Promising developments in gonococcal vaccines.

PURPOSE OF REVIEW: While effective vaccines to prevent invasive infections by Neisseria meningitidis have been deployed around the world, development of a vaccine to prevent Neisseria gonorrhoeae has lagged. After multiple failed vaccine candidates, vaccine development for N. gonorrhoeae is showing promise for the first time in several decades. This review highlights recent progress in the field. RECENT FINDINGS: Vaccines containing outer-membrane vesicles (OMV) have been used to manage outbreaks of the serogroup B N. meningitidis in a number of countries. Epidemiologic studies indicate these vaccination campaigns were associated with reductions in reported N. gonorrhoeae infections. Recently, a serogroup B N. meningitidis vaccine containing both recombinant antigens and OMV has been licensed through much of the world. Epidemiologic studies also demonstrate associations between 4CMenB immunization and reduced N. gonorrhoeae infections. Additionally, mathematical modeling studies have begun to identify potential strategies for vaccine deployment to maximize reduction of infections. SUMMARY: After several decades with little progress towards an effective gonococcal vaccine, large observational studies have provided evidence that a new generation of group B N. meningitidis vaccines containing OMV have serendipitously restarted the field. Ongoing clinical trials will soon provide definitive evidence regarding the efficacy of these vaccines in preventing N. gonorrhoeae infection.

Neisseria gonorrhoeae MtrCDE Efflux Pump During In Vivo Experimental Genital Tract Infection in Men and Mice Reveals the Presence of Within-Host Colonization Bottleneck.

The MtrCDE efflux pump of Neisseria gonorrhoeae exports a wide range of antimicrobial compounds that the gonococcus encounters at mucosal surfaces during colonization and infection. Here, we evaluate the role of this efflux pump system in strain FA1090 in human male urethral infection with a Controlled Human Infection Model. Using the strategy of competitive multi-strain infection with wild-type FA1090 and an isogenic mutant strain that does not contain a functional MtrCDE pump, we found that the presence of the efflux pump during human experimental infection did not confer a competitive advantage. This finding is in contrast to previous findings in female mice, which demonstrated that gonococci of strain FA19 lacking a functional MtrCDE pump had a significantly reduced fitness compared to the wild type strain in the lower genital tract of female mice. We conducted competitive infections in female mice with FA19 and FA1090 strains, including mutants that do not assemble a functional Mtr efflux pump, demonstrating the fitness advantage provided byt the MtrCDE efflux pump during infection of mice is strain dependent. Our data indicate that new gonorrhea treatment strategies targeting the MtrCDE efflux pump functions may not be universally efficacious in naturally occurring infections. Owing to the equal fitness of FA1090 strains in men, our experiments unexpectedly demonstrated the likely presence of an early colonization bottleneck of N. gonorrhoeae in the human male urethra. TRIAL REGISTRATION: Clinicaltrials.gov NCT03840811 .

Commensal Neisseria species share immune suppressive mechanisms with Neisseria gonorrhoeae.

Neisseria gonorrhoeae is a highly adapted human sexually transmitted pathogen that can cause symptomatic infections associated with localized inflammation as well as asymptomatic and subclinical infections, particularly in females. Gonococcal infection in humans does not generate an effective immune response in most cases, which contributes to both transmission of the pathogen and reinfection after treatment. Neisseria gonorrhoeae is known to evade and suppress human immune responses through a variety of mechanisms. Commensal Neisseria species that are closely related to N. gonorrhoeae, such as N. cinerea, N. lactamica, N. elongata, and N. mucosa, rarely cause disease and instead asymptomatically colonize mucosal sites for prolonged periods of time without evoking clearing immunologic responses. We have shown previously that N. gonorrhoeae inhibits the capacity of antigen-pulsed dendritic cells to induce CD4+ T cell proliferation in vitro. Much of the suppressive effects of N. gonorrhoeae on dendritic cells can be recapitulated either by outer-membrane vesicles released from the bacteria or by purified PorB, the most abundant outer-membrane protein in Neisseria gonorrhoeae. We show here that three commensal Neisseria species, N. cinerea, N. lactamica and N. mucosa, show a comparable capacity to suppress dendritic cell-induced T cell proliferation in vitro through mechanisms similar to those demonstrated previously for N. gonorrhoeae, including inhibition by purified PorB. Our findings suggest that some immune-evasive properties of pathogenic N. gonorrhoeae are shared with commensal Neisseria species and may contribute to the ability of both pathogens and commensals to cause prolonged mucosal colonization in humans.

