Chlamydia

Background: Ocular adnexal lymphomas may be antigendriven disorders; however, the source of the putative antigen or antigens is still unknown. Hence, we assessed whether Chlamydiae infection is associated with the development of ocular adnexal lymphomas. Methods: The presence of Chlamydia psittaci, trachomatis, and pneumoniae DNA was investigated by polymerase chain reaction in 40 ocular adnexal lymphoma samples, 20 nonneoplastic orbital biopsies, 26 reactive lymphadenopathy samples, and peripheral blood mononuclear cells (PBMCs) from 21 lymphoma patients and 38 healthy individuals. Seven patients with chlamydiapositive PBMCs were treated with the antibiotic doxycycline, and objective response was assessed in four patients with measurable lymphoma lesions. Differences in Chlamydiae DNA detection between the case patients and the control subjects were analyzed using the Fisher exact test. All statistical tests were two-sided. Results: Thirty-two of the 40 (80%) ocular adnexal lymphoma samples carried C. psittaci DNA, whereas all lymphoma samples were negative for C. trachomatis and C. pneumoniae. In contrast, none of the 20 nonneoplastic orbital biopsies (0% versus 80%; P<.001) and only three of 26 (12%) reactive lymphadenopathy samples (12% versus 80%; P<.001) carried the C. psittaci DNA. Nine of 21 (43%) patients with chlamydia-positive lymphomas carried C. psittaci DNA in their PBMCs, whereas none (0%) of the healthy PBMC donors carried C. psittaci DNA in their PBMCs (43% versus 0%; P<.001). One month after doxycycline treatment, chlamydial DNA was no longer detectable in the PBMCs of all seven treated patients, and objective response was observed in two of the four evaluable patients. Conclusion: Patients with ocular adnexal lymphoma had a high prevalence of C. psittaci infection in both tumor tissue and PBMCs. Persistent C. psittaci infection may contribute to the development of these lymphomas, as was also supported by the clinical responses observed in this study with C. psittaci-eradicating antibiotic therapy. [J Natl Cancer Inst 2004;96:586 -94]

Our understanding of both membrane traffic in mammalian cells and the cell biology of infection with intracellular pathogens has increased dramatically in recent years. In this review, we discuss the cell biology of the host-microbe interaction for four intracellular pathogens: Chlamydia spp., Legionella pneumophila, Mycobacterium spp., and the protozoan parasite Toxoplasma gondii. All of these organisms reside in vacuoles inside cells that have restricted fusion with host organelles of the endocytic cascade. Despite this restricted fusion, the vacuoles surrounding each pathogen display novel interactions with other host cell organelles. In addition to the effect of infection on host membrane traffic, we focus on these novel interactions and relate them where possible to nutrient acquisition by the intracellular organisms.

The continuous presence of bacteria or their degraded antigens in the synovium may be involved in the pathogenesis of rheumatoid arthritis (RA). The aim of this study was to determine the presence of bacterial nucleic acids and bacterial cell wall constituents in the joints of patients with RA and other forms of arthritis. Joint samples were obtained from patients with RA (n = 26), septic arthritis (n = 2), inflammatory osteoarthritis (n = 5), and gout (n = 6), and joint trauma (n = 1). Universal 16S-ribosomal RNA primers were used to detect the presence of bacterial DNA in these samples, using stringent regimens for sample collection and molecular microbiologic analysis. Automated sequencing and comparative data analysis were performed to identify the species. The presence of bacterial peptidoglycan-polysaccharide complexes in synovial tissue was detected by immunohistologic analysis with a specific antibody. The bacterial species cultured from the synovium could be identified in both of the patients with septic arthritis. DNA amplicons were also detected in the synovial fluid and/or tissue samples from 5 patients with RA and 2 patients with crystal-induced arthritis; these originated from multiple bacterial species. Staining for peptidoglycan-polysaccharide complexes was positive in the synovial tissue of both patients with septic arthritis, 16 with RA, 4 with inflammatory osteoarthritis, 4 with crystal-induced arthropathy, and 1 with joint trauma. The staining was mainly found in cells in the synovial sublining, including macrophages. The results indicate that bacterial DNA and bacterial cell wall constituents are retained in the joints of some patients with arthritis, where they might enhance synovial inflammation.

Chlamydia pneumoniae encodes a functional arginine decarboxylase (ArgDC), AaxB, that activates upon self-cleavage and converts L-arginine to agmatine. In contrast, most Chlamydia trachomatis serovars carry a missense or nonsense mutation in aaxB abrogating activity. The G115R missense mutation was not predicted to impact AaxB functionality, making it unclear whether AaxB variations in other Chlamydia species also result in enzyme inactivation. To address the impact of gene polymorphism on functionality, we investigated the activity and production of the Chlamydia AaxB variants. Because ArgDC plays a critical role in the Escherichia coli acid stress response, we studied the ability of these Chlamydia variants to complement an E. coli ArgDC mutant in an acid shock assay. Active AaxB was detected in four additional species: Chlamydia caviae, Chlamydia pecorum, Chlamydia psittaci, and Chlamydia muridarum. Of the C. trachomatis serovars, only E appears to encode active enzyme. To determine when functional enzyme is present during the chlamydial developmental cycle, we utilized an anti-AaxB antibody to detect both uncleaved and cleaved enzyme throughout infection. Uncleaved enzyme production peaked around 20 h postinfection, with optimal cleavage around 44 h. While the role ArgDC plays in Chlamydia survival or virulence is unclear, our data suggest a nichespecific function.

