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It is increasingly being recognised that the interplay between commensal and pathogenic bacteria can dictate the outcome of infection. Consequently, there is a need to understand how commensals interact with their human host and influence pathogen behaviour at epithelial surfaces. Neisseria meningitidis, a leading cause of sepsis and meningitis, exclusively colonises the human nasopharynx and shares this niche with several other Neisseria species, including the commensal Neisseria cinerea. Here, we demonstrate that during adhesion to human epithelial cells N. cinerea co-localises with molecules that are also recruited by the meningococcus, and show that, similar to N. meningitidis, N. cinerea forms dynamic microcolonies on the cell surface in a Type four pilus (Tfp) dependent manner. Finally, we demonstrate that N. cinerea colocalises with N. meningitidis on the epithelial cell surface, limits the size and motility of meningococcal microcolonies, and impairs the effective colonisation of epithelial cells by the pathogen. Our data establish that commensal Neisseria can mimic and affect the behaviour of a pathogen on epithelial cell surfaces.

Neisseria meningitidis and Neisseria gonorrhoeae are two obligate human pathogens that have evolved to be uniquely adapted to their host. The meningococcus is frequently carried asymptomatically in the nasopharynx, while gonococcal infection of the urogenital tract usually elicits a marked local inflammatory response. Other members of the Neisseria genus are abundant in the upper airway where they could engage in co-operative or competitive interactions with both these pathogens. Here, we briefly outline the potential sites of contact between Neisseria spp. in the body, with emphasis on the upper airway, and describe the growing yet circumstantial evidence for antagonism from carriage studies and human volunteer challenge models with Neisseria lactamica . Recent laboratory studies have characterized antagonistic mechanisms that enable competition between Neisseria species. Several of these mechanisms, including Multiple Adhesin family (Mafs), Two Partner Secretion Systems, and Type VI secretion system, involve direct contact between bacteria; the genetic organisation of these systems, and the domain structure of their effector molecules have striking similarities. Additionally, DNA from one species of Neisseria can be toxic to another species, following uptake. More research is needed to define the full repertoire of antagonistic mechanisms in Neisseria spp., their distribution in strains, their range of activity, and contribution to survival in vivo . Understanding the targets of effectors could reveal how antagonistic relationships between close relatives shape subsequent interactions between pathogens and their hosts.

Type VI Secretion Systems (T6SSs) are widespread in bacteria and can dictate the development and organisation of polymicrobial ecosystems by mediating contact dependent killing. In Neisseria species, including Neisseria cinerea a commensal of the human respiratory tract, interbacterial contacts are mediated by Type four pili (Tfp) which promote formation of aggregates and govern the spatial dynamics of growing Neisseria microcolonies. Here, we show that N. cinerea expresses a plasmid-encoded T6SS that is active and can limit growth of related pathogens. We explored the impact of Tfp on N. cinerea T6SS-dependent killing within a colony and show that pilus expression by a prey strain enhances susceptibility to T6SS compared to a non-piliated prey, by preventing segregation from a T6SS-wielding attacker. Our findings have important implications for understanding how spatial constraints during contact-dependent antagonism can shape the evolution of microbial communities.

LBH589 is a novel pan-histone deacetylase (HDAC) inhibitor that has potent antitumor activity in multiple myeloma and other hematological malignancies. However, its impact on the immune system has not been defined. We here evaluated the effects of LBH589 on human myeloid dendritic cells (DCs) at clinically relevant concentrations. Exposure to LBH589 affected the surface molecule expression on immature and mature DCs, which was associated with DC maturation (CD83↓), antigen presentation (human leukocyte antigen-ABC↓) and T-cell co-stimulation (CD40↓ and CD86↑). LBH589 decreased both protein and polysaccharide antigen uptake capacities by DCs. Importantly, LBH589 impaired DC function to stimulate antigen-specific immune responses, resulting in the significant reduction of invariant natural killer T-cell (CD1d-restricted) and T-cell (major histocompatibility complex-restricted) activation in innate and adaptive immunity. LBH589 also significantly repressed the production of interleukin (IL)-6, IL-10, IL-12p70, IL-23 and tumor necrosis factor-α by Toll-like receptor (TLR)3 and TLR4-induced DC activation, indicating an important role of HDAC activity in immune regulation and inflammation. RelB, a component of the nuclear factor-κ B signaling pathway, was the key component regulated by HDAC inhibition in DCs. Together, our preclinical study demonstrates that LBH589 significantly impairs the phenotype and function of DCs, indicating a need for monitoring the immune status in patients receiving HDAC inhibitor therapy. It also provides a rationale to evaluate LBH589 activity for the treatment of inflammation.

There is an urgent need to develop vaccines against pathogenic bacteria. However, this is often hindered by antigenic diversity and difficulties encountered manufacturing membrane proteins. Here we show how to use structure-based design to develop chimeric antigens (ChAs) for subunit vaccines. ChAs are generated against serogroup B Neisseria meningitidis (MenB), the predominant cause of meningococcal disease in wealthy countries. MenB ChAs exploit factor H binding protein (fHbp) as a molecular scaffold to display the immunogenic VR2 epitope from the integral membrane protein PorA. Structural analyses demonstrate fHbp is correctly folded and the PorA VR2 epitope adopts an immunogenic conformation. In mice, immunisation with ChAs generates fHbp and PorA antibodies that recognise the antigens expressed by clinical MenB isolates; these antibody responses correlate with protection against meningococcal disease. Application of ChAs is therefore a potentially powerful approach to develop multivalent subunit vaccines, which can be tailored to circumvent pathogen diversity. Factor H binding protein (fHbp) and PorA are components of experimental serogroup B N. meningitidis vaccines. Here the authors graft the VR2 loop of PorA onto an fHBp-based scaffold to demonstrate proof-of-principle of a chimeric antigen strategy and vaccination against meningococcal disease.