The role of the genital microbiota in the acquisition and pathogenesis of sexually transmitted infections.

PURPOSE OF REVIEW: There are an estimated 374 million new sexually transmitted infections (STIs) worldwide every year. Our review article examines the current evidence of how STI acquisition, transmission, and pathogenesis is impacted upon by the genital microbiota, with a focus on epidemiological, biochemical, and immunological features. RECENT FINDINGS: At least in women, a genital microbiota dominated by lactobacilli has long been considered optimal for reproductive health, while depletion of lactobacilli may lead to a genital microenvironment dominated by anaerobic pathogens, which can manifest clinically as bacterial vaginosis. Recent research efforts have characterized genital microbiota composition in greater resolution, sometimes at species-level, using proteomics, metabolomics, and deep sequencing. This has enhanced our understanding of how specific microbiota members influence acquisition or clinical manifestation of STI pathogen infection. Other advances include a steady, though still slow, increase in the number of studies that sought to determine the genital (penile or urethral) microbiota of males and how it may impact that of their female partners' genital microbiota and risk of STI acquisition. Altogether, these data enabled us to explore the concept that genital microbiota may be sexually transmitted and influence pathogenesis and clinical presentation of other STI. SUMMARY: With STI infection rates increasing worldwide, it is important now more than ever to find novel STI prevention strategies. Understanding if and how the genital microbiota is a modifiable risk factor for STI transmission, acquisition, and clinical manifestation may prove to be an important strategy in our efforts to curb morbidity in at risk populations.

Nonlytic cellular release of hepatitis A virus requires dual capsid recruitment of the ESCRT-associated Bro1 domain proteins HD-PTP and ALIX.

Although picornaviruses are conventionally considered 'nonenveloped', members of multiple picornaviral genera are released nonlytically from infected cells in extracellular vesicles. The mechanisms underlying this process are poorly understood. Here, we describe interactions of the hepatitis A virus (HAV) capsid with components of host endosomal sorting complexes required for transport (ESCRT) that play an essential role in release. We show release of quasi-enveloped virus (eHAV) in exosome-like vesicles requires a conserved export signal located within the 8 kDa C-terminal VP1 pX extension that functions in a manner analogous to late domains of canonical enveloped viruses. Fusing pX to a self-assembling engineered protein nanocage (EPN-pX) resulted in its ESCRT-dependent release in extracellular vesicles. Mutational analysis identified a 24 amino acid peptide sequence located within the center of pX that was both necessary and sufficient for nanocage release. Deleting a YxxL motif within this sequence ablated eHAV release, resulting in virus accumulating intracellularly. The pX export signal is conserved in non-human hepatoviruses from a wide range of mammalian species, and functional in pX sequences from bat hepatoviruses when fused to the nanocage protein, suggesting these viruses are released as quasi-enveloped virions. Quantitative proteomics identified multiple ESCRT-related proteins associating with EPN-pX, including ALG2-interacting protein X (ALIX), and its paralog, tyrosine-protein phosphatase non-receptor type 23 (HD-PTP), a second Bro1 domain protein linked to sorting of ubiquitylated cargo into multivesicular endosomes. RNAi-mediated depletion of either Bro1 domain protein impeded eHAV release. Super-resolution fluorescence microscopy demonstrated colocalization of viral capsids with endogenous ALIX and HD-PTP. Co-immunoprecipitation assays using biotin-tagged peptides and recombinant proteins revealed pX interacts directly through the export signal with N-terminal Bro1 domains of both HD-PTP and ALIX. Our study identifies an exceptionally potent viral export signal mediating extracellular release of virus-sized protein assemblies and shows release requires non-redundant activities of both HD-PTP and ALIX.

Experimental Urethral Infection with Neisseria gonorrhoeae.

