IL-17C is a driver of damaging inflammation during Neisseria gonorrhoeae infection of human Fallopian tube.

The human pathogen Neisseria gonorrhoeae ascends into the upper female reproductive tract to cause damaging inflammation within the Fallopian tubes and pelvic inflammatory disease (PID), increasing the risk of infertility and ectopic pregnancy. The loss of ciliated cells from the epithelium is thought to be both a consequence of inflammation and a cause of adverse sequelae. However, the links between infection, inflammation, and ciliated cell extrusion remain unresolved. With the use of ex vivo cultures of human Fallopian tube paired with RNA sequencing we defined the tissue response to gonococcal challenge, identifying cytokine, chemokine, cell adhesion, and apoptosis related transcripts not previously recognized as potentiators of gonococcal PID. Unexpectedly, IL-17C was one of the most highly induced genes. Yet, this cytokine has no previous association with gonococcal infection nor pelvic inflammatory disease and thus it was selected for further characterization. We show that human Fallopian tubes express the IL-17C receptor on the epithelial surface and that treatment with purified IL-17C induces pro-inflammatory cytokine secretion in addition to sloughing of the epithelium and generalized tissue damage. These results demonstrate a previously unrecognized but critical role of IL-17C in the damaging inflammation induced by gonococci in a human explant model of PID.

Neisseria gonorrhoeae infection in female sex workers in an STI clinic in Nairobi, Kenya.

BACKGROUND: Gonorrhea caused by Neisseria gonorrhoeae is the second most prevalent curable sexually transmitted infection worldwide. Female Sex Workers (FSWs) are at a higher risk of contracting gonorrhea due to their risky sexual behaviors like inconsistent condom use and multiple sexual partners. We determined the prevalence and risk factors associated with gonorrhea and its antimicrobial susceptibility pattern among symptomatic FSWs attending Sexual Workers Outreach Program (SWOP) city clinic in Nairobi, Kenya. METHODS: Using convenience sampling, we recruited 379 female sex workers from SWOP City clinic in Nairobi County. We administered a semi-structured questionnaire to collect data on socio-demographics and behavioral risk factors associated with gonorrhea. We also conducted three focus groups. Two endocervical swabs were collected from each participant by the attending physician for the laboratory identification of Neisseria gonorrhoeae. An antimicrobial susceptibility test was performed using the disc diffusion method. RESULTS: Twenty-four out of 379 (6.3%) participants tested positive for gonorrhea by PCR. The significant risk factors associated with gonorrhea were having multiple sexual partners in the previous two weeks, primary education, and being in the age group of 38-49 years (p < 0.05). From the qualitative data, sex work disclosure, and difficulty in engaging protected sex with their partner, and unprotected sex with their clients due to more money from the client, PREP, and alcohol use made the female sex workers vulnerable to gonorrhea exposure and or risky sexual behavior. The culture-positive sample result yielded complete (100%) resistance to all the antimicrobials used. CONCLUSION: Neisseria gonorrhoeae infection is prevalent among symptomatic FSWs in Nairobi. Multiple sexual partners, being in age group 38-49 years and having primary education were the factors associated with gonorrhea among the study participants. Based on our identification of a highly resistant isolate, we strongly recommend increasing capacity for culture-based diagnosis and susceptibility testing.

Úlcera genital imitadora / Misleading Genital Ulcer

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Inhibition of TLR2/TLR4 alleviates the Neisseria gonorrhoeae infection damage in human endometrial epithelial cells via Nrf2 and NF-Kßsignaling.