Estimation of the abundance of an uncultured http://aem.asm.org/content/62/10/3787 Updated information and services can be found at: These include: CONTENT ALERTS more» cite this article), Receive: RSS Feeds, eTOCs, free email alerts (when new articles http://journals.asm.org/site/misc/reprints.xhtml Information about commercial reprint orders: http://journals.asm.org/site/subscriptions/ To subscribe to to another ASM Journal go to: on September 30, 2014 by guest http://aem.asm.org/ Downloaded from on September 30, 2014 by guest

Chlamydiae are obligate intracellular pathogens that spend their entire growth phase sequestered in a membrane-bound vacuole called an inclusion. A set of chlamydial proteins, labelled Inc proteins, has been identi®ed in the inclusion membrane (IM). The predicted IncA, IncB and IncC amino acid sequences share very limited similarity, but a common hydrophobicity motif is present within each Inc protein. In an effort to identify a relatively complete catalogue of Chlamydia trachomatis proteins present in the IM of infected cells, we have screened the genome for open reading frames encoding this structural motif. Hydropathy plot analysis was used to screen each translated open reading frame in the C. trachomatis genome database. Forty-six candidate IM proteins (C-Incs) that satis®ed the criteria of containing a bilobed hydrophobic domain of at least 50 amino acids were identi-®ed. The genome of Chlamydia pneumoniae encodes a larger collection of C-Inc proteins, and only approximately half of the C-Incs are encoded within both genomes. In order to con®rm the hydropathy plot screening method as a valid predictor of C-Incs, antisera and/or monoclonal antibodies were prepared against six of the C. trachomatis C-Incs. Immuno¯uorescence microscopy of C. trachomatis-infected cells probed with these antibodies showed that ®ve out of six C-Incs are present in the chlamydial IM. Antisera were also produced against C. pneumoniae p186, a protein sharing identity with Chlamydia psittaci IncA and carrying a similar bilobed hydrophobic domain. These antisera labelled the inclusion membrane in C. pneumoniaeinfected cells, con®rming that proteins sharing the unique secondary structural characteristic also localize to the inclusion membrane of C. pneumoniae. Sera from patients with high-titre antibodies to C. trachomatis were examined for reactivity with each tested C-Inc protein. Three out of six tested C-Incs were recognized by a majority of these patient sera. Collectively, these studies identify and characterize novel proteins localized to the chlamydial IM and demonstrate the existence of a potential secondary structural targeting motif for localization of chlamydial proteins to this unique intracellular environment.

Background -The prevalence of microorganisms causing community-acquired pneumonia in patients who required admission to hospital was investigated and the percentage of cases whose aetiology remained unknown due to the study design and logistical problems estimated. Methods -Between January 1991 and April 1993 all patients with community-acquired pneumonia admitted to six hospitals were included in the study. Aetiological diagnosis, categorised as definite, probable and possible, was based on the results of routine microbiological and serological tests.

Chlamydia felis is an important ocular pathogen in cats worldwide. A multilocus variable-number tandem-repeat analysis (MLVA) system for the detection of tandem repeats across the whole genome of C. felis strain Fe/C-56 was developed. Nine selected genetic loci were tested by MLVA in 17 C. felis isolates, including the C. felis Baker vaccine strain, and 122 clinical samples from different geographic origins. Analysis of the results identified 25 distinct C. felis MLVA patterns. In parallel, a recently described multilocus sequence typing scheme for the typing of Chlamydia was applied to 13 clinical samples with 12 different C. felis MLVA patterns. Rare sequence differences were observed. Thus, the newly developed MLVA system provides a highly sensitive high-resolution test for the differentiation of C. felis isolates from different origins that is suitable for molecular epidemiological studies.

Gene order in bacteria is poorly conserved during evolution 1-3 . For example, although many homologous genes are shared by the proteobacteria Escherichia coli, Haemophilus influenzae and Helicobacter pylori, their relative positions are very different in each genome, except local functional clusters such as operons 3-6 . The complete sequences of the more closely related bacterial genomes, such as pairs of Chlamydia 7-9 , H. pylori 10,11 and Mycobacterium species 12 , now allow identification of the processes and mechanisms involved in genome evolution. Here we provide evidence that a substantial proportion of rearrangements in gene order results from recombination sites that are determined by the positions of the replication forks. Our observations suggest that replication has a major role in directing genome evolution. letter nature genetics • volume 26 • october 2000 195

| The chlamydiae are important obligate intracellular prokaryotic pathogens that, each year, are responsible for millions of human infections involving the eye, genital tract, respiratory tract, vasculature and joints. The chlamydiae grow in cytoplasmic vesicles in susceptible host cells, which include the mucosal epithelium, vascular endothelium, smooth muscle cells, circulating monocytes and recruited or tissue-specific macrophages. One important pathogenic strategy that chlamydiae have evolved to promote their survival is the modulation of programmed cell death pathways in infected host cells. The chlamydiae can elicit the induction of host cell death, or apoptosis, under some circumstances and actively inhibit apoptosis under others. This subtle pathogenic mechanism highlights the manner in which these highly successful pathogens take control of infected cells to promote their own survival -even under the most adverse circumstances.

