Purpose of Review: To emphasize key gaps in knowledge impacting efforts to control single infection and co-infections with Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG), the most common bacterial sexually transmitted infections (STIs) worldwide. Recent Findings: Clinical and epidemiological studies describe gaps in understanding about female rectal CT infection, screening effectiveness, pelvic inflammatory disease, and influence of the microbiome. For NG, gaps in knowledge include factors increasing incidence in men who have sex with men, correlations between treatment and antibiotic resistance, the role of pharyngeal infection, and microbiome influence. CT/NG co-infections are poorly understood, and adequate models to explore pathophysiological consequences of co-infection urgently needed. The sole existing CT/NG co-infection mouse model showed that CT/NG interactions in vivo modulate host response and NG load/shedding—encouraging further consideration of this model and potential alternatives. Summary: We stress key challenges in controlling these important STIs. Appropriate, quality-assured animal models are essential to improve understanding of the pathogenic interplay in CT/NG co-infections.

Introduction: A Neisseria meningitidis carrier study has been conducted among students at Örebro university in Sweden in 2018 and 2019. Pharyngeal samples were collected from 3489 students. PCR for the genes ctrA and crgA was run on all samples. The positive samples were then cultured on agar plates to find the N. meningitidis. In 349 of the PCR positive samples, no N. meningitidis could be isolated, which raised the question if other bacteria could have these genes. The most likely bacteria to have these genes were assumed to be other species within the genus Neisseria. Aim: To identify whether other neisserial species have the ctrA and crgA genes. Methods: The 349 samples were cultured on agar plates for two days. The species were then identified by MALDI-TOF MS. The isolated Neisserial species and some other species as controls were saved. PCR for ctrA and crgA genes were then run on these bacteria to determine whether they possessed these genes. Results: Five N. meningitidis that had been missed by the first round of culture were identified. Seventy-five other colonies of neisserial species were isolated. N. subflava (n=40) were the most common. Nine (12 %) were crgA positive but none were ctrA positive. At least one crgA positive colony was found in four of the five different non-meningococcal neisserial species isolated in this study. Conclusion: The crgA gene seems quite common among non-meningococcal neisserial species while ctrA seems to be specific for N. meningitidis

Gonorrhea is the second most common sexually transmitted infection worldwide and is caused by Gram-negative, human-specific diplococcus Neisseria gonorrhoeae. It colonizes the mucosal surface of the female reproductive tract and the male urethra. A rapid increase in antibiotic resistance makes gonorrhea a serious threat to public health worldwide. Since N. gonorrhoeae is a human-specific pathogen, animal infection models are not able to recapitulate all the features of infection. Therefore, a realistic in vitro cell culture model is urgently required for studying the gonorrhea infection. In this study, we established and characterized three independent 3D tissue models based on the porcine small intestinal submucosa (SIS) scaffold by co-culturing human dermal fibroblasts with human colorectal carcinoma, endometrial epithelial, and male uroepithelial cells. The histological, immunohistochemical, and ultra-structural analysis showed that the 3D SIS scaffold-based models closely mimic the main characteristics of the site of gonococcal infection in the human host including the formation of epithelial monolayer, underlying connective tissue, mucus production, tight junction (TJ), and microvilli. In addition, functional analysis such as transepithelial electrical resistance (TEER) and barrier permeability indicated high barrier integrity of the cell layer. We infected the established 3D tissue models with different N. gonorrhoeae strains and derivatives presenting various phenotypes regarding adhesion and invasion. The results showed disruption of TJs and growing the interleukins production in response to the infection, which depends on the type of strain and cell. In addition, the 3D tissue models supported bacterial survival, which provided an appropriate in vitro model for long-term infection study. This could be mainly because of the high resilience of the 3D tissue models based on the SIS scaffold to the infection in terms of alteration in permeability, cell destruction, and bacterial