Neisseria gonorrhoeae Population Genomics: Use of the Gonococcal Core Genome to Improve Surveillance of Antimicrobial Resistance.

Background: Gonorrhea, caused by the bacterium Neisseria gonorrhoeae, is a globally prevalent sexually transmitted infection. The dynamics of gonococcal population biology have been poorly defined due to a lack of resolution in strain typing methods.

Methods: In this study, we assess how the core genome can be used to improve our understanding of gonococcal population structure compared with current typing schemes.

-Induced Inflammatory Pyroptosis in Human Macrophages is Dependent on Intracellular Gonococci and Lipooligosaccharide.

, the human obligate pathogen responsible for the sexually transmitted disease gonorrhea, has evolved several mechanisms to evade the host immune response. One such mechanism is the modulation of host cell death pathways. In this study, we defined cell death pathways induced by in human monocyte-derived macrophages (MDMs).

Distinct roles for dietary lipids and Porphyromonas gingivalis infection on atherosclerosis progression and the gut microbiota.

Mounting evidence in humans supports an etiological role for the microbiota in inflammatory atherosclerosis. Atherosclerosis is a progressive disease characterized by accumulation of inflammatory cells and lipids in vascular tissue. While retention of lipoprotein into the sub-endothelial vascular layer is believed to be the initiating stimulus leading to the development of atherosclerosis, activation of multiple pathways related to vascular inflammation and endothelial dysfunction sustain the process by stimulating recruitment of leukocytes and immune cells into the sub-endothelial layer.

Microbiota, Immune Subversion, and Chronic Inflammation.

Several host-adapted pathogens and commensals have evolved mechanisms to evade the host innate immune system inducing a state of low-grade inflammation. Epidemiological studies have also documented the association of a subset of these microorganisms with chronic inflammatory disorders. In this review, we summarize recent studies demonstrating the role of the microbiota in chronic inflammatory diseases and discuss how specific microorganisms subvert or inhibit protective signaling normally induced by toll-like receptors (TLRs).

Microbial Degradation of Cellular Kinases Impairs Innate Immune Signaling and Paracrine TNFα Responses.

The NFκB and MAPK signaling pathways are critical components of innate immunity that orchestrate appropriate immune responses to control and eradicate pathogens. Their activation results in the induction of proinflammatory mediators, such as TNFα a potent bioactive molecule commonly secreted by recruited inflammatory cells, allowing for paracrine signaling at the site of an infection. In this study we identified a novel mechanism by which the opportunistic pathogen Porphyromonas gingivalis dampens innate immune responses by disruption of kinase signaling and degradation of inflammatory mediators.

Characterization of the Neisseria gonorrhoeae Iron and Fur Regulatory Network.

Unlabelled: The Neisseria gonorrhoeae ferric uptake regulator (Fur) protein controls expression of iron homeostasis genes in response to intracellular iron levels. In this study, using transcriptome sequencing (RNA-seq) analysis of an N. gonorrhoeae fur strain, we defined the gonococcal Fur and iron regulons and characterized Fur-controlled expression of an ArsR-like DNA binding protein.

A high-throughput method to examine protein-nucleotide interactions identifies targets of the bacterial transcriptional regulatory protein fur.

The Ferric uptake regulatory protein (Fur) is a transcriptional regulatory protein that functions to control gene transcription in response to iron in a number of pathogenic bacteria. In this study, we applied a label-free, quantitative and high-throughput analysis method, Interferometric Reflectance Imaging Sensor (IRIS), to rapidly characterize Fur-DNA interactions in vitro with predicted Fur binding sequences in the genome of Neisseria gonorrhoeae, the causative agent of the sexually transmitted disease gonorrhea. IRIS can easily be applied to examine multiple protein-protein, protein-nucleotide and nucleotide-nucleotide complexes simultaneously and demonstrated here that seventy percent of the predicted Fur boxes in promoter regions of iron-induced genes bound to Fur in vitro with a range of affinities as observed using this microarray screening technology.

Fur-mediated global regulatory circuits in pathogenic Neisseria species.

