Genomics of "chlamydia"-related bacteria

AbstractThe Chlamydiales order is an important bacterial phylum that comprises some of the most successful human pathogens such as Chlamydia trachomatis, the leading infectious cause of blindness worldwide. Since some years, several new bacteria related to Chlamydia have been discovered in clinical or environmental samples and might represent emerging pathogens. The genome sequencing of classical Chlamydia has brought invaluable information on these obligate intracellular bacteria otherwise difficult to study due to the lack of tools to perform basic genetic manipulation. The recent emergence of high-throughput sequencing technologies yielding millions of reads in a short time lowered the costs of genome sequencing and thus represented a unique opportunity to study Chlamydia-re\ated bacteria. Based on the sequencing and the analysis of Chlamydiales genomes, this thesis provides significant insights into the genetic determinants of the intracellular lifestyle, the pathogenicity, the metabolism and the evolution of Chlamydia-related bacteria. A first approach showed the efficacy of rapid sequencing coupled to proteomics to identify immunogenic proteins. This method, particularly useful for an emerging pathogen such as Parachlamydia acanthamoebae, enabled us to discover good candidates for the development of diagnostic tools that would permit to evaluate at larger scale the role of this bacterium in disease. Second, the complete genome of Waddlia chondrophila, a potential agent of miscarriage, encodes numerous virulence factors to manipulate its host cell and resist to environmental stresses. The reconstruction of metabolic pathways showed that the bacterium possesses extensive capabilities compared to related organisms. However, it is still incapable of synthesizing some essential components and thus has to import them from its host. Third, the genome comparison of Protochlamydia naegleriophila to its closest known relative Protochlamydia amoebophila revealed a particular evolutionary dynamic with the occurrence of an unexpected genome rearrangement. Fourth, a phylogenetic analysis of P. acanthamoebae and Legionella drancourtii identified several genes probably exchanged by horizontal gene transfer with other intracellular bacteria that might occur within their amoebal host. These genes often encode mechanisms for resistance to metal or toxic compounds. As a whole, the analysis of the different genomes enabled us to highlight a large diversity in size, GC percentage, repeat content as well as plasmid organization. The abundant genomic data obtained during this thesis have a wide impact since they provide the necessary bases for detailed investigations on countless aspects of the biology and the evolution of Chlamydia-related bacteria, whether in wet lab or by bioinformatical analyses.RésuméL'ordre des Chlamydiales est un important phylum bactérien qui comprend de nombreuses espèces pathogènes pour l'homme et les animaux, dont Chlamydia trachomatis, responsable du trachome, la cause majeure de cécité d'origine infectieuse à travers le monde. Durant ces dernières décennies, de nombreuses bactéries apparentées aux Chlamydia ont été découvertes dans des échantillons environnementaux ou cliniques mais leur éventuel rôle pathogène dans le développement de maladies reste peu connu. Ces bactéries sont des intracellulaires obligatoires car elles ont besoin d'une cellule hôte pour se multiplier, ce qui rend leur étude particulièrement difficile. Le développement de nouvelles technologies permettant de séquencer le génome d'un organisme rapidement et à moindre coût ainsi que l'essor des méthodes d'analyse s'y rapportant représentent une opportunité exceptionnelle d'étudier ces organismes. Dans ce contexte, cette thèse démontre l'utilité de la génomique pour développer de nouveaux outils diagnostiques ainsi que pour étudier le métabolisme de ces bactéries, leurs facteurs de virulence et leur évolution.Ainsi, une première approche a illustré l'utilité d'un séquençage rapide pour obtenir les informations nécessaires à l'identification de protéines qui sont reconnues par des anticorps humains ou animaux. Cette méthode, particulièrement utile pour un pathogène émergent tel que Parachlamydia acanthamoebae, a permis de découvrir de bons candidats pour le développement d'un outil diagnostique qui permettrait d'évaluer à plus large échelle le rôle de cette bactérie notamment dans la pneumonie. L'analyse du contenu génique de Waddlia chondrophila, un autre germe qui pourrait être impliqué dans les avortements et tes fausses-couches, a en outre mis en évidence la présence de nombreux facteurs connus qui lui permettent de manipuler son hôte. Cette bactérie possède de plus grandes capacités métaboliques que les autres Chlamydia, mais elle est incapable de synthétiser certains composants et doit donc les importer de son hôte pour subvenir à ses besoins. La comparaison du génome de Protochlamydia naegleriophila à son plus proche parent, Protochlamydia amoebophila, a dévoilé une évolution dynamique particulière avec l'occurrence d'un réarrangement majeur inattendu après la séparation de ces deux espèces. En outre, ces études ont montré l'occurrence de plusieurs transferts de gène avec d'autres organismes plus éloignés, notamment d'autres intracellulaires d'amibes, souvent pour l'acquisition de mécanismes de résistances à des composés toxiques. Les données génomiques acquises durant ce travail posent les fondements nécessaires a de nombreuses analyses qui permettront progressivement de mieux comprendre de nombreux aspects de ces bactéries fascinantes.