transmigration. During gonorrhea infection, a high level of neutrophils migrates to the site of infection. The studies also showed that N. gonorrhoeae can survive or even replicate inside the neutrophils. Therefore, studying the interaction between neutrophils and N. gonorrhoeae is substantially under scrutiny. For this purpose, we generated a 3D tissue model by triple co-culturing of human primary fibroblast cells, human colorectal carcinoma cells, and human umbilical vein endothelial cells. The tissue model was subsequently infected by N. gonorrhoeae. A perfusion-based bioreactor system was employed to recreate blood flow in the side of endothelial cells and consequently study human neutrophils transmigration to the site of infection. We observed neutrophils activation upon the infection. Furthermore, we demonstrated the uptake of N. gonorrhoeae by human neutrophils and reverse transmigration of neutrophils to the basal side carrying N. gonorrhoeae. In summary, the introduced 3D tissue models in this research represent a promising tool to investigate N. gonorrhoeae infections under close-to-natural conditions. / Tripper ist die zweithäufigste sexuell übertragbare Krankheit weltweit und wird durch Gram negative, humanspezifische Diplokokken Neisseria gonorrhoeae verursacht. Das human Pathogen besiedelt die Schleimhautoberfläche des weiblichen Fortpflanzungstraktes und der männlichen Harnröhre. Die rasante Zunahme der Antibiotikaresistenzen macht Gonorrhö zu einer ernsthaften Bedrohung für die öffentliche Gesundheit weltweit. Da N. gonorrhoeae ein humanspezifischer Erreger ist, ist es nicht möglich alle Merkmale einer Infektion in Tiermodellen nachzustellen, daher ist ein realistisches In-vitro-Zellkulturmodell für die Untersuchung der Gonorrhö-Infektion dringend erforderlich. In dieser Studie haben wir drei unabhängige 3D- Gewebemodelle etabliert und charakterisiert, die auf dem Gerüst der Schweine-Submukosa (SIS) basieren, indem wir menschliche dermale Fibroblasten mit menschlichen Darmkrebs-, Endometrialepithel- und männlichen Uroepithelzellen kultivieren. Die histologischen, immunhistochemischen und ultrastrukturellen Analysen zeigten, dass die 3D SIS-Gerüstmodelle die Hauptmerkmale der Stelle der Gonokokken Infektion im menschlichen Wirt genau nachahmen, indem sich Epithelien Monoschichten ausbildeten, unter denen sich Bindegewebe erstrechte. Zudem wiesen die Zellen enge Zell-Zell Kontakte auf und es kam zur Produktion von einer Mukosaschicht sowie Mikrovilli in den Modellen. Darüber hinaus zeigten Funktionsanalysen wie die Messung des transepithelialen elektrischen Widerstands (TEER) und die der Barriere Durchlässigkeit eine hohe Barriere Integrität der Zellschicht. Wir haben die etablierten 3D- Gewebemodelle mit verschiedenen N. gonorrhoeae-Stämmen und Derivaten infiziert, die verschiedene Phänotypen bezüglich der Adhäsion und der Invasion aufwiesen. Die Ergebnisse zeigten eine Unterbrechung der engen Zellverbindungen und eine Zunahme der Interleukin Produktion als Reaktion auf die Infektion, dessen Ausprägung allerdings stark von der Art des Stammes und des verwendeten Zelltyps abhängig ist. Darüber hinaus unterstützten die 3D- Gewebemodelle das bakterielle Überleben, was ein geeignetes In-vitro-Modell für Langzeit- Infektionsstudien liefert. Dies könnte vor allem auf die hohe Widerstandsfähigkeit der SIS- Gerüstmodelle gegen Infektionen in Bezug auf Veränderung der Permeabilität, Zellzerstörung und Bakterienwanderung zurückzuführen sein. Während der Gonorrhoe-Infektion wandert ein hoher Anteil an Neutrophilen an den Ort der Infektion. Die Studie zeigte auch, dass N. gonorrhoeae in den Neutrophilen überleben konnten oder sich sogar in diesen vermehren konnten. Deshalb wurde besonderes die Interaktion zwischen Neutrophilen und N. gonorrhoeae näher betrachtet. Zu diesem Zweck haben wir ein 3D-Gewebemodell mit Hilfe einer dreifache Co-Kultivierung von menschlichen primären Fibroblasten Zellen, menschlichen kolorektalen Karzinomzellen und menschlichen Nabelvenenendothelzellen erstellt. Das Gewebemodell wurde anschließend mit N. gonorrhoeae infiziert. Ein perfusionsbasiertes Bioreaktorsystem wurde eingesetzt, um den Blutfluss auf der Seite der Endothelzellen nachzuahmen und somit konnte die Auswanderung menschlicher Neutrophile zur Infektionsstelle untersucht werden. Darüber hinaus konnte mit diesem Modell die Aufnahme von N. gonorrhoeae in menschliche Neutrophilen und deren Rückwanderung in den Blutfluss beladen mit N. gonorrhoeae nachgewiesen werden. Zusammenfassend lässt sich sagen, dass das in dieser Forschung vorgestellte 3D-Gewebemodell ein vielversprechendes Instrument zur Untersuchung von N. gonorrhoeae-Infektionen unter naturnahen Bedingungen darstellt.