Chlamydiae are evolutionarily well-separated bacteria that live exclusively within eukaryotic host cells. They include important human pathogens such as Chlamydia trachomatis as well as symbionts of protozoa. As these bacteria are experimentally challenging and genetically intractable, our knowledge about them is still limited. In this study, we obtained the genome sequences of Simkania negevensis Z, Waddlia chondrophila 2032/99, and Parachlamydia acanthamoebae UV-7. This enabled us to perform the first comprehensive comparative and phylogenomic analysis of representative members of four major families of the Chlamydiae, including the Chlamydiaceae. We identified a surprisingly large core gene set present in all genomes and a high number of diverse accessory genes in those Chlamydiae that do not primarily infect humans or animals, including a chemosensory system in P. acanthamoebae and a type IV secretion system. In S. negevensis, the type IV secretion system is encoded on a large conjugative plasmid (pSn, 132 kb). Phylogenetic analyses suggested that a plasmid similar to the S. negevensis plasmid was originally acquired by the last common ancestor of all four families and that it was subsequently reduced, integrated into the chromosome, or lost during diversification, ultimately giving rise to the extant virulence-associated plasmid of pathogenic chlamydiae. Other virulence factors, including a type III secretion system, are conserved among the Chlamydiae to variable degrees and together with differences in the composition of the cell wall reflect adaptation to different host cells including convergent evolution among the four chlamydial families. Phylogenomic analysis focusing on chlamydial proteins with homology to plant proteins provided evidence for the acquisition of 53 chlamydial genes by a plant progenitor, lending further support for the hypothesis of an early interaction between a chlamydial ancestor and the primary photosynthetic eukaryote.

This review summarizes the dramatic changes that have occurred in the taxonomy of bacteria known as Chlamydia. Best known for the diseases they cause in humans, these intracellular bacteria also comprise many species that are responsible for a wide variety of clinically and economically important diseases in livestock and companion animals. The old taxonomy grouped most of these species into C. psittaci because systematic methods for routinely distinguishing them were not available. DNA-based testing methods are now available that distinguish different chlamydial families, genera, and species. This summary reviews these tests and a number of oligonucleotide primers that distinguish these groups using PCR and PCR-RFLP. DNA-based methods are also being used to discover new families of chlamydia-like bacteria, at least one of which is responsible for abortion in cattle (Waddlia chondrophila). This review summarizes the pathogenic roles of the Chlamydiaceae, new families, and individual species within the order Chlamydiales. These discoveries create opportunities for veterinarians to carry out epidemiological studies of chlamydiae that previously were not possible.

Free-living amoebae are increasingly being recognized to serve as vehicles of dispersal for various bacterial human pathogens and as hosts for a variety of obligate bacterial endocytobionts. Several Chlamydia-like Acanthamoeba endocytobionts constituting the recently proposed family Parachlamydiaceae are of special interest as potential human pathogens. In this study coccoid bacterial endocytobionts of a Hartmannella vermiformis isolate were analysed. Infection of H. vermiformis with these bacteria resulted in prevention of cyst formation and subsequent host-cell lysis. Transfection experiments demonstrated that the parasites were not capable of propagating within other closely related free-living amoebae but were able to infect the distantly related species Dictyostelium discoideum. Electron microscopy of the parasites revealed typical morphological characteristics of the Chlamydiales, including the existence of a Chlamydia-like life-cycle, but indicated that these endocytobionts, in contrast to Chlamydia species, do not reside within a vacuole. Comparative 16S rRNA sequence analysis showed that the endocytobiont of H. vermiformis , classified as Neochlamydia hartmannellae gen. nov., sp. nov., is affiliated to the family Parachlamydiaceae. Confocal laser scanning microscopy in combination with fluorescence in situ hybridization using rRNA-targeted oligonucleotide probes confirmed the intracellular localization of the parasites and demonstrated the absence of other bacterial species within the Hartmannella host. These findings extend our knowledge of the phylogenetic diversity of the Parachlamydiaceae and demonstrate for the first time that these endocytobionts can naturally develop within amoebae of the genus Hartmannella.

The optimal methods for the diagnosis of pharyngeal Neisseria gonorrhoeae infection are uncertain. The objective of this study was to define the performance of culture and nucleic acid amplification tests (NAATs) for the diagnosis of pharyngeal N. gonorrhoeae. In this cross-sectional study, males and females >15 years old who acknowledged performing fellatio or cunnilingus (in the previous 2 months) were recruited from three clinics (two human immunodeficiency virus clinics and one sexually transmitted diseases clinic) located in Birmingham, AL. The test performance of culture for N. gonorrhoeae, the Gen-Probe Aptima Combo 2 transcription-mediated amplification assay (TMA), the BD ProbeTec ET amplified DNA strand displacement assay (SDA), and the Roche Cobas Amplicor PCR was defined by using a rotating "gold standard" of any positive results by two or three of the three tests that excluded the test being evaluated. A total of 961 evaluable test sets were collected. On the basis of a rotating gold standard of positive results by two of three comparator tests, the sensitivity and the specificity were as follows: culture for N. gonorrhoeae, 50.0% and 99.4%, respectively; PCR, 80.3% and 73.0%, respectively; TMA, 83.6% and 98.6%, respectively; and SDA, 93.2% and 96.3%, respectively. On the basis of a rotating gold standard of positive results by three of three comparator tests, the sensitivity and specificity were as follows: culture for N. gonorrhoeae, 65.4% and 99.0%, respectively; PCR, 91.9% and 71.8%, respectively; TMA, 100% and 96.2%, respectively; and SDA, 97.1% and 94.2%, respectively. In conclusion, currently available NAATs are more sensitive than culture for the detection of pharyngeal gonorrhea in at-risk patients. PCR is substantially less specific than culture, TMA, or SDA and should not be used for the detection of pharyngeal gonorrhea.