Gonorrhea rates and antibiotic resistance are both increasing. Neisseria gonorrhoeae (Ng) is an exclusively human pathogen and is exquisitely adapted to its natural host. Ng can subvert immune responses and undergoes frequent antigenic variation, resulting in limited immunity and protection from reinfection. Previous gonococcal vaccine efforts have been largely unsuccessful, and the last vaccine to be tested in humans was more than 35 years ago. Advancing technologies and the threat of untreatable gonorrhea have fueled renewed pursuit of a vaccine as a long-term sustainable solution for gonorrhea control. Despite the development of a female mouse model of genital gonococcal infection two decades ago, correlates of immunity or protection remain largely unknown, making the gonococcus a challenging vaccine target. The controlled human urethral infection model of gonorrhea (Ng CHIM) has been used to study gonococcal pathogenesis and the basis of anti-gonococcal immunity. Over 200 participants have been inoculated without serious adverse events. The Ng CHIM replicates the early natural course of urethral infection. We are now at an inflexion point to pivot the use of the model for vaccine testing to address the urgency of improved gonorrhea control. Herein we discuss the need for gonorrhea vaccines, and the advantages and limitations of the Ng CHIM in accelerating the development of gonorrhea vaccines.

Cervicovaginal Microbiota Predicts Neisseria gonorrhoeae Clinical Presentation.

Neisseria gonorrhoeae infection of the female lower genital tract can present with a spectrum of phenotypes ranging from asymptomatic carriage to symptomatic cervical inflammation, or cervicitis. The factors that contribute to the development of asymptomatic or symptomatic infections are largely uncharacterized. We conducted a pilot study to assess differences in the cervicovaginal microbial community of patients presenting with symptomatic vs. asymptomatic N. gonorrhoeae infections to a sexually transmitted infections (STI) clinic. DNA was isolated from cervicovaginal swab specimens from women who tested positive for N. gonorrhoeae infection using a clinical diagnostic nucleic acid amplification test. We performed deep sequencing of 16S ribosomal RNA gene amplicons, followed by microbiome analyses with QIIME, and species-specific real-time PCR to assess the composition of microbial communities cohabitating the lower genital tract with the infecting N. gonorrhoeae. Specimens collected from asymptomatic individuals with N. gonorrhoeae infection and no co-infection with Chlamydia trachomatis and/or Trichomonas vaginalis carried Lactobacillus-dominant microbial communities more frequently than symptomatic patients without co-infection. When compared to asymptomatic individuals, symptomatic women had microbial communities characterized by more diverse and heterogenous bacterial taxa, typically associated with bacterial vaginosis (BV) [Prevotella, Sneathia, Mycoplasma hominis, and Bacterial Vaginosis-Associated Bacterium-1 (BVAB1)/"Candidatus Lachnocurva vaginae"]. Both symptomatic and asymptomatic N. gonorrhoeae patients with additional STI co-infection displayed a BV-like microbial community. These findings suggest that Lactobacillus-dominant vaginal microbial community may protect individuals from developing symptoms during lower genital tract infection with N. gonorrhoeae.

Nonviral sexually transmitted infections in pregnancy: current controversies and new challenges.

PURPOSE OF REVIEW: This review provides an update of nonviral, curable sexually transmitted infections (STIs) in pregnancy and summarizes our understanding of the current issues and controversies surrounding risk factors, screening, and treatment of STIs in pregnancy primarily in high-income countries (using the United States and the United Kingdom as examples). The infections covered in this review are syphilis, gonorrhea, chlamydia, trichomoniasis, and Mycoplasma genitalium infections. RECENT FINDINGS: Overall, limited modern data is available to update researchers and clinicians on the epidemiology and care of STIs in pregnancy. Though common risk factors can be identified among these STIs, like socioeconomic status and inadequate antenatal care, specific screening and treatment challenges vary by geography and pathogen. Wherever available, surveillance data and research evidence are often limited to nonpregnant patients, leading to imperfect pregnancy-specific risk estimates and obstetric lags in the development and adoption of new guidelines. We have identified three areas of opportunity that may enhance the effectiveness of current approaches and inform new ones: improved data collection and evidence-based screening practices; prompt and comprehensive therapy, including partner services, and evaluations of new treatment modalities; and equitable antenatal and sexual healthcare for all pregnant persons and their partners. SUMMARY: These findings highlight the need to revisit standards of screening and management of STIs in pregnancy in high-income countries.