Neisseria gonorrhoeae are Gram-negative bacteria with diplococcal shape. As an obligate human pathogen, it is the causative agent of gonorrhoea, a sexually transmitted disease. Gonococci colonize a variety of mucosal tissues, mainly the urogenital tract in men and women. Occasionally N. gonorrhoeae invades the bloodstream, leading to disseminated gonococcal infection. These bacteria possess a repertoire of virulence factors, which expression patterns can be adapted to the environmental conditions of the host. Through the accumulation of antibiotic resistances and in absence of vaccines, some neisserial strains have the potential to spread globally and represent a major public health threat. Therefore, it is necessary to understand the exact molecular mechanisms underlying the successful infection and progression of gonococci within their host. This deeper understanding of neisserial infection and survival mechanisms is needed for the development of new therapeutic agents. In this work, the role of host-cell sphingolipids on the intracellular survival of N. gonorrhoeae was investigated. It was shown that different classes of sphingolipids strongly interact with invasive gonococci in epithelial cells. Therefore, novel and highly specific clickable sphingolipid analogues were applied to study these interactions with this pathogen. The formation of intra- and extracellular sphingosine vesicles, which were able to target gonococci, was observed. This direct interaction led to the uptake and incorporation of sphingosine into the neisserial membrane. Together with in vitro results, sphingosine was identified as a potential bactericidal reagent as part of the host cell defence. By using different classes of sphingolipids and their clickable analogues, essential structural features, which seem to trigger the bacterial uptake, were detected. Furthermore, effects of key enzymes of the sphingolipid signalling pathway were tested in a neutrophil infection model. In conclusion, the combination of click chemistry and infection biology made it possible to shed some light on the dynamic interplay between cellular sphingosine and N. gonorrhoeae. Thereby, a possible “catch-and-kill” mechanism could have been observed. / Neisseria gonorrhoeae ist ein Gram-negatives Bakterium, welches als Diplokokke vorkommt. Als ein ausschließliches Humanpathogen sind Neisserien der Erreger für die sexuell übertragbare Infektionskrankheit Gonorrhö. Gonokokken besiedeln eine Vielzahl von Schleimhäuten, jedoch hauptsächlich den Urogenitaltrakt bei Männern und Frauen. Gelegentlich kann N. gonorrhoeae in die Blutbahn invadieren, was zu einer disseminierten Infektion führen kann. Diese Bakterien verfügen über ein Repertoire an Virulenzfaktoren, deren Expressionskombination den Umgebungsbedingungen des Wirts angepasst werden können. Durch die Anhäufung von Antibiotikaresistenzen und durch das Fehlen eines Impfstoffes, besteht die Gefahr, dass spezielle Neisserienstämme sich weltweit verbreiten und daher eine ernstzunehmende Bedrohung des Menschen sind. Daher ist es notwendig die zugrundeliegenden molekularen Mechanismen der erfolgreichen Infektion und Ausbreitung der Gonokokken im Wirt genauestens zu verstehen. Das detaillierte Wissen über die Neisserieninfektion und Überlebensmechanismen ist nötig für die Entwicklung neuer Therapieansätze. In dieser Arbeit wurde der Effekt von Sphingolipiden der Wirtszelle auf das intrazelluläre Überleben von N. gonorrhoeae untersucht. Es konnte gezeigt werden, dass unterschiedliche Klassen von Sphingolipiden stark mit invasiven Gonokokken in Epithelzellen interagieren. Um dies zu tun, wurden neue und hochspezifische clickbare Sphingolipidanaloge eingesetzt, um deren Interaktionen mit diesem Pathogen zu studieren. Die Formation von intra- als auch extrazellulären Sphingosinvesikeln, welche Gonokokken gezielt erreichten, konnte beobachtet werden. Diese direkte Interaktion führte zu einer Aufnahme und Einbau des Sphingosins in die Neisserienmembran. Zusammen mit in vitro Ergebnissen, konnte Sphingosin als potenzieller