The ferric uptake regulator (Fur) protein has been shown to function as a repressor of transcription in a number of diverse microorganisms. However, recent studies have established that Fur can function at a global level as both an activator and a repressor of transcription through both direct and indirect mechanisms. Fur-mediated indirect activation occurs via the repression of additional repressor proteins, or small regulatory RNAs, thereby activating transcription of a previously silent gene.

Pathogen-mediated proteolysis of the cell death regulator RIPK1 and the host defense modulator RIPK2 in human aortic endothelial cells.

Porphyromonas gingivalis is the primary etiologic agent of periodontal disease that is associated with other human chronic inflammatory diseases, including atherosclerosis. The ability of P. gingivalis to invade and persist within human aortic endothelial cells (HAEC) has been postulated to contribute to a low to moderate chronic state of inflammation, although how this is specifically achieved has not been well defined.

Fur-mediated activation of gene transcription in the human pathogen Neisseria gonorrhoeae.

It is well established that the ferric uptake regulatory protein (Fur) functions as a transcriptional repressor in diverse microorganisms. Recent studies demonstrated that Fur also functions as a transcriptional activator. In this study we defined Fur-mediated activation of gene transcription in the sexually transmitted disease pathogen Neisseria gonorrhoeae.

Macrophage-elicited osteoclastogenesis in response to bacterial stimulation requires Toll-like receptor 2-dependent tumor necrosis factor-alpha production.

The receptor activator of NF-kappaB ligand (RANKL) and the proinflammatory cytokines are believed to play important roles in osteoclastogenesis. We recently reported that the innate immune recognition receptor, Toll-like receptor 2 (TLR2), is crucial for inflammatory bone loss in response to infection by Porphyromonas gingivalis, the primary organism associated with chronic inflammatory periodontal disease. However, the contribution of macrophage-expressed TLRs to osteoclastogenesis has not been defined.

Roles of the host oxidative immune response and bacterial antioxidant rubrerythrin during Porphyromonas gingivalis infection.

The efficient clearance of microbes by neutrophils requires the concerted action of reactive oxygen species and microbicidal components within leukocyte secretory granules. Rubrerythrin (Rbr) is a nonheme iron protein that protects many air-sensitive bacteria against oxidative stress. Using oxidative burst-knockout (NADPH oxidase-null) mice and an rbr gene knockout bacterial strain, we investigated the interplay between the phagocytic oxidative burst of the host and the oxidative stress response of the anaerobic periodontal pathogen Porphyromonas gingivalis.

Fimbria-dependent activation of pro-inflammatory molecules in Porphyromonas gingivalis infected human aortic endothelial cells.

Epidemiological studies support that chronic periodontal infections are associated with an increased risk of cardiovascular disease. Previously, we reported that the periodontal pathogen Porphyromonas gingivalis accelerated atherosclerotic plaque formation in hyperlipidemic apoE-/- mice, while an isogenic fimbria-deficient (FimA-) mutant did not. In this study, we utilized 41 kDa (major) and 67 kDa (minor) fimbria mutants to demonstrate that major fimbria are required for efficient P.

Pathogen-accelerated atherosclerosis occurs early after exposure and can be prevented via immunization.

Here we report on early inflammatory events associated with Porphyromonas gingivalis-accelerated atherosclerosis in apolipoprotein E knockout (ApoE-/-) mice. Animals challenged with P. gingivalis presented with increased macrophage infiltration, innate immune marker expression, and atheroma without elevated systemic inflammatory mediators.

Identification of amino acid residues involved in heme binding and hemoprotein utilization in the Porphyromonas gingivalis heme receptor HmuR.

We have previously identified and characterized a heme/hemoglobin receptor, HmuR, in Porphyromonas gingivalis. To analyze the conserved amino acid residues of HmuR that may be involved in hemin/hemoprotein binding and utilization, we constructed a series of P. gingivalis A7436 hmuR mutants with amino acid replacements and characterized the ability of these mutants to utilize hemin and hemoproteins.

Gingipain-specific IgG in the sera of patients with periodontal disease is necessary for opsonophagocytosis of Porphyromonas gingivalis.

Background: Porphyromonas gingivalis is a primary etiologic agent of generalized aggressive periodontitis (GAgP), and gingipains, a group of cysteine proteinases, are critical virulence factors expressed by this organism. GAgP patients develop specific antibodies to gingipains; however, the function of these antibodies in the clearance of P. gingivalis infection is poorly understood.

