Gonorrhoea continues to seriously impact human society with an estimated 106 million new infections occurring annually. The consequence of gonorrhoea on reproductive and neonatal health is especially concerning as is its role in the spread of HIV. Current control measures rely on the identification and treatment of infected individuals and their sexual contacts. The success of this strategy, which is already inadequate, is lessened by poor diagnostic capabilities in many parts of the world and challenged by the rapid emergence of antibiotic-resistant strains. The potential of untreatable gonorrhoea is now real, and a gonorrhoea vaccine is seriously needed. Historically, gonorrhoea vaccine research has been hampered by the antigenic variability of the gonococcal surface, a lack of known protective mechanisms, and the absence of a small laboratory animal model for testing candidate vaccines and manipulating host responses. Here we discuss recent advances that have rekindled research efforts towards a gonorrhoea vaccine. Several conserved and semiconserved vaccine antigens have been identified that elicit bactericidal antibodies or inhibit target function. A mouse genital tract infection model is available for systematic testing of vaccines, and transgenic mice have been developed to relieve host restrictions. Additionally, several immunological advances have been made including the identification of mechanisms by which Neisseria gonorrhoeae suppresses the adaptive response and the demonstration that Th1 responses clear experimental infection in mice and induce a protective memory response. We also discuss important issues with respect to product development that must be considered when entering the vaccine pipeline.
Statistics from Altmetric.com
Gonorrhoea: old and new concerns, and the serious need for a vaccine
Gonorrhoea is an ancient disease that has seriously impacted reproductive and neonatal health since at least Biblical times. In the modern age, gonorrhoea is accompanied by the additional concerns of increased spread of HIV and the rapid emergence and global spread of antibiotic-resistant strains. Increased resistance to the extended-spectrum cephalosporins and treatment failures have led to the recent recommendation of dual therapy as a first-line treatment for gonorrhoea. Based on the evolutionary history of this organism, it is likely that this strategy will eventually be ineffective. This alarming situation prompted WHO to issue a global action report in 2012 that documents the disease burden of gonorrhoea in the world. Currently, 106 million cases of gonorrhoea are estimated to occur annually worldwide.1 The majority of infections are uncomplicated mucosal infections of the urogenital tract, pharynx and rectum. Ascended infections (eg, endometritis, salpingitis, and epididymitis) are more complicated, and gonococcal pelvic inflammatory disease (PID) and the related complications of infertility, ectopic pregnancy, and chronic pelvic pain constitute the major morbidity and mortality associated with gonorrhoea. Gonorrhoea during pregnancy increases the risk of ophthalmia neonatorum in neonates, an acute conjunctivitis that can lead to blindness. Pregnant women with gonorrhoea also have increased risk of premature rupture of membranes and preterm delivery, which can lead to low birthweight babies and chorioamnionitis complicated by septic abortion. Disseminated gonococcal infection (DGI) occurs in up to 0.5–3% of individuals with mucosal infections.2
Clearly, gonorrhoea continues to plague humankind, and new control strategies are seriously needed. Women and neonates suffer disproportionately from disease, and the incidence of gonorrhoea is highest in developing countries and in lower socioeconomic groups within industrialised countries. Current control measures rely on the identification and treatment of infected individuals and their sexual contacts. Unfortunately, the usefulness of this approach is threatened by antibiotic resistance and challenged by poor diagnostic capabilities in parts of the world where syndromic management is used in the absence of microbiologic culture or molecular diagnostic techniques.1 To curb the potential threat of untreatable gonorrhoea, public health agencies, professional societies and non-governmental organisations have issued a call for new antimicrobials against this pathogen. However, given the complexity of the antimicrobial development process, the time and resources needed to develop products, and the rapidity by which Neisseria gonorrhoeae develops resistance to newly introduced antibiotics, a rational case to pursue vaccine development can be made to truly have a long-term impact on public health. Here we discuss research progress and challenges in this area and describe some recent sophisticated advances in the field of vaccinology that may facilitate the development of gonorrhoea vaccines. We also discuss important issues inherent to product development and evaluation.