DNA sequences coding for 81% of the ompA gene from 24 chlamydial strains, representing all chlamydial species, were determined from DNA amplified by polymerase chain reactions. Chlamydial strains of serovars and strains with similar chromosomal restriction fragment length polymorphism had identical ompA DNA sequences. The ompA sequences were segregated into 23 different ompA alleles and aligned with each other, and phylogenetic relationships among them were inferred by neighbor-joining and maximum parsimony analyses. The neighbor-joining method produced a single phylogram which was rooted at the branch between two major clusters. One cluster included all Chlamydia trachomatis ompA alleles (trachoma group). The second cluster was composed of three major groups of ompA alleles: psittacosis group (alleles MN, 6BC, A22/M, B577, LW508, FEPN, and GPIC), pneumonia group (Chlamydia pneumoniae AR388 with the allele KOALA), and polyarthritis group (ruminant and porcine chlamydial alleles LW61...

Polymorphic membrane protein (Pmp)21 otherwise known as PmpD is the longest of 21 Pmps expressed by Chlamydophila pneumoniae. Recent bioinformatical analyses annotated PmpD as belonging to a family of exported Gram-negative bacterial proteins designated autotransporters. This prediction, however, was never experimentally supported, nor was the function of PmpD known. Here, using 1D and 2D PAGE we demonstrate that PmpD is processed into two parts, N-terminal (N-pmpD), middle (M-pmpD) and presumably third, C-terminal part (C-pmpD). Based on localization of the external part on the outer membrane as shown by immunofluorescence, immuno-electron microscopy and immunoblotting combined with trypsinization, we demonstrate that N-pmpD translocates to the surface of bacteria where it noncovalently binds other components of the outer membrane. We propose that N-pmpD functions as an adhesin, as antibodies raised against N-pmpD blocked chlamydial infectivity in the epithelial cells. In addition, recombinant N-pmpD activated human monocytes in vitro by upregulating their metabolic activity and by stimulating IL-8 release in a dosedependent manner. These results demonstrate that N-PmpD is an autotransporter component of chlamydial outer membrane, important for bacterial invasion and host inflammation.

ATP/ADP translocases catalyze the highly specific transport of ATP across a membrane in an exchange mode with ADP. Such unique transport proteins are employed by plant plastids and have among the prokaryotes so far only been identified in few obligate intracellular bacteria belonging to the Chlamydiales and the Rickettsiales. In this study, 12 phylogenetically diverse bacterial endosymbionts of free-living amoebae and paramecia were screened for the presence of genes encoding ATP/ADP transport proteins. The occurrence of ATP/ADP translocase genes was found to be restricted to endosymbionts related to rickettsiae and chlamydiae. We showed that the ATP/ADP transport protein of the Parachlamydia-related endosymbiont of Acanthamoeba sp. strain UWE25, a recently identified relative of the important human pathogens Chlamydia trachomatis and Chlamydophila pneumoniae, is functional when expressed in the heterologous host Escherichia coli and demonstrated the presence of transcripts during the chlamydial developmental cycle. These findings indicate that the interaction between Parachlamydia-related endosymbionts and their amoeba hosts concerns energy parasitism similar to the interaction between pathogenic chlamydiae and their human host cells. Phylogenetic analysis of all known ATP/ADP translocases indicated that the genes encoding ATP/ADP translocases originated from a chlamydial ancestor and were, after an ancient gene duplication, transferred horizontally to rickettsiae and plants.

To evaluate the type and etiopathogenic role of infectious agents detected in endometrial cultures obtained from women with chronic endometritis (CE). Prospective controlled study. University hospital. 2190 women undergoing hysteroscopy for different indications. Vaginal and endometrial samples were collected from 438 women with a CE diagnosis at hysteroscopy and 100 women with no signs of CE (controls). Histology and cultures for common bacteria, Neisseria gonorrhoeae and Mycoplasma, and molecular biology testing for Chlamydia were performed. We compared results of vaginal and intrauterine cultures obtained from women with and without CE. Histologic results were positive in 388 of these cases (88.6%), and at least one microorganism was found in 320 endometrial samples (73.1%). In the control group, histologic results and endometrial culture were positive in only 6% and 5% of cases, respectively. The most frequent infectious agents detected at the endometrial level were common bacteria (58% of cases). Ureaplasma urealyticum was detected in 10% and Chlamydia in only 2.7% of positive endometrial cultures. In only 143 (32.6%) cases were the same infectious agent isolated in endometrial and vaginal cultures. More than 70% of CE cases resulted from nongonococcal, nonchlamydial infections. Common bacteria and Mycoplasma were the most frequent etiologic agents. Vaginal cultures have low concordance with endometrial cultures.