The serogroup B meningococcal outer membrane vesicle-based vaccine 4CMenB induces cross-species protection against Neisseria gonorrhoeae.

There is a pressing need for a gonorrhea vaccine due to the high disease burden associated with gonococcal infections globally and the rapid evolution of antibiotic resistance in Neisseria gonorrhoeae (Ng). Current gonorrhea vaccine research is in the stages of antigen discovery and the identification of protective immune responses, and no vaccine has been tested in clinical trials in over 30 years. Recently, however, it was reported in a retrospective case-control study that vaccination of humans with a serogroup B Neisseria meningitidis (Nm) outer membrane vesicle (OMV) vaccine (MeNZB) was associated with reduced rates of gonorrhea. Here we directly tested the hypothesis that Nm OMVs induce cross-protection against gonorrhea in a well-characterized female mouse model of Ng genital tract infection. We found that immunization with the licensed Nm OMV-based vaccine 4CMenB (Bexsero) significantly accelerated clearance and reduced the Ng bacterial burden compared to administration of alum or PBS. Serum IgG and vaginal IgA and IgG that cross-reacted with Ng OMVs were induced by 4CMenB vaccination by either the subcutaneous or intraperitoneal routes. Antibodies from vaccinated mice recognized several Ng surface proteins, including PilQ, BamA, MtrE, NHBA (known to be recognized by humans), PorB, and Opa. Immune sera from both mice and humans recognized Ng PilQ and several proteins of similar apparent molecular weight, but MtrE was only recognized by mouse serum. Pooled sera from 4CMenB-immunized mice showed a 4-fold increase in serum bactericidal50 titers against the challenge strain; in contrast, no significant difference in bactericidal activity was detected when sera from 4CMenB-immunized and unimmunized subjects were compared. Our findings directly support epidemiological evidence that Nm OMVs confer cross-species protection against gonorrhea, and implicate several Ng surface antigens as potentially protective targets. Additionally, this study further defines the usefulness of murine infection model as a relevant experimental system for gonorrhea vaccine development.

Experimental Human Infection with Neisseria gonorrhoeae.

Experimental infection of male volunteers with Neisseria gonorrhoeae is safe and reproduces the clinical features of naturally acquired gonococcal urethritis. The human model is useful for testing the importance of putative gonococcal virulence factors for urethral infection in men and the model presents opportunities to examine host immune responses that may be exploited or improved in development and testing of gonococcal vaccines. In this chapter, we describe methods for production, characterization, and storage of N. gonorrhoeae stocks for experimental human challenge, preparation and delivery of inoculum suspensions, monitoring experimental infection, and statistical considerations for data analysis.

Viral DNA Binding to NLRC3, an Inhibitory Nucleic Acid Sensor, Unleashes STING, a Cyclic Dinucleotide Receptor that Activates Type I Interferon.

Immune suppression is a crucial component of immunoregulation and a subgroup of nucleotide-binding domain (NBD), leucine-rich repeat (LRR)-containing proteins (NLRs) attenuate innate immunity. How this inhibitory function is controlled is unknown. A key question is whether microbial ligands can regulate this inhibition. NLRC3 is a negative regulator that attenuates type I interferon (IFN-I) response by sequestering and attenuating stimulator of interferon genes (STING) activation. Here, we report that NLRC3 binds viral DNA and other nucleic acids through its LRR domain. DNA binding to NLRC3 increases its ATPase activity, and ATP-binding by NLRC3 diminishes its interaction with STING, thus licensing an IFN-I response. This work uncovers a mechanism wherein viral nucleic acid binding releases an inhibitory innate receptor from its target.

RIG-I Detects Kaposi's Sarcoma-Associated Herpesvirus Transcripts in a RNA Polymerase III-Independent Manner.

