Urgent Need to Understand and Prevent Gonococcal Infection: From the Laboratory to Real-World Context.

Neisseria gonorrhoeae is widespread globally. Primary prevention is unsuccessful and antimicrobial resistance threatens optimal management. There is no specific vaccine and natural infection studies show that N. gonorrhoeae can avoid and suppress immune responses. In addition to extensive variation in expression and specificity of many gonococcal surface antigens, it induces a robust inflammatory response through the Th17 pathway with a large influx of neutrophils and inflammatory cytokines but evades macrophages. The Th1- and Th2-mediated response is suppressed, resulting in low, short-lived antibody titers. Real-world evidence suggests that gonorrhea cases are reduced among recipients of N. meningitidis group B vaccines containing outer membrane vesicles (OMV). Although the first randomized trial of an OMV-containing MenB vaccine against N. gonorrhoeae infection did not show statistically significant vaccine efficacy, ongoing trials might shed further light. Several candidate vaccine antigens for a gonococcal-specific vaccine are being evaluated preclinically but only one has reached clinical trials.

Enhancing a multi-purpose artificial urine for culture and gene expression studies of uropathogenic Escherichia coli strains.

AIMS: The main objective of this study was to modify a recently reported multi-purpose artificial urine (MP-AU) for culture and gene expression studies of uropathogenic Escherichia coli (UPEC) strains. METHODS AND RESULTS: We used liquid chromatography mass spectrometry (LC-MS) to identify and adjust the metabolic profile of MP-AU closer to that of pooled human urine (PHU). Modification in this way facilitated growth of UPEC strains with growth rates similar to those obtained in PHU. Transcriptomic analysis of UPEC strains cultured in enhanced artificial urine (enhanced AU) and PHU showed that the gene expression profiles are similar, with <7% of genes differentially expressed between the two conditions. CONCLUSIONS: Enhancing an MP-AU with metabolites identified in PHU allows the enhanced AU to be used as a substitute for the culture and in vitro gene expression studies of UPEC strains.

4CMenB journey to the 10-year anniversary and beyond.

The 4-component meningococcal serogroup B (MenB) vaccine, 4CMenB, the first broadly protective, protein-based MenB vaccine to be licensed, is now registered in more than 50 countries worldwide. Real-world evidence (RWE) from the last decade confirms its effectiveness and impact, with infant immunization programs showing vaccine effectiveness of 71-95% against invasive MenB disease and cross-protection against non-B serogroups, including a 69% decrease in serogroup W cases in 4CMenB-eligible cohorts in England. RWE from different countries also demonstrates the potential for additional moderate protection against gonorrhea in adolescents. The real-world safety profile of 4CMenB is consistent with prelicensure reports. Use of the endogenous complement human serum bactericidal antibody (enc-hSBA) assay against 110 MenB strains may enable assessment of the immunological effectiveness of multicomponent MenB vaccines in clinical trial settings. Equitable access to 4CMenB vaccination is required to better protect all age groups, including older adults, and vulnerable groups through comprehensive immunization policies.

Invasive meningococcal disease, caused by the bacterium Neisseria meningitidis(meningococcus), is rare but often devastating and can be deadly. Effective vaccines are available, including vaccines against meningococcal serogroup B disease. In 2013, the 4-component meningococcal serogroup B vaccine, 4CMenB, became the first broadly protective, protein-based vaccine against serogroup B to be licensed, with the second (bivalent vaccine, MenB-FHbp) licensed the following year. 4CMenB is now registered in more than 50 countries, in the majority, for infants and all age groups. In the US, it is approved for individuals aged 10­25 years. Evidence from immunization programs in the last decade, comparing vaccinated and unvaccinated individuals and the same population before and after vaccination, confirms the effectiveness and positive impact of 4CMenB against serogroup B disease. This also demonstrates that 4CMenB can provide protection against invasive diseases caused by other meningococcal serogroups. Furthermore, N. meningitidis is closely related to the bacterium that causes gonorrhea, N. gonorrhoeae, and emerging real-world evidence suggests that 4CMenB provides additional moderate protection against gonococcal disease. The safety of 4CMenB when given to large numbers of infants, children, adolescents, and adults is consistent with the 4CMenB safety profile reported before licensure.For the future, it would be beneficial to address differences among national guidelines for the recommended administration of 4CMenB, particularly where there is supportive epidemiological evidence but no equitable access to vaccination. New assays for assessing the potential effectiveness of meningococcal serogroup B vaccines in clinical trials are also required because serogroup B strains circulating in the population are extremely diverse across different countries.

Development of a visual Adhesion/Invasion Inhibition Assay to assess the functionality of Shigella-specific antibodies.