Chlamydia: signes d'appel, diagnostic et traitement [Chlamydia: diagnostic and treatment].

Chlamydia are obligate intracellular bacteria. Three species are considered human pathogens. Chlamydophila pneumoniae is one of the most common agents of atypical community-acquired pneumonia. Chlamydophila psittaci causes psittacosis, a severe zoonotic pneumonia transmitted by birds. Finally, Chlamydia trachomatis is the etiologic agent of trachoma and urogenital infections. The latter are commonly asymptomatic or paucisymptomatic. Thus, they may remain undiagnosed for years, leading to serious late complications such as salpingitis, ectopic pregnancy and infertility. Currently, the diagnosis of chlamydial infections is essentially based on molecular methods. Treatment should use an antibiotic with good intracellular bioavailability such as tetracycline, macrolides and new generation fluoroquinolones.

The High Prevalence and Diversity of Chlamydiales DNA within Ixodes ricinus Ticks Suggest a Role for Ticks as Reservoirs and Vectors of Chlamydia-Related Bacteria.

The Chlamydiales order is composed of nine families of strictly intracellular bacteria. Among them, Chlamydia trachomatis, C. pneumoniae, and C. psittaci are established human pathogens, whereas Waddlia chondrophila and Parachlamydia acanthamoebae have emerged as new pathogens in humans. However, despite their medical importance, their biodiversity and ecology remain to be studied. Even if arthropods and, particularly, ticks are well known to be vectors of numerous infectious agents such as viruses and bacteria, virtually nothing is known about ticks and chlamydia. This study investigated the prevalence of Chlamydiae in ticks. Specifically, 62,889 Ixodes ricinus ticks, consolidated into 8,534 pools, were sampled in 172 collection sites throughout Switzerland and were investigated using pan-Chlamydiales quantitative PCR (qPCR) for the presence of Chlamydiales DNA. Among the pools, 543 (6.4%) gave positive results and the estimated prevalence in individual ticks was 0.89%. Among those pools with positive results, we obtained 16S rRNA sequences for 359 samples, allowing classification of Chlamydiales DNA at the family level. A high level of biodiversity was observed, since six of the nine families belonging to the Chlamydiales order were detected. Those most common were Parachlamydiaceae (33.1%) and Rhabdochlamydiaceae (29.2%). "Unclassified Chlamydiales" (31.8%) were also often detected. Thanks to the huge amount of Chlamydiales DNA recovered from ticks, this report opens up new perspectives on further work focusing on whole-genome sequencing to increase our knowledge about Chlamydiales biodiversity. This report of an epidemiological study also demonstrates the presence of Chlamydia-related bacteria within Ixodes ricinus ticks and suggests a role for ticks in the transmission of and as a reservoir for these emerging pathogenic Chlamydia-related bacteria.