Early vaccine studies
Gonorrhoea vaccine research was an active area of investigation four decades ago. Optimism was high with the success of capsule-based meningococcal vaccines and the promising demonstration that experimental urethral infection in chimpanzees reduced susceptibility to reinfection. Two unsuccessful field trials were conducted during this time. A parenteral heat-killed whole-cell vaccine was tested in Inuit subjects in Canada, which yielded no protection. In another trial, parenteral intradermal delivery of a purified pilin vaccine failed to protect high-risk US military personnel in Korea, most likely due to pilin antigenic variation (reviewed in3).
With time, gonorrhoea vaccine research began to wane. Chimpanzees were no longer available for vaccine studies, and investigators were frustrated by the antigenic variability of the gonococal surface and the absence of known correlates of protection. Additionally, the lack of a small laboratory animal model made it difficult to test candidate antigens and study immune responses. Many investigators turned their attention towards unravelling the genetic basis of antigenic variation in the gonococcus. This era of discovery, which was impressive in its use of the nascent tools of molecular biology, resulted in N gonorrhoeae becoming a leading paradigm of a pathogen that uses phase and antigenic variation of surface molecules to adapt to its host. The advent of molecular biology also fuelled molecular pathogenesis research, a consequence of which is the identification of surface molecules that could be targeted by a vaccine to block or interfere with infection (table 1). Many of these antigens are stably expressed and conserved, and for some semiconserved antigens, conserved functional regions have been identified against which vaccine-induced immune responses might be directed.
Current antigens under development
Potential vaccine targets in N gonorrhoeae include surface molecules that mediate adherence to, or uptake by, host cells such as PilC, the type 4 pilus-associated adhesin4 and PilQ, the secretin through which pili are extruded.5 Another colonisation factor that could be targeted is the major gonococcal porin (PorB), which is involved in invasion of host cells6 ,7 While there is antigenic heterogeneity among surface-exposed PorB loops between strains, the identification of conserved domains that confer PorB-mediated functions could direct the successful development of broadly reactive PorB antigens. Once colonisation is established, the formation of biofilms may stabilise colonisation and protect against host innate defenses. Several gonococcal factors are required for efficient biofilm formation including nitrite reductase (AniA).8 AniA is required for anaerobic growth of N gonorrhoeae in the presence of nitrite and is unusual in its surface exposure. Ani-A-specific antibodies were detected in serum from patients with PID or DGI and anaerobic growth is likely to be important in various body sites inhabited by N gonorrhoeae, particularly the female upper genital tract.9 In support of AniA as a vaccine target, Jennings and colleagues recently reported that antibodies against recombinant AniA protein lacking the immunodominant glycosylated C-terminus inhibit nitrite reductase activity.10
Nutrient acquisition systems have also been identified that could be developed as ‘nutritional vaccines’. The gonococcal transferrin (Tf) receptor (TbpA and TbpB) or lactoferrin (Lf) receptor are required for experimental urethral infection of male volunteers.11 Intranasal immunisation of mice with TbpA and TbpB proteins fused to cholera toxin subunit B induces high titre, specific vaginal IgG and IgA antibodies and bactericidal serum antibodies that inhibit growth of N gonorrhoeae in media containing Tf as the sole source of iron (reviewed in 12). Fifty percent of N gonorrhoeae strains also express a Lf receptor and the LbpA subunit of the meningococcal Lf receptor, which is relatively conserved, induces bactericidal antibodies. These antibodies have limited cross-reactivity13; however, recent structural models of the neisserial Lf receptor could facilitate the design of vaccines against conserved functional domains.14
