Abstract: Meningococci are facultative-pathogenic bacteria endowed with a set of adhesins allowing colonization of the human upper respiratory tract, leading to fulminant meningitis and septicemia. The Neisseria adhesin NadA was identified in about 50 % of N. meningitidis isolates and is closely related to the Yersinia adhesin YadA, the prototype of the oligomeric coiled-coil adhesin (Oca) family. NadA is known to be involved in cell adhesion, invasion and induction of proinflammatory cytokines. Due to the enormous diversity of neisserial cell adhesins the analysis of the specific contribution of NadA in meningococcal host interactions is limited. Therefore we used a non-invasive Y. enterocolitica mutant as carrier to study the role of NadA in host cell interaction. NadA was shown to be efficiently produced and localized in its oligomeric form on the bacterial surface of Y. enterocolitica. Additionally, NadA mediated a β1 integrin-dependent adherence with subsequent internalization of yersiniae by a β1 integrin-positive cell line. Using recombinant NadA24-210 protein and human and murine β1 integrin expressing cell lines we could demonstrate the role of the β1 integrin subunit as putative receptor for NadA. Subsequent inhibition assays revealed specific interaction of NadA24-210 with the human β1 integrin subunit. Cumulatively, these results indicate that Y. enterocolitica is a suitable toolbox system for analysis of the adhesive properties of NadA, revealing strong evidence that β1 integrins are important receptors for NadA. Thus, this study demonstrated for the first time a direct interaction between the Oca-family member NadA and human β1 integrins.
Abstract: Transcriptional regulators play an important role for the survival of Neisseria meningitidis within its human host. We have recently shown that FarR acts as transcriptional repressor of the adhesin nadA in N. meningitidis. Here, we examined the FarR regulon by microarray analyses, qRT-PCR, and electrophoretic mobility shift assays, revealing that FarR is a highly specific repressor of nadA. We demonstrate by reporter gene fusion assays that alterations of the FarR binding site within the nadA promoter are sufficient to induce transcription of nadA. Furthermore, farR expression is growth phase-dependent. The highest transcription rate was observed in the late-exponential growth phase of meningococci. Upon contact with active components of the complement system in normal human serum, expression of farR is slightly downregulated. Concluding, we present FarR as an exquisitely specialized, growth phase-dependent, possibly complement-responsive transcriptional regulator in N. meningitidis.
Abstract: Free fatty acids are important antimicrobial substances regulating the homeostasis of colonizing bacteria on epithelial surfaces. Here, we show that meningococci express a functional farAB efflux pump, which is indispensable for fatty acid resistance. However, other than in Neisseria gonorrhoeae, the transcriptional regulator FarR is not involved in regulation of this operon in Neisseria meningitidis. We tested the susceptibility of 23 meningococcal isolates against saturated and unsaturated long-chain fatty acids, proving that meningococci are generally highly resistant, with the exception of serogroup Y strains belonging to sequence type 23. Using genetically determined lipopolysaccharide (LPS)-truncated mutant strains, we show that addition of the LPS core oligosaccharide and hexa-acylation of its membrane anchor lipid A are imperative for fatty acid resistance of meningococci. The sensitivity of the serogroup Y strains is due to naturally occurring mutations within the lpxL1 gene, which is responsible for addition of the sixth acyl chain on the LPS membrane anchor lipid A. Therefore, fatty acid resistance in meningococci is provided by both the active efflux pump FarAB and by the natural permeability barrier of the Gram-negative outer membrane. The transcriptional regulator FarR is not implicated in fatty acid resistance in meningococci, possibly giving rise to a constitutively active FarAB efflux pump system and thus revealing diverse mechanisms of niche adaptation in the two closely related Neisseria species.
Abstract: Neisseria meningitidis and Neisseria gonorrhoeae are the only pathogenic species of the genus Neisseria. Although these two species are closely related, they specialized on survival in completely different environments within the human host-the nasopharynx in the case of N. meningitidis versus the urogenital tract in the case of N. gonorrhoeae. The genetic background of these differences has not yet been determined. Here, we present a comparison of all characterized transcriptional regulators in these species, delineating analogous functions and disclosing differential functional developments of these DNA-binding proteins with a special focus on the recently characterized regulator FarR and its contribution to divergent host niche adaptation in the two Neisseria spp. Furthermore, we summarize the present knowledge on two-partner secretion systems in meningococci, highlighting their overall expression among meningococcal strains in contrast to the complete absence in gonococci. Concluding, the decisive role of these two entirely different factors in host niche adaptation of the two human pathogenic Neisseria species is depicted, illuminating another piece of the puzzle to locate the molecular basis of their differences in preferred colonization sites and pathogenicity.
Abstract: Two closely related pathogenic species have evolved in the genus Neisseria: N. meningitidis and N. gonorrhoeae, which occupy different host niches and cause different clinical entities. In contrast to the pathogen N. gonorrhoeae, N. meningitidis is a commensal and only rarely becomes invasive. Little is known about the genetic background of the entirely different lifestyles in these closely related species. Meningococcal NMB1843 encodes a transcriptional regulator of the MarR family. The gonococcal homologue FarR regulates expression of farAB, mediating fatty acid resistance. We show that NmFarR also directly interacts with NmfarAB. Yet, by contrast to N. gonorrhoeae, no significant sensitivity to fatty acids was observed in a DeltafarR mutant due to intrinsic resistance of meningococci. Further analyses identified an NmFarR-repressed protein absent from N. gonorrhoeae. This protein is the meningococcus-specific adhesin and vaccine component NadA that has most likely been acquired by horizontal gene transfer. NmFarR binds to a 16 base pair palindromic repeat within the nadA promoter. De-repression of nadA resulted in significantly higher association of a DeltafarR strain with epithelial cells. Hence NmFarR has gained control over a meningococcus-specific gene involved in host colonization and thus contributed to divergent niche adaptation in pathogenic Neisseriae.
Abstract: Proinflammatory cytokines play a major role in the pathogenesis of meningococcal disease and their serum levels in patients are correlated with the outcome of infection. Dendritic cells initiate immunity against Neisseria meningitidis and are a major source of proinflammatory cytokines. Here we show that physical interaction of human DC with N. meningitidis via the class A scavenger receptor (SRA) modulates cytokine release by DC. Phagocytosis and uptake via SRA were shown to increase release of TNF-alpha, IL-1 beta and IL-6. In contrast, secretion of IL-8 is enhanced after recognition of N. meningitidis via SRA and not dependent on phagocytosis. Binding of N. meningitidis results in dephosphorylation of SRA but not in upregulation of SRA transcription. Unlike previously thought, not all meningococcal strains are recognized via SRA. A constitutively unencapsulated carriage isolate could be shown not to bind to SRA and induce proinflammatory cytokines independent of this receptor. In conclusion, recognition via SRA by dendritic cells is likely to play a central role in the immune response to N. meningitidis.