Retinoic acid-inducible gene I (RIG-I) is a cytosolic pathogen recognition receptor that initiates the innate immune response against many RNA viruses. We previously showed that RIG-I restricts Kaposi's sarcoma-associated herpesvirus (KSHV) reactivation (J. A. West et al., J Virol 88:5778-5787, 2014, https://doi.org/10.1128/JVI.03226-13). In this study, we report that KSHV stimulates the RIG-I signaling pathway in a RNA polymerase (Pol) III-independent manner and subsequently induces type I interferon (IFN) responses. Knockdown or inhibition of RNA Pol III had no effect on beta interferon (IFN-ß) induction by KSHV. By using high-throughput sequencing of RNA isolated by cross-linking immunoprecipitation (HITS-CLIP) approach, we identified multiple KSHV regions that give rise to RNA fragments binding to RIG-I, such as ORF810420-10496, Repeat region (LIR1)119059-119204, and ORF2543561-43650 The sequence dissimilarity between these fragments suggests that RIG-I detects a particular structure rather than a specific sequence motif. Synthesized ORF810420-10496 RNA stimulated RIG-I-dependent but RNA Pol III-independent IFN-ß signaling. In summary, several KSHV RNAs are sensed by RIG-I in a RNA Pol III-independent manner.IMPORTANCE Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of Kaposi's sarcoma, primary effusion lymphoma, and multicentric Castleman's disease. Innate immune responses against viral infections, especially the induction of type I interferon, are critical for limiting the replication of viruses. Retinoic acid-inducible gene I (RIG-I), a cytosolic RNA helicase sensor, plays a significant role in the induction of type I interferon responses following viral infection. Here, we identified multiple RNA regions in KSHV as potential virus ligands that bind to RIG-I and stimulate RIG-I-dependent but RNA Pol III-independent IFN-ß signaling. Our results expand the role of RIG-I by providing an example of a DNA virus activating a canonical RNA-sensing pathway.

Properly folded and functional PorB from Neisseria gonorrhoeae inhibits dendritic cell stimulation of CD4+ T cell proliferation.

Neisseria gonorrhoeae is an exclusive human pathogen that evades the host immune system through multiple mechanisms. We have shown that N. gonorrhoeae suppresses the capacity of antigen-presenting cells to induce CD4+ T cell proliferation. In this study, we sought to determine the gonococcal factors involved in this adaptive immune suppression. We show that suppression of the capacity of antigen-pulsed dendritic cells to induce T cell proliferation is recapitulated by administration of a high-molecular-weight fraction of conditioned medium from N. gonorrhoeae cultures, which includes outer membrane vesicles that are shed during growth of the bacteria. N. gonorrhoeae PorB is the most abundant protein in N. gonorrhoeae-derived vesicles, and treatment of dendritic cells with purified recombinant PorB inhibited the capacity of the cells to stimulate T cell proliferation. This immunosuppressive feature of purified PorB depended on proper folding of the protein. PorB from N. gonorrhoeae, as well as other Neisseria species and other Gram-negative bacterial species, are known to activate host Toll-like receptor 2 (TLR2) signaling. Published studies have demonstrated that purified Neisseria PorB forms proteinacious nanoparticles, termed proteosomes, when detergent micelles are removed. Unlike folded, detergent-solubilized PorB, PorB proteosomes stimulate immune responses. We now demonstrate that the formation of PorB proteosomes from structurally intact PorB eliminates the immunosuppressive property of the protein while enhancing TLR2 stimulation. These findings suggest that gonococcal PorB present in shed outer membrane vesicles plays a role in suppression of adaptive immune responses to this immune-evasive pathogen.

Novel aspects of the assembly and activation of inflammasomes with focus on the NLRC4 inflammasome.

Inflammasomes are multiprotein structures that activate caspase-1, support secretion of pro-inflammatory cytokines, IL-1ß and IL-18, and also induce inflammatory programmed cell death, termed pyoptosis. Inflammasomes are activated in response to the detection of endogenous and microbially derived danger signals and are mediated by several classes of inflammasome-forming sensors. These include several nucleotide-binding proteins of the NOD-like receptor (NLR) family, including NLRP1, NLRP3 and NLRC4, as well as the proteins Absent in Melanoma 2 (AIM2) and Pyrin. Mutations in genes encoding some of these sensors have been found to be associated with gain-of-function monogenetic inflammatory disorders in humans. Genetic, biochemical and structural studies have begun to demonstrate how these proteins sense danger signals and to shed light on the step-by-step processes that are necessary for the assembly of inflammasomes, in both physiologic responses to pathogens and potentially in autoinflammatory conditions. Recent biochemical studies of pro-caspase-1 and an adapter protein known as ASC suggest that inflammasomes act to initiate self-generating effector filaments responsible for activating caspase-1 and initiating downstream signaling. These studies have suggested a model of molecular events from sensor activation to inflammasome formation that may describe processes that are universal to inflammasome formation.