Vaccines that cripple the capacity of N gonorrhoeae to evade host innate defenses also may be effective. N gonorrhoeae expels hydrophobic antimicrobial substances that bathe mucosal surfaces (eg, fatty acids, long-chain faecal lipids, antimicrobial peptides, progesterone, bile salts) through the MtrC-MtrD-MtrE or FarA-FarB-MtrE active efflux pumps. The MtrC-MtrD-MtrE system is critical for murine infection and plays a role in antibiotic resistance.15 MtrE, the outermost component of the MtrC-MtrD-MtrE and FarA-FarB-MtrE pumps has two surface-exposed loops that could be targeted in a vaccine. Immunisation of mice with recombinant MtrE results in surface binding, bactericidal antibodies that recognise a variety of gonococcal strains (AJ DeRocco and AE Jerse, unpublished observation). Another surface-exposed factor that protects against host innate defenses is gonococcal α-2,3-sialyltransferase (Lst), which catalyses the addition of host-derived sialic acid to the lacto-neotetraose (LNT) species of lipooligosaccharide (LOS). Antibodies against purified Lst reduce the level of sialylation,16 and thus, could increase susceptibility to opsonic uptake by neutrophils and complement-mediated bacteriolysis.17 The importance of Lst during infection is supported by the demonstration that an Lst-deficient mutant is attenuated for murine genital tract infection.18
Other promising vaccine targets include the 2C7 LOS epitope. Rice and colleagues showed that antibodies against a 2C7-OS peptide mimic are highly bactericidal and promote opsonophagocytic killing of N gonorrhoeae. Recently, intraperitoneal immunisation of mice with a multiantigenic form of the 2C7-OS peptide mimic was reported to protect mice from subsequent challenge as did passive delivery of 2C7 monoclonal antibody.19 ,20
Another exciting advancement that may guide gonococcal vaccine research is the development of outer membrane vesicle (OMV) vaccines against Neisseria meningitidis group B. The 4CMenB vaccine consists of OMV with three added protein antigens identified by reverse vaccinology.21 None of the three proteins (fHBP, NHBA and NadA) in the 4CmenB vaccine are predicted to be suitable vaccine targets for gonorrhoea.22 However, similar non-biased proteomic or genomic screens to identify effective vaccine antigens could be applied to gonorrhoea vaccine research. Alternatively, gonococcal OMVs could be prepared from strains that express detoxified LOS, lack immunosuppressive or blocking antigens, and constitutively express transcriptionally regulated proteins that are known to be expressed during infection such as AniA and the Tf and Lf receptors.
Immunisation and challenge models
N gonorrhoeae is highly adapted to its human host, a characteristic that challenges animal modelling of gonorrhoea. Currently, there are two gonococcal genital tract infection models that can be used in immunisation and challenge studies. Experimental urethral infection of male volunteers is a well-characterised model that is highly relevant to natural infection.11 Modern technologies can now be applied to the human challenge model, which has been used for over 60 years, to obtain valuable information on host responses to vaccination or bacterial challenge at the protein or transcriptional level in urine or genital tract secretions. Limitations to the human model include its feasibility in most research settings, and the possibility that vaccine studies in male subjects may not accurately predict vaccine efficacy against cervical, ascending or disseminated infections.
Experimental infection of female mice can also be used to assess gonococcal vaccine efficacy.3 Female mice are transiently susceptible to N gonorrhoeae during the proestrus stage of the oestrous cycle; treatment with 17ß-estradiol and antibiotics prolongs colonisation of the lower genital tract for at least 10–12 days and ascending infection occurs in 17–20% of mice. BALB/c mice develop an inflammatory response to infection that is characterised by the influx of vaginal neutrophils. Infection persists during periods of inflammation, and similar to human infection, mice have a poor humoral response to infection, can be reinfected with the same strain, and show no evidence of memory response.18 Several host restrictions limit the capacity of experimental murine infection to mimic human neisserial infections. For example, the human carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) and the elusive pilus receptor are not expressed in mice. The gonococcus also cannot obtain iron from murine Tf or Lf. Of particular relevance to vaccine testing are host restrictions in soluble regulators of the complement cascade (factor H, C4b-binding protein), the absence of IgA1, the substrate of gonococcal IgA1 protease, and the opsonophagocytic receptor for IgA, FcαR (CD89). Transgenic mice can relieve some host restrictions and mouse strains that express human CEACAMs or Tf have been developed.18 Alternatively, purified factors, such as human fH, C4BP, or Tf could be administered to mice during the challenge phase of vaccine experiments to reproduce a more human-like state. This approach would be particularly useful for vaccines that are designed to block interactions with soluble host-restricted factors such as Tf receptor vaccines.