BACKGROUND: Neisseria gonorrhoeae (N.g) is Gram-negative bacteria and can lead to endometritis in female. Toll-like receptors regulate immune response in various diseases. However, the roles of TLR2 and TLR4 in. Neisseria gonorrhoeae-induced infection damage in human endometrial epithelia were investigated. METHODS: hEECs were infected with N.g (MOI 10 and 100) and cell viability and apoptosis were measured by CCK8 and flow cytometry assays in both infected groups with the uninfected normal hEECs as negative control. TLR2/TLR4 proteins were measured by ELISA method. Pro-inflammatory markers NLRP3, PGES (PGE2) and TNF-α were assessed by RT-qPCR (mRNA expression) and Elisa (protein concentrations). Transfection assays were performed to up- or down- regulate expression of TLR2 and TLR4 so as to study the functions of TLR2/TLR4 in. N.g-infected hEECs, followed by apoptosis and inflammation assessment. Similarly, we explored the interactions between TLR2/TLR4 and Nrf2/NF-κB/p65 by knocking down TLR2/TLR4 to detect the signaling and further regulating the signaling to evaluate TLR2/ TLR4, apoptosis and inflammation in cells. RESULTS: N.g suppressed cell viabilities and induced cell apoptosis and inflammation. TLR2/TLR4 downregulation inhibited the infection damage. Nrf2 was activated while NF-κB/p65 was depleted as TLR2/ TLR4 was knocked down. Activation of Nrf2 and inhibition of NF-κB resulted in decrease of TLR2/TLR4, which could retard apoptosis and inflammation induced by N.g infection. CONCLUSION: TLR2/TLR4 depletion could alleviate the N.g-infected hEECs via Nrf2/NF-kB signaling, suggesting that TLR2/TLR4 inhibitors might serve as a treatment to reduce N.g infection in human endometrial epithelia.

Modulation of phagocytosis-induced cell death of human neutrophils by Neisseria gonorrhoeae.

Optimal innate immune response to infection includes eradication of potential pathogens, resolution of associated inflammation, and restitution of homeostasis. Phagocytosing human polymorphonuclear leukocytes (hPMN) undergo accelerated apoptosis, a process referred to as phagocytosis-induced cell death (PICD) and an early step in their clearance from inflammatory sites. Among human pathogens that modulate hPMN apoptosis, Neisseria gonorrhoeae delays PICD, which may contribute to the exuberant neutrophilic inflammation that characterizes gonorrhea. To elucidate the mechanisms underlying delayed PICD, we compared features of hPMN cell death that followed phagocytosis of N. gonorrhoeae FA1090 wild-type (GC) or serum-opsonized zymosan (OPZ), a prototypical stimulus of PICD. Phosphatidylserine externalization required NADPH oxidase activity after ingestion of GC or OPZ, and annexin V staining and DNA fragmentation were less after phagocytosis of GC compared to OPZ. Caspase 3/7 and caspase 9 activities after phagocytosis of GC were less than that seen after ingestion of OPZ, but caspase 8 activity was the same after ingestion of GC or OPZ. When hPMN sequentially ingested GC followed by OPZ, both caspase 3/7 and 9 activities were less than that seen after OPZ alone, and the inhibition was dose dependent for GC, suggesting that ingestion of GC actively inhibited PICD. Sequential phagocytosis did not block caspase 8 activity, mitochondrial depolarization, or annexin V/propidium iodide staining compared to responses of hPMN fed OPZ alone, despite inhibition of caspases 3/7 and 9. Taken together, these data suggest that active inhibition of the intrinsic pathway of apoptosis contributes to the delay in PICD after hPMN ingestion of N. gonorrhoeae.

Clinical presentation of asymptomatic and symptomatic women who tested positive for genital gonorrhoea at a sexual health service in Melbourne, Australia.

Gonorrhoea cases in women have been rising in Australia in the 2010s but the cause of the increase is not well understood. This cross-sectional study aimed to describe the characteristics of genital gonorrhoea infection in women attending the Melbourne Sexual Health Centre, Australia. Gonorrhoea cases were diagnosed by nucleic acid amplification test (NAAT) and/or culture. Genitourinary specimens were obtained in 12 869 clinic visits in women aged 16 years or above between August 2017 and August 2018. Genital gonorrhoea was detected in 142 (1.1%) of the visits. Almost half of the cases were asymptomatic, 47.9% [95% confidence interval (CI) 39.8-56.1%]; yellow, green or pus-like vaginal discharge was present in 11.3% (95% CI 7.0-17.6%) and other genital symptoms in 40.8% (95% CI 33.1-49.1%) of the cases. The mean time between last sexual contact and onset of symptoms was 7.3 days and between the onset of symptoms to presentation to the clinic was 12.1 days. Half of the cases of genital gonorrhoea among women are asymptomatic and these cases would have been missed by testing of only symptomatic women. Further epidemiological and behavioural research is required to understand the temporal changes in sexual practices among women in Australia.

