Strategies to Tackle Antimicrobial Resistance: The Example of Escherichia coli and Pseudomonas aeruginosa.

Traditional antimicrobial treatments consist of drugs which target different essential functions in pathogens. Nevertheless, bacteria continue to evolve new mechanisms to evade this drug-mediated killing with surprising speed on the deployment of each new drug and antibiotic worldwide, a phenomenon called antimicrobial resistance (AMR). Nowadays, AMR represents a critical health threat, for which new medical interventions are urgently needed. By 2050, it is estimated that the leading cause of death will be through untreatable AMR pathogens. Although antibiotics remain a first-line treatment, non-antibiotic therapies such as prophylactic vaccines and therapeutic monoclonal antibodies (mAbs) are increasingly interesting alternatives to limit the spread of such antibiotic resistant microorganisms. For the discovery of new vaccines and mAbs, the search for effective antigens that are able to raise protective immune responses is a challenging undertaking. In this context, outer membrane vesicles (OMV) represent a promising approach, as they recapitulate the complete antigen repertoire that occurs on the surface of Gram-negative bacteria. In this review, we present Escherichia coli and Pseudomonas aeruginosa as specific examples of key AMR threats caused by Gram-negative bacteria and we discuss the current status of mAbs and vaccine approaches under development as well as how knowledge on OMV could benefit antigen discovery strategies.

Neisserial adhesin A (NadA) binds human Siglec-5 and Siglec-14 with high affinity and promotes bacterial adhesion/invasion.

Neisserial adhesin A (NadA) is a meningococcal surface protein included as recombinant antigen in 4CMenB, a protein-based vaccine able to induce protective immune responses against Neisseria meningitidis serogroup B (MenB). Although NadA is involved in the adhesion/invasion of epithelial cells and human myeloid cells, its function in meningococcal physiology is still poorly understood. To clarify the role played by NadA in the host-pathogen interaction, we sought to identify its cellular receptors. We screened a protein microarray encompassing 2,846 human and 297 mouse surface/secreted recombinant proteins using recombinant NadA as probe. Efficient NadA binding was revealed on the paired sialic acid-binding immunoglobulin-type lectins receptors 5 and 14 (Siglec-5 and Siglec-14), but not on Siglec-9 therein used as control. The interaction was confirmed by biochemical tools with the determination of the KD value in the order of nanomolar and the identification of the NadA binding site by hydrogen-deuterium exchange coupled to mass spectrometry. The N-terminal domain of the Siglec-5 that recognizes the sialic acid was identified as the NadA binding domain. Intriguingly, exogenously added recombinant soluble Siglecs, including Siglec-9, were found to decorate N. meningitidis surface in a NadA-dependent manner. However, Siglec-5 and Siglec-14 transiently expressed in CHO-K1 cells endorsed NadA binding and increased N. meningitidis adhesion/invasion while Siglec-9 did not. Taken together, Siglec-5 and Siglec-14 satisfy all features of NadA receptors suggesting a possible role of NadA in the acute meningococcal infection.IMPORTANCEBacteria have developed several strategies for cell colonization and immune evasion. Knowledge of the host and pathogen factors involved in these mechanisms is crucial to build efficacious countermoves. Neisserial adhesin A (NadA) is a meningococcal surface protein included in the anti-meningococcus B vaccine 4CMenB, which mediates adhesion to and invasion of epithelial cells. Although NadA has been shown to bind to other cell types, like myeloid and endothelial cells, it still remains orphan of a defined host receptor. We have identified two strong NadA interactors, Siglec-5 and Siglec-14, which are mainly expressed on myeloid cells. This showcases that NadA is an additional and key player among the Neisseria meningitidis factors targeting immune cells. We thus provide novel insights on the strategies exploited by N. meningitidis during the infection process, which can progress to a severe illness and death.

OpcA and PorB are novel bactericidal antigens of the 4CMenB vaccine in mice and humans.

The ability of Neisseria meningitidis Outer Membrane Vesicles (OMV) to induce protective responses in humans is well established and mainly attributed to Porin A (PorA). However, the contribution of additional protein antigens to protection remains to be elucidated. In this study we dissected the immunogenicity of antigens originating from the OMV component of the 4CMenB vaccine in mice and humans. We collected functional data on a panel of strains for which bactericidal responses to 4CMenB in infants was attributable to the OMV component and evaluated the role of 30 OMV-specific protein antigens in cross-coverage. By using tailor-made protein microarrays, the immunosignature of OMV antigens was determined. Three of these proteins, OpcA, NspA, and PorB, triggered mouse antibodies that were bactericidal against several N. meningitidis strains. Finally, by genetic deletion and/or serum depletion studies, we demonstrated the ability of OpcA and PorB to induce functional immune responses in infant sera after vaccination. In conclusion, while confirming the role of PorA in eliciting protective immunity, we identified two OMV antigens playing a key role in protection of infants vaccinated with the 4CMenB vaccine against different N. meningitidis serogroup B strains.

Synergistic activity of antibodies in the multicomponent 4CMenB vaccine.

INTRODUCTION: Vaccines based on multiple antigens often induce an immune response, which is higher than that triggered by each single component, with antibodies acting cooperatively and synergistically in tackling the infection. AREAS COVERED: An interesting example is the antibody response induced by the 4CMenB vaccine, currently licensed for the prevention of Neisseria meningitidis serogroup B (MenB). It contains four antigenic components: Factor H binding protein (fHbp), Neisseria adhesin A (NadA), Neisserial Heparin Binding Antigen (NHBA), and Outer Membrane Vesicles (OMV). Monoclonal and polyclonal antibodies raised by vaccination with 4CMenB show synergistic activity in complement-dependent bacterial killing. This review summarizes published and unpublished data and provides evidence of the added value of multicomponent vaccines. EXPERT OPINION: The ability of 4CMenB vaccine to elicit antibodies targeting multiple surface-exposed antigens is corroborated by the recent data on real-world evidences. Bactericidal activity is generally mediated by antibodies that bind to antigens highly expressed on the bacterial surface and immunologically related. However, simultaneous binding of antibodies to various surface-exposed antigens can overcome the threshold density of antigen-antibody complexes needed for complement activation. The data discussed in this review highlight the interplay between antibodies targeting major and minor antigens and their effect on functionality. CLINICAL TRIAL REGISTRATION: www.clinicaltrials.gov identifiers of studies with original data mentioned in the article: NCT00937521, NCT00433914, NCT02140762, and NCT02285777.