Iron and heme utilization in Porphyromonas gingivalis.

Porphyromonas gingivalis is a Gram-negative anaerobic bacterium associated with the initiation and progression of adult periodontal disease. Iron is utilized by this pathogen in the form of heme and has been shown to play an essential role in its growth and virulence. Recently, considerable attention has been given to the characterization of various secreted and surface-associated proteins of P.

Coordinate expression of the Porphyromonas gingivalis lysine-specific gingipain proteinase, Kgp, arginine-specific gingipain proteinase, RgpA, and the heme/hemoglobin receptor, HmuR.

Heme utilization in Porphyromonas gingivalis requires the participation of an outer membrane hemin/hemoglobin receptor, HmuR, the lysine-specific gingipain proteinase (Kgp) and arginine-specific gingipain proteinase (Rgp). In this study, the expression of hmuR , kgp and rgpA genes in response to growth with different heme sources was examined by reverse transcription-polymerase chain reaction and enzyme-linked immunoassay. Coordinate regulation of hmuR , kgp and rgpA gene expression was evaluated through utilization of P.

Innate immune recognition of invasive bacteria accelerates atherosclerosis in apolipoprotein E-deficient mice.

Background: Infectious diseases have emerged as potential risk factors for cardiovascular disease (CVD). Epidemiological studies support a connection between periodontal disease, a chronic inflammatory disease of the supporting tissues of the teeth, and CVD.

Methods And Results: To directly test the connection between periodontal disease and atherosclerosis, apoE-/- mice were orally challenged with the periodontal disease pathogen Porphyromonas gingivalis or an invasion-impaired P gingivalis fimbriae-deficient mutant (FimA-).

Porphyromonas gingivalis-specific immunoglobulin G prevents P. gingivalis-elicited oral bone loss in a murine model.

Active immunization with Porphyromonas gingivalis whole-cell preparations has been shown to prevent P. gingivalis infection and oral bone loss. Employing passive antibody transfer and opsonization, we demonstrate with this study that immunization-elicited P.

Identification of iron-activated and -repressed Fur-dependent genes by transcriptome analysis of Neisseria meningitidis group B.

Iron is limiting in the human host, and bacterial pathogens respond to this environment by activating genes required for bacterial virulence. Transcriptional regulation in response to iron in Gram-negative bacteria is largely mediated by the ferric uptake regulator protein Fur, which in the presence of iron binds to a specific sequence in the promoter regions of genes under its control and acts as a repressor. Here we describe DNA microarray, computational and in vitro studies to define the Fur regulon in the human pathogen Neisseria meningitidis group B (strain MC58).

Antimicrobial activity of magainin analogues against anaerobic oral pathogens.

Magainins are a family of potent antimicrobial cationic peptides that possess antimicrobial activity against a wide range of target organisms. In this study, the antimicrobial activity of synthetic magainin-mimetic compounds MSI-751 and MSI-774 was investigated against the periodontal pathogens Porphyromonas gingivalis, Fusobacterium nucleatum, Actinobacillus actinomycetemcomitans, Eikenella corrodens, Prevotella loescheii and Prevotella intermedia. P.

The gonococcal fur regulon: identification of additional genes involved in major catabolic, recombination, and secretory pathways.

In this study, we have characterized the in vitro binding of Neisseria gonorrhoeae Fur to several well-defined iron transport genes, as well as to additional genes involved in major catabolic, secretory, and recombination pathways of gonococci. The gonococcal Fur protein was recombinantly expressed in Escherichia coli HBMV119. Fur was isolated from inclusion bodies and partially purified by ion-exchange chromatography.

Role for fimbriae and lysine-specific cysteine proteinase gingipain K in expression of interleukin-8 and monocyte chemoattractant protein in Porphyromonas gingivalis-infected endothelial cells.

Recent cross-sectional and prospective epidemiological studies have demonstrated an association between periodontal disease and atherosclerosis and human coronary heart disease. Previously, we have established that the periodontal pathogen Porphyromonas gingivalis is capable of invading aortic, heart, and human umbilical vein endothelial cells (HUVEC). Since atherosclerosis is a chronic inflammatory response initiated at the vascular wall, interactions of P.