und antibakterieller Bestandteil des zellulären Abwehrsystems identifiziert werden. Weiterhin wurde durch die Verwendung unterschiedlicher Sphingolipidklassen und deren clickbaren Analoge wichtige Strukturen erkannt, die die bakterielle Aufnahme auslösen. Des Weiteren wurden die Auswirkungen von Schlüsselenzymen des Sphingolipidsignalwegs in einem Infektionsmodell mit Neutrophilen getestet. Abschließend ist zu sagen, dass die Kombination aus Click Chemie und Infektionsbiologie es ermöglicht hat, die dynamischen Wechselwirkungen zwischen zellulären Sphingosin und N. gonorrhoeae zu beleuchten. Dadurch konnte ein möglicher „catch-and-kill”-Mechanismus entdeckt werden.

Neisseria gonorrhoeae, the causative agent of gonorrhea, is the second most prevalent sexually transmitted infection that leads to substantial morbidity and economic burden worldwide. Improperly treated or untreated gonorrhea can lead to severe and life-threatening complications, including abortion, infertility, pelvic pain, and maternal death. Neisseria gonorrhoeae has developed resistance to the formally and currently used antibiotics. The Centers for Disease Control and Prevention (CDC) have listed multi-drug resistant N. gonorrhoeae as an urgent threat that promptly requires the development of novel therapeutic agents. Traditional drug discovery and development is a time-consuming and costly process associated with high risks. To address the dire need to replenish the dry pipeline of anti-gonorrhea medications, drug repurposing is a promising approach. In this study, an FDA-approved drug library was screened, and 14 drugs were found to exhibit promising anti-gonococcal activity. Interestingly, three extremely potent and narrow-spectrum novel candidates, itraconazole, isavuconazole, and ravuconazole, are azole antifungals, and their activities were further investigated in vitro. Of the three azoles, ravuconazole displayed the most potent activity against N. gonorrhoeae clinical isolates. The time-kill assay revealed that the three azoles showed bactericidal activity. All three azole drugs showed a low frequency of resistance. Besides, isavuconazole and ravuconazole have a longer post-antibiotic effect than azithromycin. All three azoles cleared the burden of intracellular N. gonorrhoeae completely, which is superior to ceftriaxone. In conclusion, itraconazole, isavuconazole, and ravuconazole merit future investigation for the development of anti-gonorrheal therapeutics. This study provided unexplored avenues and promising opportunities that can be further evaluated to combat N. gonorrhoeae infection. / Master of Science / Neisseria gonorrhoeae, the causative agent of gonorrhea, is the second most prevalent sexually transmitted infection that leads to substantial morbidity and economic burden worldwide. Improperly treated or untreated gonorrhea can lead to severe and life-threatening complications, including abortion, infertility, pelvic pain, and maternal death. Due to the increasing prevalence of drug resistance against the formally and currently used antibiotics, the Centers for Disease Control and Prevention (CDC) have classified multi-drug resistant N. gonorrhoeae as an urgent-threat pathogen. Therefore, the discovery of new anti-gonorrheal therapeutics is an urgent need. Drug repurposing is the process of discovering new therapeutic uses for approved or investigational drugs that go beyond the original medical indication. To address the dire need to replenish the dry pipeline of anti-gonorrheal drugs, repurposing FDA-approved drugs is a promising approach as it significantly reduces the time and expense associated with traditional drug development. By screening an FDA-approved drug library, 14 drugs were found to display promising anti-gonococcal activity. Interestingly, three (itraconazole, isavuconazole, and ravuconazole) out of 14 identified drugs were azole antifungal drugs, and their activities were further investigated in vitro. All three azole drugs showed bactericidal activity, meaning that they killed bacteria, had a low propensity