Is Switzerland doing enough regarding the prevention of syphilis, gonorrhea and chlamydia infections ?

Sexually transmitted infections are a major problem for medicine and for public health services worldwide. More than 30 sexually transmittable pathogenic micro-organisms are known, including bacteria, viruses, fungi, protozoa and ectoparasites. According to estimates from the World Health Organisation more than 333 million of bacterial sexually transmitted infections occur worldwide per year. Sexually transmitted infections, by their nature, affect individuals, within partnerships and larger sexual networks, and in turn populations. This report focuses on three bacterial sexually transmitted infections in Switzerland that are Chlamydia trachomatis, Neisseria gonorrhea and Treponema pallidum (syphilis) in Switzerland. The prevalence of these infections has been increasing alarmingly for a decade. All three infections can be asymptomatic and their diagnosis and treatment can therefore occur too late or worse not at all, even though treatments are available. This is an important problem as untreated sexually transmitted infections may cause complications such as ascending infections, infertility, ectopic pregnancies and serious long-term neurological sequels. The consequences of these infections should not be underestimated. They constitute a significant public health burden as well as serious financial burden. The increases in chlamydia, syphilis and gonorrhea infections have also been observed in many European countries. Countries, where rising numbers of sexually transmitted infections have been observed, have reacted in different ways. Some have developed clinical guidelines or implemented screening programs, while others are still in their observational phase. The aim of this mémoire is to assess whether Switzerland is doing enough regarding the prevention of chlamydial, syphilis and gonorrheal infections. After first describing the infections, surveillance systems of sexually transmitted infections are assessed, then the epidemiological trends of these three infections are described, and finally the prevention measures implemented in Switzerland to respond to the increasing number of infections are described. The reaction of the United Kingdom to the same problem is reported for comparison. [Author, p. 7]

O impacto da infecção por Chlamydia em populações indígenas da Amazônia brasileira

A disseminação das bactérias do gênero Chlamydia no Brasil, inclusive na região Amazônica, é pouco conhecida. Este estudo soroepidemiológico incluiu 2.086 amostras de soro de populações indígenas da Amazônia brasileira, empregando metodologia de triagem pela imunofluorescência indireta para pesquisa de anticorpos. Usou-se o sorotipo L2 da C. trachomatis como substrato; a seguir, para os quinze sorotipos de C. trachomatis e para a C. pneumoniae, discriminou-se a sororreatividade pela microimunofluorescência específica. A prevalência média de anticorpos para Chlamydia foi de 48,6%. Sua variação entre as comunidades indicou as que não tiveram contato com as bactérias e aquelas em que quase todos os testados tiveram. Por meio da titulação dos anticorpos IgG e a presença de IgM específica nas amostras com títulos altos viu-se que 6,1% dos infectados persistiam com a infecção, servindo de reservatórios à disseminação das espécies de Chlamydia. Pela resposta à C. trachomatis, evidenciou-se a circulação dos sorotipos A, B, Ba, D, E, G, H, I e L1. Ademais, constatou-se que há C. pneumoniae na região. As duas espécies causariam impacto significativo no hospedeiro humano.

Prevalência de coinfecção e fatores associados ao Papilomavírus Humano (HPV) e à Chlamydia tracomatis em mulheres em idade reprodutiva atendidas em Program de Prevençao de Câncer do Colo Uterino na região sudeste do Brasil: sub-titulo / (se houver)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Use of E. coli OMVs as delivery system to elicit neutralizing antibodies against Chlamydia infectivity: the HtrA protein example

