Summary 

Neisseria gonorrhoeae, the causative agent of the sexually transmitted infection gonorrhoea, represents an emerging global health problem with increasing antibiotic resistance. Vaccine development is complicated by the fact that gonorrhoea infections fail to engender protective immunity. A successful vaccine against gonorrhoea should therefore include multiple antigens and induce a protective immune response beyond that generated through natural infections.

Identification of suitable vaccine targets remains a bottleneck and identification of conserved antigens through comparative genomics overlooks post-translational modifications. Here, we aim at providing proof-of-concept for protein glycans as important vaccine antigens against gonorrhoea. The glycoproteins identified in N. gonorrhoeae are mainly lipoproteins or transmembrane proteins localized in the periplasm or on the cell surface, and we have established that the attached glycans are antigenic and immunogenic.

We have interesting results that protein glycans are protecting the gonococci against bactericidal activity and indirect results that antibodies against these glycans might be bactericidal. We will confirm and validate these results in different strains and investigate the molecular mechanisms behind these observations by using a panel of mutant strains that express different glycans or glycoproteins. Recent studies have shown decreased rates of gonorrhea after vaccination with meningococcal outer membrane vesicle (OMV) vaccines, and we will identify the glycoproteins and attached glycans within these OMV vaccines.

Project outcomes

Neisseria gonorrhoeae represents a global health problem with significant morbidity and mortality and increasing number of antimicrobial resistant isolates. Surveillance remains limited in several regions, and here we present the first WGS study from Ethiopia with seventy N. gonorrhoeae isolates. Endocervical and urethral swabs were collected among symptomatic patients at STI clinics of health facilities in Bahir Dar, Ethiopia. Among the isolates, multi-locus sequence typing ST-1587 was dominant (54%) and the most common NG-STAR sequence types were ST-5621 (18.6%), ST-1203 (14.3%) and ST-5666 (10%). All isolates were resistant to Tetracycline, Ciprofloxacin, and most isolates (95.7%) were resistance to Penicillin-G, while all isolates were susceptible for Cefixime, Ceftriaxone, Azithromycin and Spectinomycin. The genome-based AMR prediction was matched with phenotypic observations for all six antimicrobials and the genotypic AMR determinants was identified. This study contributes to increased knowledge on antimicrobial susceptibility pattern and mechanisms in Ethiopia.

Vaccine development has been difficult, and even repeated gonorrhoea infections fail to engender protective immunity. An effective vaccine should therefore include several antigens to induce a strong and protective immune response.

Here, we have investigated the potential for protein glycans as vaccine antigens. Species of Neisseria encompass broad-spectrum O-linked protein glycosylation where the glycoproteins are mainly transmembrane proteins or lipoproteins localized on the cell surface or in the periplasm^1-4,6. Several of the N. gonorrhoeae glycoproteins elicit bactericidal or functional blocking antibodies and have previously been suggested as vaccine candidates: PilE⁷, Mip^8,9, AniA¹⁰,  PilQ^11,12, MetQ^13,14, PorB¹⁵, ZnuD^16,17 and PotF3¹⁸. We suggest that since glycans impact on the immunogenicity, as shown for the PilE protein¹, glycan function and diversity ought to be considered when including glycoproteins in future vaccines.

Multiple studies have reported decreased rates of gonorrhoea after vaccination with meningococcal serogroup B (MenB) outer membrane vesicles (OMV) – based vaccines^19-21. However, we found that meningococcal OMV vaccination did not significantly induce bactericidal activity against N. gonorrhoeae. Meningococcal OMV vaccines contain various outer membrane proteins that are known to be glycosylated, and we are identifying these glycoproteins and glycans. In addition, we examined the potential impact of bacterial protein glycosylation on susceptibility to complement-mediated killing of bacteria (serum bactericidal activity) elicited by human antibodies. We observed significantly higher bactericidal titres against glycosylation null mutant N. meningitidis versus the wild type strains conceivably through masking protein epitopes important for bactericidal killing. This effect was less evident for the N. gonorrhoeae strains studied.

An effective gonorrhoea vaccine will control this epidemic and reduce the need for antibiotic treatment and it is therefore important to explore new vaccine antigens. We believe that protein glycans fulfil key criteria for an optimal vaccine antigen in that they are surface-exposed, immunogenic, and possess limited antigenic variability in N. gonorrhoeae. However, further studies are necessary to understand the complex molecular mechanisms involved, as well as the involved regulatory mechanisms.