to develop resistance, and completely cleared the burden of intracellular N. gonorrhoeae. Besides, our findings suggested that isavuconazole and ravuconazole possessed exceptional activity in the suppression of bacterial growth following brief antibiotic exposure. In conclusion, the three azole drugs exhibited potent anti-gonococcal activity and merited further investigation. This study provided unexplored avenues and promising opportunities that can be further evaluated to combat multidrug-resistant N. gonorrhoeae. Neisseria gonorrhoeae, the causative agent of gonorrhea, is the second most prevalent sexually transmitted infection that leads to substantial morbidity and economic burden worldwide. Improperly treated or untreated gonorrhea can lead to severe and life-threatening complications, including abortion, infertility, pelvic pain, and maternal death. Due to the increasing prevalence of drug resistance against the formally and currently used antibiotics, the Centers for Disease Control and Prevention (CDC) have classified multi-drug resistant N. gonorrhoeae as an urgent-threat pathogen. Therefore, the discovery of new anti-gonorrheal therapeutics is an urgent need. Drug repurposing is the process of discovering new therapeutic uses for approved or investigational drugs that go beyond the original medical indication. To address the dire need to replenish the dry pipeline of anti-gonorrheal drugs, repurposing FDA-approved drugs is a promising approach as it significantly reduces the time and expense associated with traditional drug development. By screening an FDA-approved drug library, 14 drugs were found to display promising anti-gonococcal activity. Interestingly, three (itraconazole, isavuconazole, and ravuconazole) out of 14 identified drugs were azole antifungal drugs, and their activities were further investigated in vitro. All three azole drugs showed bactericidal activity, meaning that they killed bacteria, had a low propensity to develop resistance, and completely cleared the burden of intracellular N. gonorrhoeae. Besides, our findings suggested that isavuconazole and ravuconazole possessed exceptional activity in the suppression of bacterial growth following brief antibiotic exposure. In conclusion, the three azole drugs exhibited potent anti-gonococcal activity and merited further investigation. This study provided unexplored avenues and promising opportunities that can be further evaluated to combat multidrug-resistant N. gonorrhoeae.

No description available.

Neisseria meningitidis B libera vesículas de membrana externa, conhecidas pela sigla OMV. Essas possuem os mesmos componentes da membrana externa da bactéria e podem ser utilizadas como antígenos em vacinas contra a meningite B. As vesículas devem, também, expressar proteínas da membrana externa (OMP) e proteínas reguladoras do íon ferro (IRP). O objetivo deste trabalho é estudar a cinética de crescimento bacteriano, consumo das fontes de carbono e nitrogênio - especialmente os limitantes de crescimento ? e produção de OMV visando melhorar a produção desse antígeno. Realizaram-se cultivos descontínuos em biorreator, com duração de 20 h, empregando meio de Catlin com limitação de ferro e modificações nas concentrações de lactato, aminoácidos e glicerol. As condições do cultivo foram: 4,2 L de meio, temperatura de 36°C, pressão de 0,5 atm, vazão de ar 1 L/min, agitação entre 250-850 rpm, controle de oxigênio dissolvido em 10% de saturação. Constatou-se que o lactato é a principal fonte de carbono limitante, embora somente se tem a hipótese de que o glicerol age como protetor mecânico. O ácido L-glutâmico é a principal fonte de nitrogênio consumida durante o cultivo. As OMV começaram a ser liberadas quantitativamente no início da fase estacionária de crescimento. Sendo que a melhor condição para a produção de OMV, valor 162,3 mg/L, é aquela em que as concentrações iniciais de lactato e aminoácidos foram duplicadas, 15,00 g/L e 2,93 g/L respectivamente. Através da análise do padrão eletroforético, confirmou-se a presença das principais