistant gonococcal strains in a murine genital tract infection model. The PK determined in uninfected mice displayed a clear dose-response in plasma levels following single doses of ceftriaxone (CRO) (intraperitoneal) or cefixime (CFM) (oral). The observed doses required for efficacy against ESC-susceptible (ESC
) strain FA1090 were 5 mg/kg of body weight (CRO) and 12 mg/kg (CFM); these doses had estimated therapeutic times (the time that the free drug concentration remains above the MIC
) of 24 h and 37 h, respectively. No single dose of CRO or CFM was effective against ESC-resistant (ESC
) strain H041. However, fractionation (three times a day every 8 h TIDq8h) of a 120-mg/kg dose of CRO resulted in estimated therapeutic times in the range of 23 h and cleared H041 infection in a majority (90%) of mice, comparable to the findings for gentamicin. In contrast, multiple CFM doses of 120 or 300 mg/kg administered TIDq8h cleared infection in ≤50% of mice, with the therapeutic times estimated from single-dose PK data being 13 and 27 h, respectively. This study reveals a clear relationship between plasma ESC levels and bacterial clearance rates in the gonorrhea mouse model. The PK/PD relationships observed in mice reflected those observed in humans, with
efficacy against an ESC
strain requiring doses that yielded an
in excess of 20 to 24 h. PK data also accurately predicted the failure of single doses of ESCs against an ESC
strain and were useful in designing effective dosing regimens.
There is a pressing need for a gonorrhea vaccine due to the high disease burden associated with gonococcal infections globally and the rapid evolution of antibiotic resistance in Neisseria ...gonorrhoeae (Ng). Current gonorrhea vaccine research is in the stages of antigen discovery and the identification of protective immune responses, and no vaccine has been tested in clinical trials in over 30 years. Recently, however, it was reported in a retrospective case-control study that vaccination of humans with a serogroup B Neisseria meningitidis (Nm) outer membrane vesicle (OMV) vaccine (MeNZB) was associated with reduced rates of gonorrhea. Here we directly tested the hypothesis that Nm OMVs induce cross-protection against gonorrhea in a well-characterized female mouse model of Ng genital tract infection. We found that immunization with the licensed Nm OMV-based vaccine 4CMenB (Bexsero) significantly accelerated clearance and reduced the Ng bacterial burden compared to administration of alum or PBS. Serum IgG and vaginal IgA and IgG that cross-reacted with Ng OMVs were induced by 4CMenB vaccination by either the subcutaneous or intraperitoneal routes. Antibodies from vaccinated mice recognized several Ng surface proteins, including PilQ, BamA, MtrE, NHBA (known to be recognized by humans), PorB, and Opa. Immune sera from both mice and humans recognized Ng PilQ and several proteins of similar apparent molecular weight, but MtrE was only recognized by mouse serum. Pooled sera from 4CMenB-immunized mice showed a 4-fold increase in serum bactericidal50 titers against the challenge strain; in contrast, no significant difference in bactericidal activity was detected when sera from 4CMenB-immunized and unimmunized subjects were compared. Our findings directly support epidemiological evidence that Nm OMVs confer cross-species protection against gonorrhea, and implicate several Ng surface antigens as potentially protective targets. Additionally, this study further defines the usefulness of murine infection model as a relevant experimental system for gonorrhea vaccine development.
The concept of immunizing against gonorrhea has received renewed interest because of the recent emergence of strains of
that are resistant to most currently available antibiotics, an occurrence that ...threatens to render gonorrhea untreatable. However, despite efforts over many decades, no vaccine has yet been successfully developed for human use, leading to pessimism over whether this goal was actually attainable. Several factors have contributed to this situation, including extensive variation of the expression and specificity of many of the gonococcal surface antigens, and the ability of
to resist destruction by complement and other innate immune defense mechanisms. The natural host restriction of
for humans, coupled with the absence of any definable state of immunity arising from an episode of gonorrhea, have also complicated efforts to study gonococcal pathogenesis and the host's immune responses. However, recent findings have elucidated how the gonococcus exploits and manipulates the host's immune system for its own benefit, utilizing human-specific receptors for attachment to and invasion of tissues, and subverting adaptive immune responses that might otherwise be capable of eliminating it. While no single experimental model is capable of providing all the answers, experiments utilizing human cells and tissues
, various
animal models, including genetically modified strains of mice, and both experimental and observational human clinical studies, have combined to yield important new insight into the immuno-pathogenesis of gonococcal infection. In turn, these have now led to novel approaches for the development of a gonococcal vaccine. Ongoing investigations utilizing all available tools are now poised to make the development of an effective human vaccine against gonorrhea an achievable goal within a foreseeable time-frame.