The obligate intracellular pathogen Chlamydia trachomatis is a gram negative bacterium which infects epithelial cells of the reproductive tract. C. trachomatis is the leading cause of bacterial sexually transmitted disease worldwide and a vaccine against this pathogen is highly needed. Many evidences suggest that both antigen specific-Th1 cells and antibodies may be important to provide protection against Chlamydia infection. In a previous study we have identified eight new Chlamydia antigens inducing CD4-Th1 and/or antibody responses that, when combined properly, can protect mice from Chlamydia infection. However, all selected recombinant antigens, upon immunization in mice, elicited antibodies not able to neutralize Chlamydia infectivity in vitro. With the aim to improve the quality of the immune response by inducing effective neutralizing antibodies, we used a novel delivery system based on the unique capacity of E. coli Outer Membrane Vesicles (OMV) to present membrane proteins in their natural composition and conformation. We have expressed Chlamydia antigens, previously identified as vaccine candidates, in the OMV system. Among all OMV preparations, the one expressing HtrA Chlamydia antigen (OMV-HtrA), showed to be the best in terms of yield and quantity of expressed protein, was used to produce mice immune sera to be tested in neutralization assay in vitro. We observed that OMV-HtrA elicited specific antibodies able to neutralize efficiently Chlamydia infection in vitro, indicating that the presentation of the antigens in their natural conformation is crucial to induce an effective immune response. This is one of the first examples in which antibodies directed against a new Chlamydia antigen, other than MOMP (the only so far known antigen inducing neutralizing antibodies), are able to block the Chlamydia infectivity in vitro. Finally, by performing an epitope mapping study, we investigated the specificity of the antibody response induced by the recombinant HtrA and by OMV-HtrA. In particular, we identified some linear epitopes exclusively recognized by antibodies raised with the OMV-HtrA system, detecting in this manner the antigen regions likely responsible of the neutralizing effect.

The role of reinfection and partner notification in the efficacy of Chlamydia screening programs

Repeated Chlamydia trachomatis infections after treatment are common. One reason is reinfection from untreated partners in ongoing sexual partnerships. Mathematical models that are used to predict the impact of screening on reducing chlamydia prevalence often do not incorporate reinfection and might overestimate the expected impact. We describe a pair compartmental model that explicitly incorporates sexual partnership duration and reinfection. The pair model predicts a weaker impact of screening when compared directly with a model that does not accommodate partnerships. Effective management of sex partners to prevent reinfection might need to be strengthened in chlamydia control programs.

Predicting the population impact of chlamydia screening programmes: comparative mathematical modelling study

BACKGROUND: Published individual-based, dynamic sexual network modelling studies reach different conclusions about the population impact of screening for Chlamydia trachomatis. The objective of this study was to conduct a direct comparison of the effect of organised chlamydia screening in different models. METHODS: Three models simulating population-level sexual behaviour, chlamydia transmission, screening and partner notification were used. Parameters describing a hypothetical annual opportunistic screening program in 16-24 year olds were standardised, whereas other parameters from the three original studies were retained. Model predictions of the change in chlamydia prevalence were compared under a range of scenarios. RESULTS: Initial overall chlamydia prevalence rates were similar in women but not men and there were age and sex-specific differences between models. The number of screening tests carried out was comparable in all models but there were large differences in the predicted impact of screening. After 10 years of screening, the predicted reduction in chlamydia prevalence in women aged 16-44 years ranged from 4% to 85%. Screening men and women had a greater impact than screening women alone in all models. There were marked differences between models in assumptions about treatment seeking and sexual behaviour before the start of the screening intervention. CONCLUSIONS: Future models of chlamydia transmission should be fitted to both incidence and prevalence data. This meta-modelling study provides essential information for explaining differences between published studies and increasing the utility of individual-based chlamydia transmission models for policy making.