proteínas de superfície, inclusive das IRPs. A integridade da OMV foi constatada por microscopia eletrônica. Assim, o antígeno obtido mostra-se passível de utilização na composição de vacina anti-meningocócica. / Neisseria meningitidis B liberates outer membrane vesicles known by the abbreviation OMV. These vesicles have the same components of the outer membrane of the bacteria and may be used as antigens in vaccines against meningitis B. The vesicles must also express outer membrane proteins (OMP) and iron regulated proteins (IRP). The aim of this paper is to study bacterial growth kinetics, carbon and nitrogen sources consumption ? specially those which limit growth ? and OMV production, seeking to improve the production of this antigen. Discontinuous bioreactor cultivations were carried out for a period of 20 hours in Catlin medium with iron restriction and modifications in lactate, amino acid, and glycerol concentrations. Cultivation conditions were: 4,2 L of medium, temperature at 36ºC, 0,5 atm, air flow rate of 1 L/min, agitation between 250-850 rpm, and dissolved oxygen control at 10% of saturation. It was verified that lactate is the main limiting carbon source, although there is just a hypothesis that glycerol acts as a mechanic protector. The L-glutamic acid is the main source of nitrogen consumed during the cultivation. The OMV started to be liberated quantitatively at the beginning of the stationary phase of growth. The best condition for production of OMV, value 162,3 mg/L, is that where the initial concentrations of lactate and amino acids were duplicated, 15,00 g/L and 2,93 g/L, respectively. Through an analysis of the electroforetic pattern, the presence of the main surface proteins was confirmed, including the IRPs. The integrity of the OMV was testified by electronic microscopy. So, the antigen thus obtained may be used in the antimeningococcal vaccine composition.

INTRODUÇÃO: Neisseria meningitidis é uma das principais causas de meningite bacteriana em todo o mundo. No Brasil, a vacina meningocócica conjugada C foi introduzida no calendário básico do Programa Nacional de Imunizações em 2010 para a população abaixo de 2 anos de idade. Este estudo teve como objetivos avaliar a taxa de portadores sadios de meningococo em nasofaringe entre estudantes de medicina através de métodos fenotípicos e genotípicos. Além disso, foram analisados os fatores de risco associados ao estado de portador sadio. MÉTODOS: Trezentos e setenta e cinco (375) estudantes da Faculdade de Medicina da Universidade de São Paulo foram submetidos à coleta de swabs de nasofaringe, de outubro a novembro de 2010: desta casuística, 190 estudantes foram submetidos à coleta de dois swabs. Os swabs foram encaminhados ao Laboratório de Investigação Médica (LIM) de Bacteriologia para isolamento e caracterização das cepas: o primeiro swab foi submetido a cultivo em meio seletivo, testes bioquímicos e reação em cadeia da polimerase com primers específicos para dois genes de Neisseria meningitidis, ctrA e crgA. O segundo swab foi submetido à extração direta de DNA, sem cultivo prévio. Todas as cepas isoladas foram genogrupadas com primers específicos direcionados aos genogrupos A, B, C, Y e W. RESULTADOS: A prevalência geral de portadores sadios foi 45,6%. A taxa de portadores obtida após cultivo em meio seletivo e testes bioquímicos foi 6,1%. A taxa de colonização resultante da PCR dos produtos de cultivo em meio seletivo foi 15.4%. Destas cepas, a maioria pertencia ao genogrupo B. A partir da extração direta de DNA de 190 amostras clínicas, obtivemos 69,5% de positividade de portadores sadios, sendo o genogrupo C predominante nestas amostras. A PFGE mostrou que todas as cepas isoladas eram policlonais. Não houve relação estatisticamente significativa entre os fatores de risco avaliados e o estado de portador sadio. CONCLUSÕES: Este estudo avaliou a prevalência de portadores sadios em adultos jovens no Brasil antes da introdução da vacina meningocócica C conjugada no calendário vacinal de rotina. A prevalência variou de 6,1% a 69,5% dentre