The essential gonococcal outer-membrane porin, PorB, manipulates the innate immune response via recruitment of negative regulators of the complement system, and by repressing the killing mechanisms ...of macrophages and neutrophils.Gonococcal PorB in its native folded conformation suppresses the capability of dendritic cells to stimulate proliferation of T cells.Although neisserial PorB is well described, there is conflicting information on the immunomodulatory properties of PorB from different Neisseria species.Studying PorB as protein aggregates or in its native folded state can generate conflicting results, an important consideration for future research.As PorB is the most abundant protein in gonococcal outer-membrane vesicles, so the immunomodulatory properties of this porin should be carefully considered in developing a successful gonorrhoea vaccine.
Neisseria gonorrhoeae is a human-specific pathogen responsible for the sexually transmitted infection, gonorrhoea. N. gonorrhoeae promotes its survival by manipulating both innate and adaptive immune responses. The most abundant gonococcal outer-membrane protein is PorB, an essential porin that facilitates ion exchange. Importantly, gonococcal PorB has several immunomodulatory properties. To subvert the innate immune response, PorB suppresses killing mechanisms of macrophages and neutrophils, and recruits negative regulators of complement to the gonococcal cell surface. For manipulation of adaptive immune responses, gonococcal PorB suppresses the capability of dendritic cells to stimulate proliferation of T cells. As gonococcal PorB is highly abundant in outer-membrane vesicles, consideration of the immunomodulatory properties of this porin is critical when designing gonococcal vaccines.
Neisseria gonorrhoeae is a human-specific pathogen responsible for the sexually transmitted infection, gonorrhoea. N. gonorrhoeae promotes its survival by manipulating both innate and adaptive immune responses. The most abundant gonococcal outer-membrane protein is PorB, an essential porin that facilitates ion exchange. Importantly, gonococcal PorB has several immunomodulatory properties. To subvert the innate immune response, PorB suppresses killing mechanisms of macrophages and neutrophils, and recruits negative regulators of complement to the gonococcal cell surface. For manipulation of adaptive immune responses, gonococcal PorB suppresses the capability of dendritic cells to stimulate proliferation of T cells. As gonococcal PorB is highly abundant in outer-membrane vesicles, consideration of the immunomodulatory properties of this porin is critical when designing gonococcal vaccines.
Lysozymes are nearly omnipresent as the first line of immune defense against microbes in animals. They exert bactericidal action through antimicrobial peptide activity and peptidoglycan hydrolysis. ...Gram-negative bacteria developed several weapons to battle lysozymes, including inhibitors of c-type lysozymes in the MliC/PliC family and the Neisseria adhesin complex protein (ACP). Until the recent discovery of ACP, no proteinaceous lysozyme inhibitors were reported for the genus Neisseria, including the important human pathogen N. gonorrhoeae. Here, we describe a previously unrecognized gonococcal virulence mechanism involving a protein encoded by the open reading frame ngo1063 that acts to counteract c-type Iysozyme and provides a competitive advantage in the murine model of gonorrhea. We named this protein SliC as a surface-exposed lysozyme inhibitor of c-type lysozyme. SliC displays low overall primary sequence similarity to the MliC/PliC inhibitors, but we demonstrate that it has a parallel inhibitory mechanism. Our studies provide the first evidence that bacterial proteinaceous lysozyme inhibitors protect against host lysozyme during infection based on lack of attenuation of the ΔsliC mutant in lysozyme knock-out mice, and that the conserved residues involved in lysozyme inhibition, S83 and K103, are functionally indispensable during infection in wild type mice. Recombinant SliC completely abrogated the lytic activity of human and chicken c-type lysozymes, showing a preference towards human lysozyme with an IC50 of 1.85 μM and calculated KD value of 9.2 ± 1.9 μM. In contrast, mutated SliC bearing S83A and K103A substitutions failed to protect fluorescein-labeled cell-wall from lysozyme-mediated hydrolysis. Further, we present data revealing that SliC is a surface-displayed lipoprotein released in membrane vesicles that is expressed throughout all phases of growth, in conditions relevant to different niches of the human host, and during experimental infection of the murine genital tract. SliC is also highly conserved and expressed by diverse gonococcal isolates as well as N. meningitidis, N. lactamica, and N. weaveri. This study is the first to highlight the importance of an anti-lysozyme strategy to escape the innate immune response during N. gonorrhoeae infection.