The immune response to gonorrhoea
Induction of vigorous innate, but not adaptive, responses
Symptomatic gonococcal infections are characterised by purulent exudates that contain neutrophils with intracellular diplococci. Inapparent infections are common, including most pharyngeal and rectal infections and 50–80% of female genital tract infections. Proinflammatory cytokines and chemokines are elevated in experimentally infected men with urethritis, but were not detected at levels higher than uninfected controls in naturally infected women unless coinfected with another sexually transmitted pathogen. The reason for the high rate of inapparent infections is not known; body site may play a role, and in women, hormonal influences may affect symptomology (reviewed in3). Recently, Russell and colleagues demonstrated that the Th17 pathway was responsible for the inflammatory response induced by N gonorrhoeae in the mouse model. Th17 responses have also been detected in humans with gonorrhoea, and are induced by N gonorrhoeae in human monocyte-derived dendritic cells along with IL-10. Induction of the Th17 pathway by N gonorrhoeae is consistent with other mostly extracellular pathogens that elicit a strong neutrophil response (reviewed in12).
By contrast with the innate response, the adaptive response to mucosal N gonorrhoeae infections is poor. Repeated infections are common and there is no definitive evidence of protective responses in humans. While partial serotype-specific immunity was detected in repeatedly infected women in Kenya, there was no evidence of serotype-specific immunity in less exposed subjects in a rural setting in the USA. The ratio of bactericidal antibodies to antibodies against the reduction-modifiable protein (Rmp), which blocks the bactericidal activity of PorB-specific or LOS-specific antibodies, has been proposed as a correlate of protection, and in one study, bactericidal antibodies were associated with reduced risk of salpingitis (reviewed in3).
Identification of immunosuppression mechanisms
The lack of knowledge of protective responses against gonorrhoea is a major challenge for gonorrhoea vaccine development. Appreciation of the immunosuppressive nature of gonorrhoea, further complicating vaccine development, has been relatively recent on the time-line of gonorrhoea research beginning with reports of Opa protein-mediated immunosuppression of T-cell proliferation and induction of B-cell death.23 The recognition of several other immunosuppressive mechanisms followed. These mechanisms include: (1) T-cell-independent polyclonal activation of IgD+CD27+ B cells to produce low-affinity IgM without inducing a memory response; (2) induction of pyronecrosis in macrophages, which is NLRP3 inflammasome-dependent and occurs by gonococcal-induced upregulation of cathepsin B; (3) inhibition of T-cell proliferation due to upregulation of IL-10 and programmed death ligand 1 (PDL-1) in dendritic cells; (4) suppression of Th1- and Th2-driven adaptive immune responses by mechanisms dependent on TGF-β and IL-10 as well as type 1 regulatory T cells (reviewed in12).
Continued characterisation of the immunosuppressive mechanisms used by N gonorrhoeae is critical for vaccine development. The bacterial factors responsible for triggering these pathways should be excluded from a vaccine and understanding these pathways may identify steps that can be subverted to induce a protective response. Recently, Russell and colleagues reported that antibody-mediated neutralisation of TGF-β and IL-10 in mice resulted in Th1-dependent and Th2-dependent responses and high titre serum and vaginal-specific antibodies. Moreover, mice also developed a memory response and were protected against reinfection. Similarly, localised administration of IL-12 incorporated in sustained-release microspheres induced Th1-dependent responses, clearance of infection and a protective memory response (reviewed in 12). These studies suggest that induction of a Th1 response may be the key to inducing protective immunity against gonorrhoea. This hypothesis was also proposed by Sparling and colleagues based on protection studies on mice immunised with viral replicon particles (VRP) expressing rPorB.3 While it has been assumed that B cells are most important for vaccine-mediated clearance of N gonorrhoeae, T helper cell responses drive antibody generation and the immunological memory response and may also be effective against intracellular gonococci.