os diferentes métodos utilizados, com uma prevalência geral de 45,6%. Comparando-se os diferentes métodos, os métodos genotípicos mostraram-se superiores na detecção de portadores sadios, especialmente quando o DNA é extraído diretamente da amostra clínica / BACKGROUND: Neisseria meningitidis is one of the leading causes of bacterial meningitis worldwide. In Brazil, the meningococcal C conjugate vaccine was introduced as a part of basic vaccination schedule in the public health system in 2010 to the population under 2 years old. The objectives of this study were to evaluate the rate of asymptomatic carriers of meningococci in the nasopharynx among medical students by phenotypic and genotypic methods. Furthermore, the risk factors for carriage were also analyzed. METHODS: Three hundred seventy-five (375) students from University of São Paulo Medical School underwent nasopharyngeal swab collection from October to November, 2010. From those students, 190 of them underwent collection of two swabs; they were sent to the laboratory for isolation and characterization of strains: the first one were submitted to culture on solid plates with selective medium, biochemical tests, and Polymerase Chain Reaction (PCR) with specific primers for two genes of Neisseria meningitidis, ctrA and crgA. From the second swab, direct DNA extraction to PCR reactions were performed, with no previous culture. All strains isolated were genogrouped with specific primers targeting A, B, C, Y and W groups. RESULTS: The overall carriage rate was 45,6%. Carriage rate by culture on solid plates and biochemical tests was 6.1%. The rate obtained from PCR targeting genes ctrA and crgA of colony growth on selective medium was 15.4%. From those strains, the majority belongs to genogroup B. With direct extraction of DNA from 190 clinical samples, we obtained a positivity of 69.5% of meningococcal carriage, and the genogroup C was the main genogroup from those samples. The PFGE showed that the samples were polyclonal. CONCLUSIONS: This study has evaluated the rate of asymptomatic carriers in young people in Brazil before the vaccine introduction in the vaccine routine schedule of infants. The prevalence of nasopharyngeal carriage by meningoccoci among medical students was 6.1% for the phenotypic method, 15.4% by PCR after growth in selective medium and 69.5% by PCR from DNA extracted from clinical samples. In the comparison of the phenotypic and genotypic methods, it was observed that genotypic are superior to phenotypic methods, especially when direct DNA extraction from clinical specimen is performed

Neisseria meningitidis possède un PTS incomplet. Constitué des composants générales EI et HPr et de deux EIIAs (EIIANtr et EIIAMan), ce système ne permet pas le transport des sucres chez cette bactérie. Cependant, nous avons confirmé que la cascade de phosphorylation (EI HPr EIIANtr) est fonctionnelle et que HPr est aussi phosphorylée sur sa Ser-46 par une HprK/P.Dans l’objectif d’étudier l’effet de HPr sur la virulence de N. meningitidis, nous avons construit un mutant ΔptsH chez N. meningitidis 2C4-3. Le mutant ΔptsH a montré une faible survie chez la souris, une faible production de la capsule, une meilleure adhérence aux cellules épithéliales et un niveau élevé de cellules apoptotiques, par rapport à la souche sauvage. HPr semble intervenir dans la virulence de N. meningitidis en interagissant avec la protéine CrgA. L’interaction HPr/CrgA est plus forte quand HPr est phosphorylée sur sa Ser-46 par HprK/P.N. meningitidis utilise le glucose et le maltose comme seuls sucres. Nous avons identifié une perméase à glucose (GlcP) et une perméase à maltose (MalY), responsables du transport de ces sucres. Une perméase putative à gluconate (GntP) a été également identifiée chez N. meningitidis 2C4-3. Cette perméase n’assure pas le transport du gluconate, dans les conditions testées. La délétion de gntP chez N. meningitidis 2C4-3 induit une meilleure croissance sur glucose et une bonne survie du mutant ΔgntP chez la souris, par rapport à la souche sauvage. La