Antimicrobial susceptibility profile of Gonococcal isolates obtained from men presenting with urethral discharge in Addis Ababa, Ethiopia: Implications for national syndromic treatment guideline.

BACKGROUND: Neisseria gonorrhoeae (gonococcus) is the etiologic agent for the sexually transmitted Infection gonorrhea, a disease with a significant global public health impact. The treatment regimen for gonorrhea has been changed frequently over the past few decades due to the organism's propensity for developing antibiotic resistance. This study investigated antimicrobial susceptibility patterns of quinolones, third-generation cephalosporin, and other relevant antimicrobials found in N. gonorrhoeae isolated from men presenting with urethral discharge at selected healthcare facilities in Addis Ababa, Ethiopia, with the aim of revising the national treatment regimen based on the information generated from this study. METHODS: A total of 599 male patients presenting with urethral discharge were included in the current study. Urethral discharge specimens were cultured on Modified Thayer Martín media and suspected gonococcal colonies were confirmed using Oxidase and Superoxol tests followed by identification through a commercial kit (API-NHR). Antimicrobial susceptibility testing was performed by the Kirby-Bauer disc diffusion method using ciprofloxacin (5µg), ceftriaxone (30µg), cefixime (5µg), cefoxitin (30 µg), penicillin (10µg) and spectinomycin (100 µg) on enriched GC agar. Minimum Inhibitory Concentration (MIC) was also carried out using concentration gradient strips (E-tests) of the same antimicrobial agents. RESULTS: The prevalence of gonococcal isolates in the current study was 69%. Out of the 361 gonococcal isolates, close to 68% were fluoroquinolone non-susceptible, with 60% resistant and 7% having an intermediate status. However, all tested isolates were susceptible to ceftriaxone. In addition, all of the isolates have shown reduced non-susceptibility to spectinomycin and cefoxitin. CONCLUSION: The prevalence of gonococcal isolates in men presenting with urethral discharge at selected healthcare facilities in Addis Ababa, Ethiopia was found to be high. The high level of fluoroquinolone resistance observed in gonococcal isolates recovered in this study necessitates revision of the national syndromic treatment guideline.

Complement interactions with the pathogenic Neisseriae: clinical features, deficiency states, and evasion mechanisms.

Neisseria gonorrhoeae causes the sexually transmitted infection gonorrhea, while Neisseria meningitidis is an important cause of bacterial meningitis and sepsis. Complement is a central arm of innate immune defenses and plays an important role in combating Neisserial infections. Persons with congenital and acquired defects in complement are at a significantly higher risk for invasive Neisserial infections such as invasive meningococcal disease and disseminated gonococcal infection compared to the general population. Of note, Neisseria gonorrhoeae and Neisseria meningitidis can only infect humans, which in part may be related to their ability to evade only human complement. This review summarizes the epidemiologic and clinical aspects of Neisserial infections in persons with defects in the complement system. Mechanisms used by these pathogens to subvert killing by complement and preclinical studies showing how these complement evasion strategies may be used to counteract the global threat of meningococcal and gonococcal infections are discussed.

Factors associated with time to presentation for individuals with symptomatic uncomplicated genital gonorrhoea: a cross sectional cohort study of GToG trial participants.

OBJECTIVES: To determine the variation in the time from onset of symptoms to clinical presentation (time to presentation [TTP]) in a cohort of sexual health attendees with symptomatic uncomplicated genital gonorrhoea and to identify factors associated with TTP. METHODS: Participants were recruited from 14 clinics across England into the 'Gentamicin for the Treatment of Gonorrhoea (GToG)' trial between October 2014 and November 2016. Multivariable analysis was performed using prospectively collected demographic, behavioural and clinical data in a subset of the GToG study cohort presenting with genital discharge and/or dysuria who tested positive for Neisseria gonorrhoeae using a nucleic acid amplification test. The results were expressed as geometric mean ratios (GMR) with 95% CI for time to presentation after onset of symptoms. RESULTS: 316 participants (269 men and 47 women) with a median age of 27.6 years (IQR 23.0-34.8) were included. 194 (61%) were Caucasian, 29 (9%) Black African, 27 (9%) Asian and 66 (21%) of other ethnicities. Median TTP was 3 days for men (IQR 2-7) and 14 days for women (IQR 7-21). Participants reported genital discharge (297/316 [94%]), dysuria (251/316 [79%]), both genital discharge and dysuria (232/316 [73%]) and other concurrent symptoms 76/316 (24%) (e.g., rectal bleeding or genital itching). 45/316 (14%) participants reported sexual contact while symptomatic, of whom TTP was more than 7 days in 32/45 (71%). A longer TTP was associated with gender (female cf. male, GMR 2.34 [1.67 to 3.26]), no prior history of gonorrhoea (GMR 1.46 [1.15 to 1.86]), 'regular' or 'ex-regular' sexual relationship (regular cf. one off GMR 1.35 [1.05 to 1.72]); ex-regular cf. one off GMR 1.88 [1.12 to 3.14]), and being heterosexual (GMR 1.69 [1.31 to 2.19]). CONCLUSION: Specific demographic and behavioural factors are associated with a longer TTP in individuals with symptomatic genital gonorrhoea. Detailed knowledge of these factors can be used to prioritise and optimise gonorrhoea management and prevention.