Immunity to intracellular microbial pathogens, including Chlamydia species, is controlled primarily by cellmediated effector mechanisms, yet, the absence of antibodies results in inefficient microbial clearance. We investigated the hypothesis that certain Fc receptor functions promote the rapid induction of elevated T helper type 1 (Th1) response, which effectively clears chlamydiae. FcR Ϫ/Ϫ mice exhibited a delayed and reduced frequency of Chlamydia-specific Th1 cells, compared to FcR +/+ mice. In vitro, antichlamydial antibodies increased the rate of Th1 activation by FcR +/+ but not FcR Ϫ/Ϫ antigen-presenting cells. FcR Ϫ/Ϫ dendritic cells and the T cell-associated IgG2A and IgA mediate enhanced Th1 activation by antibodies. Immunization with chlamydia-antibody complexes induced elevated and protective Th1 response. These results provide a mechanistic basis for requiring both T cell and humoral immune responses in protective immunity and vaccine evaluation. Findings offer a paradigm in host defense wherein different effector components function indirectly to maximize the principal effector mechanism.

The family Chlamydiaceae (order Chlamydiales, phylum Chlamydiae) comprises important, obligate intracellular bacterial pathogens of humans and animals. Subdivision of the family into the two genera Chlamydia and Chlamydophila has been discussed controversially during the past decade. Here, we have revisited the current classification in the light of recent genomic data and in the context of the unique biological properties of these microorganisms. We conclude that neither generally used 16S rRNA sequence identity cut-off values nor parameters based on genomic similarity consistently separate the two genera. Notably, no easily recognizable phenotype such as host preference or tissue tropism is available that would support a subdivision. In addition, the genus Chlamydophila is currently not well accepted and not used by a majority of research groups in the field. Therefore, we propose the classification of all 11 currently recognized Chlamydiaceae species in a single genus, the genus Chlamydia. Finally, we provide emended descriptions of the family Chlamydiaceae, the genus Chlamydia, as well as the species Chlamydia abortus, Chlamydia caviae and Chlamydia felis.

Gram-negative bacteria secrete a variety of proteins to the cell surface and beyond, a process with many inherent difficulties. An exceptionally widespread answer to these problems is the type V (or autotransporter) secretion pathway. By exploiting the data made available by bacterial genome sequencing, we have discovered that the previously described polymorphic proteins of Chlamydia spp. resemble members of the autotransporter family, and we suggest that they follow the same secretion pathway. Research Update Research Update Research Update their polymorphisms, surface exposure, antigenicity and phase-variable nature, it is reasonable to assume they play a role in the virulence of the Chlamydiales.

An unusually high proportion of proteins encoded in Chlamydia genomes are most similar to plant proteins, leading to proposals that a Chlamydia ancestor obtained genes from a plant or plant-like host organism by horizontal gene transfer. However, during an analysis of bacterial-eukaryotic protein similarities, we found that the vast majority of plant-like sequences in Chlamydia are most similar to plant proteins that are targeted to the chloroplast, an organelle derived from a cyanobacterium. We present further evidence suggesting that plant-like genes in Chlamydia, and other Chlamydiaceae, are likely a reflection of an unappreciated evolutionary relationship between the Chlamydiaceae and the cyanobacteria-chloroplast lineage. Further analyses of bacterial and eukaryotic genomes indicates the importance of evaluating organellar ancestry of eukaryotic proteins when identifying bacteria-eukaryote homologs or horizontal gene transfer and supports the proposal that Chlamydiaceae, which are obligate intracellular bacterial pathogens of animals, are not likely exchanging DNA with their hosts.

Cell surface heparan sulfate (HS), a polysaccharide composed of alternating uronic acid and glucosamine residues, represents a common link that many sexually transmitted infections (STIs) require for infection. Variable modifications within the monomeric units of HS chains together with their unique structural conformations generate heterogeneity, which expands the ability of HS to bind a diverse array of host and microbial proteins. Recent advances made in the field of glycobiology have critically enhanced our understanding of HS and its interactions with microbes and their significance in important human diseases. The role of HS has been elaborated for several STIs to include those caused by herpes simplex virus, human immunodeficiency virus, human papillomavirus, and Chlamydia. In addition, gonorrhea, syphilis, and yeast infections are also dependent on the presence of HS on human target cells. Critical steps such as pathogen adhesion or binding to host cells followed by internalization to enhance intracellular survival and possible spread to other cells are mediated by HS. In addition, HS guided cell signaling plays a role in the development of angiogenesis and inflammation associated with many STIs. Past and ongoing investigations are providing new push for the development of HS-mimetics and analogs as novel prevention strategies against many different STIs. This review article summarizes the significance of HS in STIs and describes how emerging new products that target HS can be used to control the spread of STIs.