Immunisation of the genital tract
Currently, the only licenced STI vaccines are against human papilloma virus and, therefore, there are few paradigms for successful vaccination of the genital tract. Understudied areas on gonorrhoea vaccine research include controlled comparisons of different immunisation routes used to induce mucosal responses and the testing of mucosal adjuvants known to enhance immune responses in the genital tract.24 ,25 Other important questions that are especially relevant to STI vaccines are the potential for gender-biased responses, and the influence of coinfection by other sexually transmitted pathogens on the efficacy of vaccines against a single pathogen. Little work has been done in this area.
Application of new vaccine technology
Advances in vaccinology have provided several new approaches that could be applied to gonorrhoea vaccines. Conventional approaches to vaccines such as killed/inactivated vaccines, subunit vaccines, conjugated vaccines, or live, attenuated vaccines, have strived to replicate the type of immunity elicited by natural infection while disassociating it from a pathogenic event. Newer approaches to rational vaccine design, which may include de novo constructs, are directed towards optimisation of immune responses and/or target delivery. Examples are vector-based vaccines, which can optimise antigen expression on a standardised platform that can incorporate multiple antigens. These platforms could form the basis of the delivery system, or could be used to increase antigen yield for subsequent purification. Advances in manufacturing and stabilisation include protein-coated microcrystal, nanoparticles, and glassification. New forms of product delivery could reduce the reliance on needle-based injection, such as a patch delivery system, microneedles, or orally delivered vaccines.
Entry into the vaccine pipeline
Each potential product must be evaluated through a product development pathway that includes preclinical development, clinical evaluation for safety and efficacy, and manufacturing scale-up to eventually reach licensure (figure 1). Many challenges are inherent in the development process including the potential timeline of a decade or more and the significant final resources necessary which can be in the billions of dollars. In the long run, the development of vaccines to prevent infections has been a significant public health advance and has proved to be cost saving or cost effective for many diseases. While it is unlikely that a gonorrhoea vaccine will be 100% effective, it is predicted that an efficacy level of 70% would reduce a considerable amount of disease and transmission worldwide. The most significant public health impact of a gonorrhoea vaccine would be on reproductive health. Therefore, a vaccine that protects against upper reproductive tract infections might be advanced through the product pipeline, but perhaps not a vaccine that protects only against urethritis. This rationale is based on a changing landscape, however, due to the emergence of multiple drug-resistant gonococcal strains. The potential rise of untreatable gonorrhoea could change the cost benefit analysis of a gonorrhoea vaccine as well as the public health message used to educate the public and providers when a vaccine is introduced. The case needs to be made for a vaccine to protect against gonococcal infection, because with the growing threat of antimicrobial resistance in N gonorrheae, the time to act may be now.
The impact of gonorrhoea on reproductive health and the threat of potentially untreatable gonorrhoea support the need for a gonorrhoea vaccine.
Progress in antigen discovery, gonococcal immunobiology, and animal modelling, together with the recent success of meningococcal outer membrane-based vaccines have reinvigorated gonorrhoea vaccine research.
Further investigation is needed in the areas of mucosal adjuvants, gender-biased responses, and the impact of coinfection on vaccine-induced protection.
Entry of gonorrhoea vaccines into the product pipeline will require a heavy investment of time and money, but the potential benefit should outweigh the risks.
We thank Dr Amanda DeRocco for helpful reading of the manuscript.
Handling editor Jackie A Cassell.
Contributors AEJ and CDD collaborated in the writing of this review. AEJ contributed sections about the history of gonorrhoea vaccine development and the current status and technical challenges of research in this area. CDD contributed the public health perspective on gonorrhoea vaccines, new advances in the field of vaccinology, and information pertaining to the product pipeline. Both authors edited the manuscript before submission.
Funding Funding for this work was provided to AEJ by grant number U19 AI31496 from the National Institute of Health.
Competing interests None.
Provenance and peer review Commissioned; externally peer reviewed.
If you wish to reuse any or all of this article please use the link below which will take you to the Copyright Clearance Center’s RightsLink service. You will be able to get a quick price and instant permission to reuse the content in many different ways.