Mechanistic Insight Into the Activation of the NLRP3 Inflammasome by Neisseria gonorrhoeae in Macrophages.

Gonorrhea is a type III legal communicable disease caused by Neisseria gonorrhoeae (NG), one of the most common sexually transmitted bacteria worldwide. NG infection can cause urethritis or systemic inflammation and may lead to infertility or other complications. The NACHT, LRR, and PYD domains-containing protein 3 (NLRP3) inflammasome is a protein complex composed of NLRP3, apoptosis-associated speck-like protein and caspase-1 and is an important part of the cellular machinery controlling the release of interleukin (IL)-1ß and IL-18 and the pathogenesis of numerous infectious diseases. It has been reported that NG infection activates the NLRP3 inflammasome; however, the underlying mechanism remain unclear. In this report, the signaling pathways involved in the regulation of NG-mediated NLRP3 inflammasome activation in macrophages were studied. The results indicated that viable NG, but not heat-killed or freeze/thaw-killed NG, activated the NLRP3 inflammasome in macrophages through toll-like receptor 2, but not toll-like receptor 4. NG infection provided the priming signal to the NLRP3 inflammasome that induced the expression of NLRP3 and IL-1ß precursor through the nuclear factor kappa B and mitogen-activated protein kinase pathways. In addition, NG infection provided the activation signal to the NLRP3 inflammasome that activated caspase-1 through P2X7 receptor-dependent potassium efflux, lysosomal acidification, mitochondrial dysfunction, and reactive oxygen species production pathways. Furthermore, we demonstrated that NLRP3 knockout increased phagocytosis of bacteria by macrophages and increases the bactericidal activity of macrophages against NG. These findings provide potential molecular targets for the development of anti-inflammatory drugs that could ameliorate NG-mediated inflammation.

The novel interaction between Neisseria gonorrhoeae TdfJ and human S100A7 allows gonococci to subvert host zinc restriction.

Neisseria gonorrhoeae causes the sexually-transmitted infection gonorrhea, a global disease that is difficult to treat and for which there is no vaccine. This pathogen employs an arsenal of conserved outer membrane proteins called TonB-dependent transporters (TdTs) that allow the gonococcus to overcome nutritional immunity, the host strategy of sequestering essential nutrients away from invading bacteria to handicap infectious ability. N. gonorrhoeae produces eight known TdTs, of which four are utilized for acquisition of iron or iron chelates from host-derived proteins or xenosiderophores produced by other bacteria. Of the remaining TdTs, two of them, TdfH and TdfJ, facilitate zinc uptake. TdfH was recently shown to bind Calprotectin, a member of the S100 protein family, and subsequently extract its zinc, which is then internalized by N. gonorrhoeae. Like Calprotectin, other S100s are also capable of binding transition metals such as zinc and copper, and thus have demonstrated growth suppression of numerous other pathogens via metal sequestration. Considering the functional and structural similarities of the TdTs and of the S100s, as well as the upregulation in response to Zn limitation shown by TdfH and TdfJ, we sought to evaluate whether other S100s have the ability to support gonococcal growth by means of zinc acquisition and to frame this growth in the context of the TdTs. We found that both S100A7 and S10012 are utilized by N. gonorrhoeae as a zinc source in a mechanism that depends on the zinc transport system ZnuABC. Moreover, TdfJ binds directly to S100A7, from which it internalizes zinc. This interaction is restricted to the human version of S100A7, and zinc presence in S100A7 is required to fully support gonococcal growth. These studies highlight how gonococci co-opt human nutritional immunity, by presenting a novel interaction between TdfJ and human S100A7 for overcoming host zinc restriction.