Chlamydia species are the causative agents of trachoma, various forms of pneumonia, and the most common sexually transmitted diseases. Although the infection cycle has been extensively characterized in epithelial cells, where the Chlamydia entry-vacuoles avoid fusion with host-cell lysosomes, the cellular immune response has received less attention. Moreover, despite the abundant presence of dendritic cells (DC) in the sites of

Epidemiological data indicate that infection of cattle with chlamydiae such as Chlamydophila (C.) pecorum, C. abortus, C. psittaci and Chlamydia suis, is ubiquitous with mixed infections occurring frequently. The apparent lack of association between infection and clinical disease has resulted in debate as to the pathogenic significance of these organisms, and their tendency to sub-clinical and/or persistent infection presents a challenge to the study of their potential effects. However, recent evidence indicates that chlamydial infections have a substantial and quantifiable impact on livestock productivity with chronic, recurrent infections associated with pulmonary disease in calves and with infertility and sub-clinical mastitis in dairy cows. Data also suggest these infections manifest clinically when they coincide with a number of epidemiological risk factors. Future research should: (1) use relevant animal models to clarify the pathogenesis of bovine chlamydioses; (2) quantify the impact of chlamydial infection at a herd level and identify strategies for its control, including sub-unit vaccine development; and (3) evaluate the zoonotic risk of bovine chlamydial infections which will require the development of species-specific serodiagnostics.

Obligate intracellular bacterial pathogens of the genus Chlamydia are reported to enter host cells by both clathrin-dependent and clathrin-independent processes. C. trachomatis serovar K recently was shown to enter cells via caveolae-like lipid raft domains. We asked here how widespread raft-mediated entry might be among the Chlamydia. We show that C. pneumoniae, an important cause of respiratory infections in humans that additionally is associated with cardiovascular disease, and C. psittaci, an important pathogen in domestic mammals and birds that also infects humans, each enter host cells via cholesterol-rich lipid raft microdomains. Further, we show that C. trachomatis serovars E and F also use these domains to enter host cells. The involvement of these membrane domains in the entry of these organisms was indicated by the sensitivity of their entry to the raft-disrupting agents Nystatin and filipin, and by their intracellular association with caveolin-1, a 22-kDa protein associated with the formation of caveolae in rafts. In contrast, caveolin-marked lipid raft domains do not mediate entry of C. trachomatis serovars A, 36B, and C, nor of LGV serovar L2 and MoPn. Finally, we show that entry of each of these chlamydial strains is independent of cellular expression of caveolin-1. Thus, entry via the Nystatin and filipin-sensitive pathway is dependent on lipid rafts containing cholesterol, rather than invaginated caveolae per se.

Serology is commonly used for the diagnosis of acute Chlamydia pneumoniae infections and also for the diagnosis of complicated Chlamydia trachomatis infections. Furthermore, recent sero-epidemiological studies have linked C. pneumoniae infection with several diseases traditionally considered non-infectious. The objectives of this mini-review are to critically review and discuss some selected analytical and methodological aspects, controversies and current problems in chlamydial serodiagnosis. To illustrate our views we present some original data of the comparison of current technologies. The review of the literature revealed high variability in methodologies applied to different studies. This observation was supported by our own data, which explains occasional conflicting clinical interpretation. Although the microimmunofluorescence (MIF) technique is generally considered as the gold standard for serodiagnosis of chlamydial infections, assay conditions are highly variable and hence pose a major problem in the interpretation of the results. For instance, many recent studies linking C. pneumoniae and atherosclerosis have utilized MIF techniques with variable threshold criteria for the positivity, in combination with selection bias of cases and controls possibly leading to conflicting results. Variability of assay conditions is also a common problem with Western blots, and interpretation is problematic when both anti-C. pneumoniae and anti-C. trachomatis antibodies are present. Furthermore, there is a lot of disagreement in serological criteria applied to recently emerged enzyme immunoassay (EIA) techniques when these assays are used for acute and non-acute clinical conditions and their association with Chlamydiae. In conclusion, standardization of serological techniques and the development of uniform criteria for interpretation of serologic findings is necessary to increase our knowledge of the biology of Chlamydiae, pathogenesis of any chlamydial infection and chronic infections in particular.

We report here the identification of a bacterial protein capable of interacting with mammalian death receptors in vitro and in vivo. The protein is encoded in the genome of Chlamydia trachomatis and has homologues in other Chlamydia species. This protein, which we refer to as "Chlamydia protein associating with death domains" (CADD), induces apoptosis in a variety of mammalian cell lines when expressed by transient gene transfection. Apoptosis induction can be blocked by Caspase inhibitors, indicating that CADD triggers cell death by engaging the host apoptotic machinery. CADD interacts with death domains of tumor necrosis factor (TNF) family receptors TNFR1, Fas, DR4, and DR5 but not with the respective downstream adaptors. In infected epithelial cells, CADD is expressed late in the infectious cycle of C. trachomatis and co-localizes with Fas in the proximity of the inclusion body. The results suggest a role for CADD modulating the apoptosis pathways of cells infected, revealing a new mechanism of host-pathogen interaction.