Our work addresses two processes, the maintenance of plasmids and antibiotic tolerance; both contribute to the development of antimicrobial resistance in bacteria that cause human disease. Here, we found a single nucleotide change in the vapBC toxin:antitoxin system that stabilizes the large virulence plasmid of Shigella sonnei . The mutation is in the vapB antitoxin gene and makes the antitoxin more likely to be degraded, releasing the VapC toxin to efficiently kill cells without the plasmid (and thus unable to produce more antitoxin as an antidote). We found that vapBC mutations in E. coli that lead to antibiotic tolerance (a precursor to resistance) also operate by the same mechanism ( i.e. , generating VapB that is prone to cleavage); free VapC during tolerance will arrest bacterial growth and prevent susceptibility to antibiotics. This work shows the mechanistic links between plasmid maintenance and tolerance, and has applications in biotech and in the design and evaluation of vaccines against shigellosis.

Neisseria meningitidis remains an important cause of severe sepsis and meningitis worldwide. The bacterium is only found in human hosts, and so must continually coexist with the immune system. Consequently, N meningitidis uses multiple mechanisms to avoid being killed by antimicrobial proteins, phagocytes, and, crucially, the complement system. Much remains to be learnt about the strategies N meningitidis employs to evade aspects of immune killing, including mimicry of host molecules by bacterial structures such as capsule and lipopolysaccharide, which poses substantial problems for vaccine design. To date, available vaccines only protect individuals against subsets of meningococcal strains. However, two promising vaccines are currently being assessed in clinical trials and appear to offer good prospects for an effective means of protecting individuals against endemic serogroup B disease, which has proven to be a major challenge in vaccine research.

We can now collect cell-count data across whole animal brains quantifying recent neuronal activity, gene expression, or anatomical connectivity. This is a powerful approach since it is a multiregion measurement, but because the imaging is done postmortem, each animal only provides one set of counts. Experiments are expensive, and since cells are counted by imaging and aligning a large number of brain sections, they are time-intensive. The resulting datasets tend to be undersampled with fewer animals than brain regions. As a consequence, these data are a challenge for traditional statistical approaches. We present a ‘standard’ partially pooled Bayesian model for multiregion cell-count data and apply it to two example datasets. These examples demonstrate that hierarchical Bayesian methods are well suited to these data. In both cases, the Bayesian model outperformed standard parallel t -tests. Overall, inference for cell-count data is substantially improved by the ability of the Bayesian approach to capture nested data and by its rigorous handling of uncertainty in undersampled data.

Neisseria meningitidis is a leading cause of sepsis and meningitis. The bacterium recruits factor H (fH), a negative regulator of the complement system, to its surface via fH binding protein (fHbp), providing a mechanism to avoid complement-mediated killing. fHbp is an important antigen that elicits protective immunity against the meningococcus and has been divided into three different variant groups, V1, V2 and V3, or families A and B. However, immunisation with fHbp V1 does not result in cross-protection against V2 and V3 and vice versa. Furthermore, high affinity binding of fH could impair immune responses against fHbp. Here, we investigate a homologue of fHbp in Neisseria gonorrhoeae, designated as Gonococcal homologue of fHbp (Ghfp) which we show is a promising vaccine candidate for N. meningitidis. We demonstrate that Gfhp is not expressed on the surface of the gonococcus and, despite its high level of identity with fHbp, does not bind fH. Substitution of only two amino acids in Ghfp is sufficient to confer fH binding, while the corresponding residues in V3 fHbp are essential for high affinity fH binding. Furthermore, immune responses against Ghfp recognise V1, V2 and V3 fHbps expressed by a range of clinical isolates, and have serum bactericidal activity against N. meningitidis expressing fHbps from all variant groups.

Neisseria meningitis remains a leading cause of sepsis and meningitis, and vaccines are required to prevent infections by this important human pathogen. Factor H binding protein (fHbp) is a key antigen that elicits protective immunity against the meningococcus and recruits the host complement regulator, fH. As the high affinity interaction between fHbp and fH could impair immune responses, we sought to identify non-functional fHbps that could act as effective immunogens. This was achieved by alanine substitution of fHbps from all three variant groups (V1, V2 and V3 fHbp) of the protein; while some residues affected fH binding in each variant group, the distribution of key amino underlying the interaction with fH differed between the V1, V2 and V3 proteins. The atomic structure of V3 fHbp in complex with fH and of the C-terminal barrel of V2 fHbp provide explanations to the differences in the precise nature of their interactions with fH, and the instability of the V2 protein. To develop transgenic models to assess the efficacy of non-functional fHbps, we determined the structural basis of the low level of interaction between fHbp and murine fH; in addition to changes in amino acids in the fHbp binding site, murine fH has a distinct conformation compared with the human protein that would sterically inhibit binding to fHbp. Non-functional V1 fHbps were further characterised by binding and structural studies, and shown in non-transgenic and transgenic mice (expressing chimeric fH that binds fHbp and precisely regulates complement system) to retain their immunogenicity. Our findings provide a catalogue of non-functional fHbps from all variant groups that can be included in new generation meningococcal vaccines, and establish proof-in-principle for clinical studies to compare their efficacy with wild-type fHbps.