fonction réelle de la perméase GntP chez N. meningitidis reste inconnue et suscite des études ultérieures. / Neisseria meningitidis has an incomplete PTS composed of general proteins EI and HPr and two EIIAs (EIIANtr and EIIAMan). This system does not allow the transport of sugars in this bacterium. However, we confirmed that the phosphorylation cascade (EI HPr EIIANtr) is functional and HPr is also phosphorylated at its Ser-46 by an HprK/P.In order to study the effect of HPr on meningococcal virulence, we constructed a ΔptsH mutant in N. meningitidis 2C4-3. Compared to the wild-type strain, the ΔptsH mutant showed poor survival in mice, low production of capsule, better adherence to epithelial cells and high levels of apoptotic cells. HPr appears to be involved in the virulence of N. meningitidis by interacting with CrgA protein. The HPr/CrgA interaction is stronger when HPr is phosphorylated at its Ser-46 by HprK/P.N. meningitidis uses glucose and maltose as the only sugars. We identified permeases for glucose (GlcP) and maltose (MalY), which catalyze the uptake of these sugars.A putative gluconate permease (GntP) has also been identified in N. meningitidis 2C4-3. Under the conditions tested, this permease did not catalyze the transport of gluconate. Compared to the wild-type strain, deletion of gntP in N. meningitidis 2C4-3 induced faster growth on glucose and a better survival of the mutant in mice. To underst and the function of the GntP permease in N. meningitidis further studies will have to be carried out.

Beställningsfrekvensen för detektion av Trichomonas vaginalis samt Mycoplasma genitalium i Jönköpings län är låg jämfört med den för Chlamydia trachomatis och Neisseria gonorrhoeae.  Både T. vaginalis och M. genitalium har associerats med infektion av humant papillomvirus (HPV) samt kan bland annat orsaka salpingit med infertilitet som potentiell komplikation. Patogenerna har även beskrivits öka risken för transmission av HIV. Syftet med studien var att detektera T. vaginalis samt M. genitalium med realtids-Polymerase Chain Reaction (PCR) för uppskattning av prevalens hos individer provtagna för C. trachomatis, N. gonorrhoeae samt HPV i Jönköpings län. Hos individer över 25 år, provtagna för C. trachomatis och N. gonorrhoeae, uppskattades prevalensen till 5,5 % för M. genitalium samt 0,13 % för T. vaginalis. Hos samma individer var prevalensen av C. trachomatis och N. gonorrhoeae 4,5 % respektive 0,13 %. Prevalensen hos individer provtagna för HPV uppskattades till 2,3 % för M. genitalium samt 0,26 % för T. vaginalis. De slutsatser som dras är att relevans finns för en mer frekvent beställning av M. genitalium samt att analys för detektion av endast en patogen ej är optimal. Multiplex analys för detektion av sexuellt överförbara patogener föreslås. / The request for detection of Trichomonas vaginalis and Mycoplasma genitalium in Jönköping County is low compared to Chlamydia trachomatis and Neisseria gonorrhoeae. Both T. vaginalis and M. genitalium have been associated with Human Papilloma Virus (HPV) infection and can cause infections such as salpingitis, potentially resulting in infertility. The pathogens have also been described to increase the risk of HIV transmission. The aim of this study was to detect T. vaginalis and M. genitalium by real-time Polymerase Chain Reaction (PCR) to estimate the prevalence among individuals tested for C. trachomatis, N. gonorrhoeae and HPV in Jönköping County. In individuals above the age of 25 years, tested for C. trachomatis and N. gonorrhoeae, the prevalence was estimated to 5,5 % for M. genitalium and 0,13 % for T. vaginalis. In the same group the prevalence of C. trachomatis and N. gonorrhoeae was 4,5 % and 0,13 % respectively. The prevalence in individuals tested for HPV was estimated to 2,3 % for M. genitalium and 0,26 % for T. vaginalis. Relevance of a more frequent request for detection of M. genitalium was concluded and single pathogen detection was not deemed to be optimal. Multiplex analysis for detection of sexually transmitted pathogens is encouraged.