Microscopic examination of Gram-stained smears for anogenital gonorrhoea in men who have sex with men is cost-effective: evidence from a modelling study.

OBJECTIVE: To assess the cost-effectiveness of three testing strategies with or without light microscopic Gram-stained smear (GSS) evaluation for the detection of anogenital gonorrhoea among men who have sex with men (MSM) at the Amsterdam STI clinic using a healthcare payer perspective. METHODS: Three testing strategies for MSM were compared: (1) GSS in symptomatic MSM only (currently practised strategy), (2) no GSS and (3) GSS in symptomatic and asymptomatic MSM. The three testing protocols include testing with nucleic acid amplification test to verify the GSS results in (1) and (3), or as the only test in (2). A transmission model was employed to calculate the influence of the testing strategies on the prevalence of anogenital gonorrhoea over 10 years. An economic model combined cost data on medical consultations, tests and treatment and utility data to estimate the number of epididymitis cases and quality-adjusted life years (QALY) associated with gonorrhoea. Incremental cost-effectiveness ratios (ICERs) for the testing scenarios were estimated. Uncertainty and sensitivity analyses were performed. RESULTS: No GSS testing compared with GSS in symptomatic MSM only (current strategy) resulted in nine extra epididymitis cases (95% uncertainty interval (UI): 2-22), 72 QALYs lost (95% UI: 59-187) and €7300 additional costs (95% UI: -€185 000 (i.e.cost-saving) to €407 000) over 10 years. GSS testing in both symptomatic and asymptomatic MSM compared with GSS in symptomatic MSM only resulted in one prevented epididymitis case (95% UI: 0-2), 1.1 QALY gained (95% UI: 0.1-3.3), €148 000 additional costs (95% UI: €86 000 to-€217 000) and an ICER of €177 000 (95% UI: €67 000-to €705 000) per QALY gained over 10 years. The results were robust in sensitivity analyses. CONCLUSIONS: GSS for symptomatic MSM only is cost-effective compared with no GSS for MSM and with GSS for both symptomatic and asymptomatic MSM.

Pathogenesis of Neisseria gonorrhoeae and the Host Defense in Ascending Infections of Human Fallopian Tube.

Neisseria gonorrhoeae is an obligate human pathogen that causes mucosal surface infections of male and female reproductive tracts, pharynx, rectum, and conjunctiva. Asymptomatic or unnoticed infections in the lower reproductive tract of women can lead to serious, long-term consequences if these infections ascend into the fallopian tube. The damage caused by gonococcal infection and the subsequent inflammatory response produce the condition known as pelvic inflammatory disease (PID). Infection can lead to tubal scarring, occlusion of the oviduct, and loss of critical ciliated cells. Consequences of the damage sustained on the fallopian tube epithelium include increased risk of ectopic pregnancy and tubal-factor infertility. Additionally, the resolution of infection can produce new adhesions between internal tissues, which can tear and reform, producing chronic pelvic pain. As a bacterium adapted to life in a human host, the gonococcus presents a challenge to the development of model systems for probing host-microbe interactions. Advances in small-animal models have yielded previously unattainable data on systemic immune responses, but the specificity of N. gonorrhoeae for many known (and unknown) host targets remains a constant hurdle. Infections of human volunteers are possible, though they present ethical and logistical challenges, and are necessarily limited to males due to the risk of severe complications in women. It is routine, however, that normal, healthy fallopian tubes are removed in the course of different gynecological surgeries (namely hysterectomy), making the very tissue most consequentially damaged during ascending gonococcal infection available for laboratory research. The study of fallopian tube organ cultures has allowed the opportunity to observe gonococcal biology and immune responses in a complex, multi-layered tissue from a natural host. Forty-five years since the first published example of human fallopian tube being infected ex vivo with N. gonorrhoeae, we review what modeling infections in human tissue explants has taught us about the gonococcus, what we have learned about the defenses mounted by the human host in the upper female reproductive tract, what other fields have taught us about ciliated and non-ciliated cell development, and ultimately offer suggestions regarding the next generation of model systems to help expand our ability to study gonococcal pathogenesis.