The mucosa is colonized with commensal Neisseria. Some of these niches are sites of infection for the STD pathogen Neisseria gonorrhoeae (Ngo). Given the antagonistic behavior of commensal bacteria ...toward their pathogenic relatives, we hypothesized that commensal Neisseria may negatively affect Ngo colonization. Here, we report that commensal species of Neisseria kill Ngo through a mechanism based on genetic competence and DNA methylation state. Specifically, commensal-triggered killing occurs when the pathogen takes up commensal DNA containing a methylation pattern that it does not recognize. Indeed, any DNA will kill Ngo if it can enter the cell, is differentially methylated, and has homology to the pathogen genome. Consistent with these findings, commensal Neisseria elongata accelerates Ngo clearance from the mouse in a DNA-uptake-dependent manner. Collectively, we propose that commensal Neisseria antagonizes Ngo infection through a DNA-mediated mechanism and that DNA is a potential microbicide against this highly drug-resistant pathogen.
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•Commensal Neisseria kill STD pathogen N. gonorrhoeae by releasing DNA into the environment•Killing requires DNA entry and recombination and a foreign DNA methylation pattern•Commensal N. elongata accelerates clearance of N. gonorrhoeae from the mouse vagina•A N. gonorrhoeae DNA uptake mutant resists this clearance
Kim et al. report that the STD pathogen Neisseria gonorrhoeae is killed when it takes up DNA released by related commensal bacteria. Neisseria gonorrhoeae employs the DNA uptake system to acquire antibiotic resistance genes and evade the immune response, thereby suggesting that this system may be the pathogen’s Achilles heel.
Declining gonococcal susceptibility to ceftriaxone and azithromycin has raised the possibility of untreatable gonorrhea in the future and reignited interest in gonococcal vaccine development. Despite ...decades of research, previous gonococcal vaccine candidates have been ineffective. A growing body of data suggests that meningococcal group B outer-membrane vaccines may be cross-protective against Neisseria gonorrhoeae. Clinical trials of a licensed vaccine against Neisseria meningitidis serogroup B containing an outer-membrane vaccine component are underway to determine its efficacy against N. gonorrhoeae. Other experimental gonococcal vaccine candidates are in the preclinical phases. Population impact of future gonococcal vaccines with different levels of efficacy and duration of protection in various populations is being evaluated using modeling studies. Despite recent progress, gaps in gonococcal vaccine research remain. Research is needed to evaluate vaccine efficacy in preventing gonococcal infections acquired via various anatomic routes and among patients coinfected with other sexually transmitted infections. Studies that model the impact of a future vaccine on high-burden populations such as men who have sex with men and estimate both vaccine cost-effectiveness and the incremental cost-effectiveness ratio of vaccination to antimicrobial resistance and treatment costs are warranted. This narrative review examines the current state of gonococcal vaccine research, the possible impact of a gonococcal vaccine on gonorrhea rates based on modeling studies, gaps in the gonococcal vaccine literature, and public health implications of a future gonococcal vaccine on reducing the gonorrhea burden in the United States.
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