Chlamydiae are obligate intracellular bacteria found as symbionts and pathogens in a wide range of eukaryotes, including protists, invertebrates, and vertebrates. It was recently proposed that an ancient chlamydial symbiont facilitated the establishment of primary plastids in a tripartite symbiosis with cyanobacteria and early eukaryotes. In this review, we summarize recent advances in understanding of the lifestyle and the evolutionary history of extant Chlamydiae. We reconstruct and describe key features of the ancient chlamydial symbiont. We propose that it was already adapted to an intracellular lifestyle before the emergence of Archaeplastida, and that several observations are compatible with an essential contribution of Chlamydiae to the evolution of algae and plants.

Background: Chlamydiaceae are a family of obligate intracellular pathogens causing a wide range of diseases in animals and humans, and facing unique evolutionary constraints not encountered by free-living prokaryotes. To investigate genomic aspects of infection, virulence and host preference we have sequenced Chlamydia psittaci, the pathogenic agent of ornithosis. Results: A comparison of the genome of the avian Chlamydia psittaci isolate 6BC with the genomes of other chlamydial species, C. trachomatis, C. muridarum, C. pneumoniae, C. abortus, C. felis and C. caviae, revealed a high level of sequence conservation and synteny across taxa, with the major exception of the human pathogen C. trachomatis. Important differences manifest in the polymorphic membrane protein family specific for the Chlamydiae and in the highly variable chlamydial plasticity zone. We identified a number of psittaci-specific polymorphic membrane proteins of the G family that may be related to differences in host-range and/or virulence as compared to closely related Chlamydiaceae. We calculated non-synonymous to synonymous substitution rate ratios for pairs of orthologous genes to identify putative targets of adaptive evolution and predicted type III secreted effector proteins. Conclusions: This study is the first detailed analysis of the Chlamydia psittaci genome sequence. It provides insights in the genome architecture of C. psittaci and proposes a number of novel candidate genes mostly of yet unknown function that may be important for pathogen-host interactions.

Growing evidence suggests that a novel member of the Chlamydiales order, Waddlia chondrophila, is a potential agent of miscarriage in humans and abortion in ruminants. Due to the lack of genetic tools to manipulate chlamydia, genomic analysis is proving to be the most incisive tool in stimulating investigations into the biology of these obligate intracellular bacteria. 454/Roche and Solexa/Illumina technologies were thus used to sequence and assemble de novo the full genome of the first representative of the Waddliaceae family, W. chondrophila. The bacteria possesses a 291169312bp chromosome and a 159593 bp low-copy number plasmid that might integrate into the bacterial chromosome. The Waddlia genome displays numerous repeated sequences indicating different genome dynamics from classical chlamydia which almost completely lack repetitive elements. Moreover, W. chondrophila exhibits many virulence factors also present in classical chlamydia, including a functional type III secretion system, but also a large complement of specific factors for resistance to host or environmental stresses. Large families of outer membrane proteins were identified indicating that these highly immunogenic proteins are not Chlamydiaceae specific and might have been present in their last common ancestor. Enhanced metabolic capability for the synthesis of nucleotides, amino acids, lipids and other co-factors suggests that the common ancestor of the modern Chlamydiales may have been less dependent on their eukaryotic host. The fine-detailed analysis of biosynthetic pathways brings us closer to possibly developing a synthetic medium to grow W. chondrophila, a critical step in the development of genetic tools. As a whole, the availability of the W. chondrophila genome opens new possibilities in Chlamydiales research, providing new insights into the evolution of members of the order Chlamydiales and the biology of the Waddliaceae.

Background: Chlamydiae are obligate intracellular bacteria that multiply in a vacuolar compartment, the inclusion. Several chlamydial proteins containing a bilobal hydrophobic domain are translocated by a type III secretion (TTS) mechanism into the inclusion membrane. They form the family of Inc proteins, which is specific to this phylum. Based on their localization, Inc proteins likely play important roles in the interactions between the microbe and the host. In this paper we sought to identify and analyze, using bioinformatics tools, all putative Inc proteins in published chlamydial genomes, including an environmental species. Results: Inc proteins contain at least one bilobal hydrophobic domain made of two transmembrane helices separated by a loop of less than 30 amino acids. Using bioinformatics tools we identified 537 putative Inc proteins across seven chlamydial proteomes. The amino-terminal segment of the putative Inc proteins was recognized as a functional TTS signal in 90% of the C. trachomatis and C. pneumoniae sequences tested, validating the data obtained in silico. We identified a macro domain in several putative Inc proteins, and observed that Inc proteins are enriched in segments predicted to form coiled coils. A surprisingly large proportion of the putative Inc proteins are not constitutively translocated to the inclusion membrane in culture conditions.

There are increasing concerns about treatment failure following treatment for rectal chlamydia with 1 g of azithromycin. A systematic review and meta-analysis was conducted to investigate the efficacy of 1 g of azithromycin as a single dose or 100 mg of doxycycline twice daily for 7 days for the treatment of rectal chlamydia. Medline, Embase, PubMed, Cochrane Controlled Trials Register, Australia New Zealand Clinical Trial Register and ClinicalTrials.gov were searched to the end of April 2014. Studies using 1 g of azithromycin or 7 days of doxycycline for the treatment of rectal chlamydia were eligible. Gender, diagnostic test, serovar, symptomatic status, other sexually transmitted infections, follow-up time, attrition and microbial cure were extracted. Meta-analysis was used to calculate pooled (i) azithromycin and doxycycline efficacy and (ii) efficacy difference. All eight included studies were observational. The random-effects pooled efficacy for azithromycin (based on eight st...