Chlamydia Control: A Comparative Review from the USA and UK

Chlamydia trachomatis infection (chlamydia) is the most common notifiable bacterial sexually transmitted infection (STI) worldwide. In the United States of America (USA) in 2009, 1,244,180 cases of chlamydia were reported to the Centers for Disease Control and Prevention (CDC), the largest number of cases ever reported to CDC for any notifiable disease [1]. It has been estimated, from population prevalence surveys, that approximately 2 % of sexually active adults aged 18–44 years old in the UK [2] and 2.2 % (CI, 1.8–2.8 %) of the US population aged 14–39 years [3] are infected with chlamydia. This level of prevalence in the USA translates into an estimated 2,291,000 (95 % confidence interval, CI, 1,857,000–2,838,000) chlamydia infections each year [3]. Globally, the World Health Organization (WHO) estimates that there are about 92 million new cases of chlamydia each year [4].

Genital chlamydia prevalence in Europe and in non-European high income countries: systematic review and meta-analysis

Background: Accurate information about the prevalence of Chlamydia trachomatis is needed to assess national prevention and control measures. Methods: We systematically reviewed population-based cross-sectional studies that estimated chlamydia prevalence in European Union/European Economic Area (EU/EEA) Member States and non-European high income countries from January 1990 to August 2012. We examined results in forest plots, explored heterogeneity using the I2 statistic, and conducted random effects meta-analysis if appropriate. Metaregression was used to examine the relationship between study characteristics and chlamydia prevalence estimates. Results: We included 25 population-based studies from 11 EU/EEA countries and 14 studies from five other high income countries. Four EU/EEA Member States reported on nationally representative surveys of sexually experienced adults aged 18-26 years (response rates 52-71%). In women, chlamydia point prevalence estimates ranged from 3.0-5.3%; the pooled average of these estimates was 3.6% (95% CI 2.4, 4.8, I2 0%). In men, estimates ranged from 2.4-7.3% (pooled average 3.5%; 95% CI 1.9, 5.2, I2 27%). Estimates in EU/EEA Member States were statistically consistent with those in other high income countries (I2 0% for women, 6% for men). There was statistical evidence of an association between survey response rate and estimated chlamydia prevalence; estimates were higher in surveys with lower response rates, (p=0.003 in women, 0.018 in men). Conclusions: Population-based surveys that estimate chlamydia prevalence are at risk of participation bias owing to low response rates. Estimates obtained in nationally representative samples of the general population of EU/EEA Member States are similar to estimates from other high income countries.

Effect of coinfection with STDs and of STD treatment on HIV shedding in genital-tract secretions - Systematic review and data synthesis

Objective: To determine whether coinfection with sexually transmitted diseases (STD) increases HIV shedding in genital-tract secretions, and whether STD treatment reduces this shedding. Design: Systematic review and data synthesis of cross-sectional and cohort studies meeting. predefined quality criteria. Main Outcome Measures: Proportion of patients with and without a STD who had detectable HIV in genital secretions, HIV toad in genital secretions, or change following STD treatment. Results: Of 48 identified studies, three cross-sectional and three cohort studies were included. HIV was detected significantly more frequently in participants infected with Neisseria gonorrhoeae (125 of 309 participants, 41%) than in those without N gonorrhoeae infection (311 of 988 participants, 32%; P = 0.004). HIV was not significantly more frequently detected in persons infected with Chlamydia trachomatis (28 of 67 participants, 42%) than in those without C trachomatis infection (375 of 1149 participants, 33%; P = 0.13). Median HIV load reported in only one study was greater in men with urethritis (12.4 x 10(4) versus 1.51 x 10(4) copies/ml; P = 0.04). In the only cohort study in which this could be fully assessed, treatment of women with any STD reduced the proportion of those with detectable HIV from 39% to 29% (P = 0.05), whereas this proportion remained stable among controls (15-17%), A second cohort study reported fully on HIV load; among men with urethritis, viral load fell from 12.4 to 4.12 x 10(4) copies/ml 2 weeks posttreatment, whereas viral load remained stable in those without urethritis. Conclusion: Few high-quality studies were found. HIV is detected moderately more frequently in genital secretions of men and women with a STD, and HIV load is substantially increased among men with urethritis, Successful STD treatment reduces both of these parameters, but not to control levels. More high-quality studies are needed to explore this important relationship further.

