Role of lipooligosaccharide in Neisseria gonorrhoeae immune response

In 2017, Neisseria gonorrhoeae (Ng) was listed by the World Health Organization (WHO) as a high-priority pathogen for which new therapeutics and prophylactic tools are urgently needed. In this panorama, renewed interest has been directed towards the development of GMMA- or OMV-based vaccines. GMMA (Generalized Module for Membrane Antigens) are outer membrane vesicles (OMV) shed from a strain genetically designed with different aims; among them, removing or reducing the expression of undesired antigens as in the case of lipopolysaccharide or lipooligosaccharide (LPS/LOS) removal to decrease the GMMA reactogenicity. Moreover, GMMA can be exploited for the expression new antigens and/or to enhance the expression of desired ones, as well as to increase the release of vesicles, thus improving the production yield. Lipooligosaccharide (LOS) is the most abundant antigen on the gonococcal membrane and plays a crucial role in pathogenesis being involved in bacterial adhesion to human cells and serum resistance. Moreover, antibody responses to LOS can mediate complement activation and bactericidal and opsonic activity. Despite these promising properties, the heterogeneity of LOS structures remains a challenge. Indeed, the glycan extensions of LOS oligosaccharide chains are determined by the expression of glycosyltransferases, some of which encoded by phase variable genes (lgtA, lgtC, lgtD, and lgtG). Therefore, the impact of different LOS epitopes expressed on a GMMA-based vaccine on functional immunogenicity requires further examination. In this study, GMMA derived from a mutant strain engineered to express a highly truncated LOS have been tested in vivo, showed low bactericidal activity for the majority of tested strains, suggesting a role for anti-LOS antibodies in the hSBA functional response. Indeed, the corresponding GMMA from the parental isogenic strain having a functional lgtF gene, are able to elicit a functional response, thus confirming LOS truncation is abolishing the response. In addition, a library of gonococcal isogenic mutant strains suitably engineered to express distinct LOS structures was used to dissect the contribution of the different LOS epitopes on GMMA immunogenicity. In vivo mice studies highlighted a correlation of different LOS structure and GMMA-mediated cross-functional activity. Moreover, to investigate the role of LOS in the absence of other antigens, liposomes formulated ad hoc to mimic a vesicle have been exploited as naked LOS carriers, underlining the importance of further exploring these lipid-based vesicles as carriers to deliver anti-gonococcal antigens. In conclusion, data indicated that GMMA-mediated cross-strain bactericidal antibody responses were referable to defined LOS epitopes, suggesting that LOS an important role in the elicitation of functional antibodies following vescicle-based immunization. This result paves the way towards the design of new strategies to develop or improve vaccines against this concerning anti-microbial resistant pathogen.