In order to exert their antibacterial activity, antibiotics exhibits a peculiar behaviour: it has a very long halflife of about 50 h accompanied by very low serum must reach the infectious focus. Infection may develop in the interstitial spaces of tissues or inside the cells. levels and extremely high tissue and intracellular levels, particularly in PMNs, alveolar macrophages and The physicochemical properties of drugs are the main factors conditioning their distribution in body tissues fibroblasts, from which it is slowly released. The C/E ratio in alveolar macrophages is about 300 after 48 h, and their penetration into cells. For the treatment of respiratory infections antibiotics must reach adequate but values may increase to >1000 3-4 days following administration, because of the extremely low serum concentrations in lung parenchyma, bronchial tissues, bronchial secretions, and cells. The behaviour of all levels (Ζ0.01 g/ml). 12-16 Correlation between intracellular accumulation and classes of antibiotics in the respiratory tract has been thoroughly investigated. 1 A recently developed micro-antimicrobial activity has been submitted to careful analysis. The presence of large amounts of antibiotics lavage technique, performed via fibreoptic bronchoscopy, allows the sampling of alveolar epithelial within cells does not always ensure higher antibacterial activity. Cell penetration of macrolides correlates with lining fluid and alveolar macrophages in order to determine antibiotic concentrations following sys-extracellular concentration and antimicrobial activity correlates with intracellular accumulation in ex-temic administration in humans. 2 Information regarding the pharmacokinetic behaviour of antibiotics perimental models. Fluoroquinolones, which accumulate in cells to a lesser extent than macrolides, in the respiratory tract has been of great importance in anticipating and assessing their clinical efficacy in nevertheless show an excellent antimicrobial activity, while clindamycin, which shows a C/E ratio somewhat respiratory infections. Clindamycin, rifampicin, tetracyclines, macrolides superior to fluoroquinolones, displays low activity against intracellular Staphylococcus aureus. 5 Evidently and fluoroquinolones accumulate in phagocytes. 3,4 Macrolides show the greatest capacity for intracellular the nature of the parasite, its growth rate, the different experimental conditions and the mechanism of action accumulation, localizing within the cytosol and lysosomes. In polymorphonuclear neutrophils (PMN), of the antibiotic can greatly influence the outcome of experiments performed in vitro and in animal models. the cellular/extracellular (C/E) ratio varies from 2 to 14 for erythromycin and josamycin and reach the Atypical pneumonia is a clinical syndrome without a precise definition, first described by Reimann in highest values for dirithromycin, clarithromycin and azithromycin. 5-10 In alveolar macrophages the C/E 1938, 17 who realised that some patients with pneumonia had clinical picture and natural history that ratio of dirithromycin is about 80, that of clarithromycin can range according to the time of sample differ from that seen in patients with pneumococcal infection. The syndrome includes a subacute onset collection from about 40 to >100. 10,11 Azithromycin with low-moderate grade fever, dry cough, prominent constitutional symptoms, absent or moderate leuko

Bovine vaginal cytobrush specimens were analyzed for the presence of Chlamydia spp. by a high-sensitivity, high-specificity quantitative PCR. The 53% prevalence of low-level Chlamydia psittaci and C. pecorum genital infection detected in virgin heifers suggests predominantely extragenital transmission of Chlamydia in cattle and conforms to the high seroprevalence of anti-Chlamydia antibodies.

A retrospective study on reptile tissues presenting with granulomatous inflammation was performed to detect the possible presence of mycobacteria and chlamydiae in these lesions. Ninety cases including 48 snakes, 27 chelonians, and 15 lizards were selected. Mycobacteria were detected by Ziehl-Neelsen (ZN) staining and a broad-range polymerase chain reaction (PCR) followed by DNA sequencing. To detect chlamydiae, immunohistochemistry with monoclonal antibodies against chlamydial lipopolysaccharide (LPS) and a Chlamydiales order-specific PCR and sequencing were applied. Acid-fast bacilli were found in 14 cases (15.6%) by ZN staining and in 23 cases (25.6%) by PCR. Sequence analysis revealed the presence of Mycobacteria other than Mycobacterium tuberculosis complex (MOTT). Chlamydial LPS antigen was observed within granulomas from five samples (5.6%), whereas the PCR screen revealed 58 positive cases (64.4%). Of these, 9 cases (10%) showed 98-99% similarity to Chlamydophila (Cp.) pneumoniae and 49 cases (54.4%) displayed a high similarity (88-97%) to the newly described &amp;quot;Chlamydia-like&amp;quot; microorganisms Parachlamydia acanthamoebae and Simkania negevensis. Results from this study confirm, on the one hand, that MOTT are probably the most important infectious etiology for granulomatous inflammation in reptiles. On the other hand, they indicate that chlamydia infects reptiles and that Cp. pneumoniae should be considered an etiological agent of granulomatous lesions of reptiles. Because both MOTT and Cp. pneumoniae are human pathogens, the potential of zoonotic transmission from reptiles to humans has to be considered. In contrast, the significance of Chlamydia-like isolates remains completely open, and further studies are needed to evaluate their role.