Human Immune Responses and the Natural History of Neisseria gonorrhoeae Infection.

The intimate relationship between humans and Neisseria gonorrhoeae infections span centuries, which is evidenced in case reports from studies dating back to the late 1700s and historical references that predate medical literature. N. gonorrhoeae is an exclusive human pathogen that infects the genital tract of both men and women as well as other mucosal surfaces including the oropharynx and rectum. In symptomatic infections, N. gonorrhoeae induces a robust inflammatory response at the site of infection. However, infections can also present asymptomatically complicating efforts to reduce transmission. N. gonorrhoeae infections have been effectively treated with antibiotics since their use was introduced in humans. Despite the existence of effective antibiotic treatments, N. gonorrhoeae remains one of the most common sexually transmitted pathogens and antibiotic resistant strains have arisen that limit treatment options. Development of a vaccine to prevent infection is considered a critical element of controlling this pathogen. The efforts to generate an effective gonococcal vaccine is limited by our poor understanding of the natural immunologic responses to infection. It is largely accepted that natural protective immunity to N. gonorrhoeae infections in humans does not occur or is very rare. Previous studies of the natural history of infection as well as some of the humoral and cellular immune responses to infection offer a window into the issues surrounding N. gonorrhoeae vaccine development. In this review, we summarize the current body of knowledge pertaining to human immune responses to gonococcal infections and the role of these responses in mediating protection from N. gonorrhoeae.

The NLRC4 Inflammasome.

15 years ago, the fundamental biology of an inflammatory signaling complex eventually dubbed "the inflammasome" began to unravel in chronologic parallel with the discovery that many inflammatory diseases were associated with its hyperactivity. Though the genetic origins of Familial Mediterranean Fever (FMF, caused my mutations in MEFV) were discovered first, it would take nearly two decades before the mechanistic connections to a PYRIN inflammasome were made. In the interim, the intensive study of the NLRP3 inflammasome, and the diseases associated with its hyperactivation, have largely dictated the paradigm of inflammasome composition and function. Despite impressive gains, focusing on NLRP3 left gaps in our understanding of inflammasome biology. Foremost among these gaps were how inflammasomes become activated and the connections between inflammasome structure and function. Fortunately, work in another inflammasome inducer, NLRC4, grew to fill those gaps. The current understanding of the NLRC4 inflammasome is perhaps the most comprehensive illustration of the inflammasome paradigm: trigger (e.g. cytosolic flagellin), sensor (NAIP), nucleator (NLRC4), adaptor (ASC), and effector (CASP1). Detailed work has also identified observations that challenge this paradigm. Simultaneously, the features unique to each inflammasome offer a lesson in contrast, providing perspectives on inflammasome activation, regulation, and function. In this review, we endeavor to highlight recent breakthroughs related to NLRC4 inflammasome structure and activation, important in vivo work in infection and systemic inflammation, and the characterization of a spectrum of human NLRC4-associated autoinflammatory diseases.

NLRX1 promotes immediate IRF1-directed antiviral responses by limiting dsRNA-activated translational inhibition mediated by PKR.

NLRX1 is unique among the nucleotide-binding-domain and leucine-rich-repeat (NLR) proteins in its mitochondrial localization and ability to negatively regulate antiviral innate immunity dependent on the adaptors MAVS and STING. However, some studies have suggested a positive regulatory role for NLRX1 in inducing antiviral responses. We found that NLRX1 exerted opposing regulatory effects on viral activation of the transcription factors IRF1 and IRF3, which might potentially explain such contradictory results. Whereas NLRX1 suppressed MAVS-mediated activation of IRF3, it conversely facilitated virus-induced increases in IRF1 expression and thereby enhanced control of viral infection. NLRX1 had a minimal effect on the transcription of IRF1 mediated by the transcription factor NF-kB and regulated the abundance of IRF1 post-transcriptionally by preventing translational shutdown mediated by the double-stranded RNA (dsRNA)-activated kinase PKR and thereby allowed virus-induced increases in the abundance of IRF1 protein.