Introduction: Shigella is the etiologic agent of a bacillary dysentery known as shigellosis, which causes millions of infections and thousands of deaths worldwide each year due to Shigella's unique lifestyle within intestinal epithelial cells. Cell adhesion/invasion assays have been extensively used not only to identify targets mediating host-pathogen interaction, but also to evaluate the ability of Shigella-specific antibodies to reduce virulence. However, these assays are time-consuming and labor-intensive and fail to assess differences at the single-cell level. Objectives and methods: Here, we developed a simple, fast and high-content method named visual Adhesion/Invasion Inhibition Assay (vAIA) to measure the ability of anti-Shigellaantibodies to inhibit bacterial adhesion to and invasion of epithelial cells by using the confocal microscope Opera Phenix. Results: We showed that vAIA performed well with a pooled human serum from subjects challenged with S. sonnei and that a specific anti-IpaD monoclonal antibody effectively reduced bacterial virulence in a dose-dependent manner. Discussion: vAIA can therefore inform on the functionality of polyclonal and monoclonal responses thereby supporting the discovery of pathogenicity mechanisms and the development of candidate vaccines and immunotherapies. Lastly, this assay is very versatile and may be easily applied to other Shigella species or serotypes and to different pathogens.

Genetic characterization and estimated 4CMenB vaccine strain coverage of 284 Neisseria meningitidis isolates causing invasive meningococcal disease in Argentina in 2010-2014.

Meningococcal (Neisseria meningitidis) serogroup B (MenB) strain antigens are diverse and a limited number of strains can be evaluated using the human serum bactericidal antibody (hSBA) assay. The genetic Meningococcal Antigen Typing System (gMATS) was developed to predict the likelihood of coverage for large numbers of isolates by the 4CMenB vaccine, which includes antigens Neisseria adhesin A (NadA), Neisserial Heparin-Binding Antigen (NHBA), factor H-binding protein (fHbp), and Porin A (PorA). In this study, we characterized by whole-genome analyses 284 invasive MenB isolates collected from 2010 to 2014 by the Argentinian National Laboratories Network (52-61 isolates per year). Strain coverage was estimated by gMATS on all isolates and by hSBA assay on 74 randomly selected isolates, representative of the whole panel. The four most common clonal complexes (CCs), accounting for 81.3% of isolates, were CC-865 (75 isolates, 26.4%), CC-32 (59, 20.8%), CC-35 (59, 20.8%), and CC-41/44 (38, 13.4%). Vaccine antigen genotyping showed diversity. The most prevalent variants/peptides were fHbp variant 2, NHBA peptides 24, 21, and 2, and PorA variable region 2 profiles 16-36 and 14. The nadA gene was present in 66 (23.2%) isolates. Estimated strain coverage by hSBA assay showed 78.4% of isolates were killed by pooled adolescent sera, and 51.4% and 64.9% (based on two different thresholds) were killed by pooled infant sera. Estimated coverage by gMATS (61.3%; prediction interval: 55.5%, 66.7%) was consistent with the infant hSBA assay results. Continued genomic surveillance is needed to evaluate the persistence of major MenB CCs in Argentina.

The most common clinical manifestations of invasive meningococcal disease include meningitis and septicemia, which can be deadly, and many survivors suffer long-term serious after-effects. Most cases of invasive meningococcal disease are caused by six meningococcal serogroups (types), including serogroup B. Although vaccines are available against meningococcal serogroup B infection, these vaccines target antigens that are highly diverse. Consequently, the effectiveness of vaccination may vary from country to country because the meningococcal serogroup B strains circulating in particular regions carry different forms of the target vaccine antigens. This means it is important to test serogroup B strains isolated from specific populations to estimate the percentage of strains that a vaccine is likely to be effective against (known as 'vaccine strain coverage'). The genetic Meningococcal Antigen Typing System (gMATS) was developed to predict strain coverage by the four-component meningococcal serogroup B vaccine, 4CMenB, against large numbers of serogroup B strains. In this study, we analyzed 284 invasive meningococcal serogroup B isolates collected between 2010 and 2014 in Argentina. Genetic analyses showed that the vaccine antigens of the isolates were diverse and some genetic characteristics had not been found in isolates from other countries. However, vaccine strain coverage estimated by gMATS was consistent with that reported in other parts of the world and with strain coverage results obtained for a subset via another method, the human serum bactericidal antibody (hSBA) assay. These results highlight the need for continued monitoring of circulating bacterial strains to assess the estimated strain coverage of meningococcal serogroup B vaccines.

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.

Bactericidal killing of meningococcal W strains isolated in Argentina by the sera of adolescents and infants immunized with 4-component meningococcal serogroup B vaccine (4CMenB).

Invasive meningococcal disease (IMD) is a life-threatening disease caused by meningococcal serogroups A, B, C, W, X, and Y, of which B and W are most common in Argentina. The 4-component meningococcal serogroup B (4CMenB) vaccine contains three purified recombinant protein antigens (Neisseria adhesin A [NadA], factor H binding protein [fHbp], and Neisserial Heparin Binding Antigen [NHBA]) and outer membrane vesicles (OMV), which is derived from the New Zealand epidemic strain and contains Porin A 1.4. These antigens are present and conserved in strains that belong to other serogroups. In this study, we show that 10/11 (91%) meningococcal serogroup W (MenW) strains selected to be representative of MenW isolates that caused IMD in Argentina during 2010-2011 were killed in bactericidal assays by the sera of adolescents and infants who had been immunized with the 4CMenB vaccine. We also show that MenW strains that caused IMD in Argentina during 2018-2021 were genetically similar to the earlier strains, indicating that the 4CMenB vaccine would likely still provide protection against current MenW strains. These data highlight the potential of 4CMenB vaccination to protect adolescents and infants against MenW strains that are endemic in Argentina.