Microbiological Analysis from a Phase 2 Randomized Study in Adults Evaluating Single Oral Doses of Gepotidacin in the Treatment of Uncomplicated Urogenital Gonorrhea Caused by Neisseria gonorrhoeae.

We evaluated microbiological correlates for the successful treatment of Neisseria gonorrhoeae isolates from a phase 2 study of gepotidacin, a novel triazaacenaphthylene antibacterial, for therapy of uncomplicated urogenital gonorrhea. Culture, susceptibility testing, genotypic characterization, and frequency of resistance (FoR) were performed for selected isolates. Microbiological success was defined as culture-confirmed eradication of N. gonorrhoeae Against 69 baseline urogenital isolates, gepotidacin MICs ranged from ≤0.06 to 1 µg/ml (MIC90 = 0.5 µg/ml). For gepotidacin, the ratio of the area under the free-drug concentration-time curve to the MIC (fAUC/MIC) was associated with therapeutic success. Success was 100% (61/61) at fAUC/MICs of ≥48 and decreased to 63% (5/8) for fAUC/MICs of ≤25. All 3 isolates from microbiological failures were ciprofloxacin resistant, had a baseline gepotidacin MIC of 1 µg/ml, and carried a preexisting ParC D86N mutation, a critical residue for gepotidacin binding. In a test-of-cure analysis, the resistance to gepotidacin emerged in 2 isolates (MICs increased ≥32-fold) with additional GyrA A92T mutations, also implicated in gepotidacin binding. Test-of-cure isolates had the same sequence type as the corresponding baseline isolates. For 5 selected baseline isolates, all carrying a ParC D86N mutation, the in vitro FoR to gepotidacin was low (10-9 to 10-10); the resistant mutants had the same A92T mutation as the 2 isolates in which resistance emerged. Five participants with isolates harboring the ParC D86N mutation were treatment successes. In summary, fAUC/MICs of ≥48 predicted 100% microbiological success, including 3 isolates with the ParC D86N mutation (fAUC/MICs ≥ 97). Pharmacokinetic/pharmacodynamic determinations may help to evaluate new therapies for gonorrhea; further study of gepotidacin is warranted. (This study has been registered at ClinicalTrials.gov under identifier NCT02294682.).

Sexually transmitted rectal infections in a cohort of 'men having sex with men'.

PURPOSE: We assessed the prevalence and predictors of Chlamydia trachomatis, Neisseriagonorrhoeae and Mycoplasmagenitalium rectal infections in a population of 'men having sex with men' (MSM). METHODOLOGY: From January to November 2017, 165 MSM attending a STI outpatients clinic in Bologna (Italy) and reporting unsafe anal intercourses were enrolled. An ano-rectal swab was collected from each patient: chlamydial and gonococcal infections were diagnosed by a commercial NAAT, whereas an in-house quantitative PCR was used for M. genitalium detection. In addition, 131 urine samples and 84 pharyngeal swabs underwent testing for C. trachomatis and N. gonorrhoeae. A molecular C. trachomatis typing, a serological screening for anti-Chlamydia IgG and IgA, as well as the assessment of HIV, HCV and syphilis infections, were performed.Results/Key findings. The prevalence of C. trachomatis, N. gonorrhoeae and M. genitalium rectal infections was 27.2, 25.4 and 4.8 %, respectively. Globally, 63.1 % of cases were asymptomatic and up to 80 % of chlamydial and gonococcal infections would have been missed if the rectal site had not been tested. All the patients with rectal M. genitalium carriage were asymptomatic and characterized by low bacterial loads (<2500 DNA copies/reaction). Lymphogranuloma venereum (LGV) prevalence was 12.1 % with a considerable proportion of asymptomatic infections (35 %). The presence of symptoms, age >30, HIV-positivity and elevated levels of anti-Chlamydia antibodies were the most significant predictors of LGV. CONCLUSIONS: Sexually transmitted rectal infections are frequent and often asymptomatic among MSM. LGV prevalence is high in our country and there is increasing evidence of symptomless cases.