Neisseria gonorrhoeae Lytic Transglycosylases LtgA and LtgD Reduce Host Innate Immune Signaling through TLR2 and NOD2.

Neisseria gonorrhoeae releases anhydro peptidoglycan monomers during growth through the action of two lytic transglycosylases encoded in the N. gonorrhoeae genome, LtgA and LtgD. Because peptidoglycan and peptidoglycan components activate innate immune signaling, we hypothesized that the activity of LtgA and LtgD would influence the host responses to gonococcal infection. N. gonorrhoeae lacking LtgA and LtgD caused increased host production of inflammatory cytokines IL-1ß and TNF-α. Culture supernatants from ΔltgA/ΔltgD N. gonorrhoeae contain more shed outer membrane-associated proteins and multimeric peptidoglycan fragments rather than monomers. These culture supernatants were more potent activators of host TLR2 and NOD2 signaling when compared to supernatants from the isogenic parental N. gonorrhoeae strain. Purified peptidoglycan monomers containing anhydro muramic acid produced by LtgA were poor stimulators of NOD2, whereas peptidoglycan monomers containing reducing muramic acid produced by host lysozyme were potent stimulators of NOD2. These data indicate that LtgA and LtgD reduce recognition of N. gonorrhoeae by TLR2 and NOD2.

Inflammasome Activation Can Mediate Tissue-Specific Pathogenesis or Protection in Staphylococcus aureus Infection.

Staphylococcus aureus is a Gram-positive coccus that interacts with human hosts on a spectrum from quiet commensal to deadly pathogen. S. aureus is capable of infecting nearly every tissue in the body resulting in cellulitis, pneumonia, osteomyelitis, endocarditis, brain abscesses, bacteremia, and more. S. aureus has a wide range of factors that promote infection, and each site of infection triggers a different response in the human host. In particular, the different patterns of inflammasome activation mediate tissue-specific pathogenesis or protection in S. aureus infection. Although still a nascent field, understanding the unique host-pathogen interactions in each infection and the role of inflammasomes in mediating pathogenesis may lead to novel strategies for treating S. aureus infections. Reviews addressing S. aureus virulence and pathogenesis (Thammavongsa et al. 2015), as well as epidemiology and pathophysiology (Tong et al. 2015), have recently been published. This review will focus on S. aureus factors that activate inflammasomes and their impact on innate immune signaling and bacterial survival.

ADAM10 Cell Surface Expression but Not Activity Is Critical for Staphylococcus aureus α-Hemolysin-Mediated Activation of the NLRP3 Inflammasome in Human Monocytes.

The Staphylococcus aureus toxin, α-hemolysin, is an important and well-studied virulence factor in staphylococcal infection. It is a soluble monomeric protein that, once secreted by the bacterium, forms a heptameric pore in the membrane of a broad range of host cell types. Hemolysin was recently discovered to bind and activate a disintegrin and metalloprotease 10 (ADAM10). In epithelial and endothelial cells, ADAM10 activation is required for the toxin's activity against these cells. In host monocytic cells, α-hemolysin activates the nucleotide-binding domain and leucine-rich repeat containing gene family, pyrin domain containing 3 (NLRP3) inflammasome leading to production of pro-inflammatory cytokines and cell death. We now show that ADAM10 is critical for α-hemolysin-mediated activation of the NLRP3 inflammasome in human monocytes as siRNA knockdown or chemical blockade of ADAM10-α-hemolysin interaction leads to diminished inflammasome activation and cell death by reducing the available ADAM10 on the cell surface. Unlike epithelial cell and endothelial cell damage, which requires α-hemolysin induced ADAM10 activation, ADAM10 protease activity was not required for NLRP3 inflammasome activation. This work confirms the importance of ADAM10 in immune activation by α-hemolysin, but indicates that host cell signal induction by the toxin is different between host cell types.