Trichomonas vaginalis is associated with pelvic inflammatory disease in women infected with human immunodeficiency virus

We assessed the association between the causative agents of vaginal discharge and pelvic inflammatory disease (PID) among women attending a rural sexually transmitted disease clinic in South Africa; the role played by coinfection with human immunodeficiency virus type 1 (HIV-1) was studied. Vaginal and cervical specimens were obtained to detect Neisseria gonorrhoeae, Chlamydia trachomatis, Trichomonas vaginalis, and bacterial vaginosis. HIV-1 infection was established by use of serum antibody tests. A total of 696 women with vaginal discharge were recruited, 119 of whom had clinical PID. Patients with trichomoniasis had a significantly higher risk of PID than did women without trichomoniasis (P = .03). PID was not associated with any of the other pathogens. When the patients were stratified according to HIV-1 status, the risk of PID in HIV-1-infected patients with T. vaginalis increased significantly (P = .002); no association was found in patients without HIV-1. T. vaginalis infection of the lower genital tract is associated with a clinical diagnosis of PID in HIV-1-infected women.

Association between HIV-1 infection, the etiology of genital ulcer disease, and response to syndromic management

Background: Reports on the effect of HIV-1 infection on healing rates of ulcers are conflicting. Goal: The goal was to determine the etiology and response to treatment of genital ulcer disease (GUD) in relation to HIV-1 infection. Study Design: This was a cohort study of patients with GUD treated with local syndromic management protocols. Results: Among the 587 recruited, the prevalences of infections due to HSV, Treponema pallidum, Chlamydia trachomatis (lymphogranuloma venereum [LGV]), Haemophilus ducreyi, Calymmatobacterium granulomatis, and HIV-1 were 48%, 14%, 11%, 10%, 1%, and 75%, respectively. The prevalence of T pallidum was higher among men (P = 0.03), and an association was seen among HIV-1-seronegatives on univariate and multivariate analyses (P < 0.001; P = 0.01). The prevalence of C trachomatis (LGV) was higher among females (P = 0.004), and an association was seen among HIV-1-seropositives on univariate analysis (P = 0.04). At follow-up, 40/407 (10%) showed a decreased healing tendency, not associated with ulcer etiology or HIV-1 seropositivity. Conclusion: Response to syndromic management of GUD was acceptable and not associated with HIV-1 coinfection.

Bacterial triggers and autoimmune rheumatic diseases

Autoimmune rheumatic diseases are generally considered as a multifactorial aetiology, mainly genetic susceptibility combined with environmental triggers of which bacteria are considered one of the most prominent. Among the rheumatic diseases where bacterial agents are more clearly involved as triggers are: reactive arthritis (ReA), rheumatic fever (RF) and Lyme disease. The role of bacterial infections in inducing other seronegative spondyloarthritis and antiphospholipid antibody syndrome has been hypothesized but is still not proven. The classic form of ReA is associated with the presence of HLA-B27 and is triggered by the urethritis or enteritis causing pathogens Chlamydia trachomatis and the enterobacteria Salmonella, Shigella, and Yersinia, respectively. But several other pathogens such as Brucella, Leptospira, Mycobacteria, Neisseria, Staphylococcus and Streptococcus have also been reported to cause ReA. RF is due to an autoimmune reaction triggered by an untreated throat infection by Streptococcus pyogenes in susceptible individuals. Carditis is the most serious manifestation of RF and HLA-DR7 is predominantly observed in the development of valvular lesions. Lyme disease is a tick-transmitted disease caused by the spirochete Borrelia burgdorferi. Knowledge is limited about how this spirochete interacts with human tissues and cells. Some data report that Borrelia burgdorferi can manipulate resident cells towards a pro- but also anti-inflammatory reaction and persist over a long period of time inside the human body or even inside human cells.