Anti-chlamydial activities of cell-permeable hydrophobic dipeptide-containing derivatives.

The obligate intracellular bacteria chlamydia is major human pathogen that causes millions of trachoma, sexually transmitted infections and pneumonia worldwide. We serendipitously found that both calpain inhibitors z-Val-Phe-CHO and z-Leu-Nle-CHO showed marked inhibitory activity against chlamydial growth in human epithelial HeLa cells, whereas other calpain inhibitors not. These peptidomimetic inhibitors consist of N-benzyloxycarbonyl group and hydrophobic dipeptide derivatives. Both compounds strongly restrict the chlamydial growth even addition at the 12 h post infection. Notably, inhibitors-mediated growth inhibition of chlamydia was independent on host calpain activity. Electron microscopic analysis revealed that z-Val-Phe-CHO inhibited chlamydial growth by arresting bacterial cell division and RB-EB re-transition, but not by changing into persistent state. We searched and found that z-Leu-Leu-CHO and z-Phe-Ala-FMK also inhibited chlamydial growth. Neither biotin-hydrophobic dipeptide nor morpholinoureidyl-hydrophobic dipeptide shows inhibitory effects on chlamydial intracellular growth. Our results suggested the possibility of some chemical derivatives based on z-hydrophobic dipeptide group for future therapeutic usage to the chlamydial infectious disease.

Chlamydia trachomatis impairs host base excision repair by downregulating polymerase ß.

Chlamydia trachomatis infections have been associated with ovarian cancer by several epidemiological studies. Here, we show that C. trachomatis-infected primary human ovarian epithelial cells display elevated oxidative DNA damage. Base excision repair, an important cellular mechanism to repair oxidative DNA lesions, was impaired in infected primary ovarian and in several other types of cells. Polymerase ß was downregulated in infected cells associated with upregulation of microRNA-499a (miR-499a). Stabilising polymerase ß by inhibiting miR-499a significantly improved repair. Moreover, downregulation of tumour suppressor p53 also resulted in attenuated repair in these cells. Thus, our data show that downregulation of polymerase ß by direct inhibition through miR-499a and downregulation of p53 debilitate the host-cell base excision repair during C. trachomatis infection.

Prevalence of Trichomonas vaginalis by PCR in men attending a primary care urology clinic in South Korea.

Trichomonas vaginalis, a causative agent of trichomoniasis, may trigger symptomatic or asymptomatic nongonococcal urethritis and chronic prostatitis in men. Despite the availability of highly sensitive diagnostic tests, such as nucleic acid amplification tests, including PCR, few prospective studies present data on male T. vaginalis infection in South Korea. In the present study, the prevalence of T. vaginalis and associated clinical conditions were evaluated in 201 male patients from a primary care urology clinic in South Korea. The prevalence of T. vaginalis infection in our cohort was 4% (8/201) by PCR. T. vaginalis infection was common in men older than 40 years (median age, 52 years). Among the 8 Trichomonas-positive patients, 87.5% (7/8) had prostatic diseases, such as prostatitis and benign prostatic hyperplasia, and 25.0% (2/8) and 12.5% (1/8) were coinfected with Chlamydia trachomatis and Mycoplasma genitalium, respectively. Our results suggest that T. vaginalis infection is not rare in men attending primary care urology clinics in South Korea, especially in those older than 40 years, in whom it may explain the presence of prostatic disease. The possibility of T. vaginalis infection should be routinely considered in older male patients with prostatic diseases in South Korea.

Prevalence of Trichomonas vaginalis by PCR in Men Attending a Primary Care Urology Clinic in South Korea

Trichomonas vaginalis, a causative agent of trichomoniasis, may trigger symptomatic or asymptomatic nongonococcal urethritis and chronic prostatitis in men. Despite the availability of highly sensitive diagnostic tests, such as nucleic acid amplification tests, including PCR, few prospective studies present data on male T. vaginalis infection in South Korea. In the present study, the prevalence of T. vaginalis and associated clinical conditions were evaluated in 201 male patients from a primary care urology clinic in South Korea. The prevalence of T. vaginalis infection in our cohort was 4% (8/201) by PCR. T. vaginalis infection was common in men older than 40 years (median age, 52 years). Among the 8 Trichomonas-positive patients, 87.5% (7/8) had prostatic diseases, such as prostatitis and benign prostatic hyperplasia, and 25.0% (2/8) and 12.5% (1/8) were coinfected with Chlamydia trachomatis and Mycoplasma genitalium, respectively. Our results suggest that T. vaginalis infection is not rare in men attending primary care urology clinics in South Korea, especially in those older than 40 years, in whom it may explain the presence of prostatic disease. The possibility of T. vaginalis infection should be routinely considered in older male patients with prostatic diseases in South Korea.

Fierce competition between Toxoplasma and Chlamydia for host cell structures in dually infected cells.

The prokaryote Chlamydia trachomatis and the protozoan Toxoplasma gondii, two obligate intracellular pathogens of humans, have evolved a similar modus operandi to colonize their host cell and salvage nutrients from organelles. In order to gain fundamental knowledge on the pathogenicity of these microorganisms, we have established a cell culture model whereby single fibroblasts are coinfected by C. trachomatis and T. gondii. We previously reported that the two pathogens compete for the same nutrient pools in coinfected cells and that Toxoplasma holds a significant competitive advantage over Chlamydia. Here we have expanded our coinfection studies by examining the respective abilities of Chlamydia and Toxoplasma to co-opt the host cytoskeleton and recruit organelles. We demonstrate that the two pathogen-containing vacuoles migrate independently to the host perinuclear region and rearrange the host microtubular network around each vacuole. However, Toxoplasma outcompetes Chlamydia to the host microtubule-organizing center to the detriment of the bacterium, which then shifts to a stress-induced persistent state. Solely in cells preinfected with Chlamydia, the centrosomes become associated with the chlamydial inclusion, while the Toxoplasma parasitophorous vacuole displays growth defects. Both pathogens fragment the host Golgi apparatus and recruit Golgi elements to retrieve sphingolipids. This study demonstrates that the productive infection by both Chlamydia and Toxoplasma depends on the capability of each pathogen to successfully adhere to a finely tuned developmental program that aims to remodel the host cell for the pathogen's benefit. In particular, this investigation emphasizes the essentiality of host organelle interception by intravacuolar pathogens to facilitate access to nutrients.

Chlamydia trachomatis ompA genotypes in male patients with urethritis in Greece: conservation of the serovar distribution and evidence for mixed infections with Chlamydophila abortus.

PCR amplification and nucleotide sequencing of the ompA gene of Chlamydia trachomatis were used to determine the prevalence and distribution of genotypes in 51 urine and urethral specimens from Greek male patients with urethritis, that were positive by the COBAS Amplicor test. A single C. trachomatis serovar was identified in 43 of the 51 amplified samples. Serovars F and E were the most prevalent (both 12, 28%), followed by D (9, 21%), G (4, 9%), B and K (both 2, 5%) and H and J (both 1, 2%). Over one third of the samples bared a variant ompA genotype that had been previously identified in other areas worldwide. Two results in this study, both observed for the first time, were of particular interest. First, the emergence of the unique variant genotype D/Ep6 (X77364.2) identified in 3 urethral samples. Second, the ompA genotype OCLH196 of the animal pathogen Chlamydophila abortus as well as a 23S rRNA gene fragment of this species detected by the assay ArrayTube™ was found in 7 urethral samples. The implications resulting from this observation for the health of the general population are discussed.

Drosophila melanogaster S2 cells: a model system to study Chlamydia interaction with host cells.

Chlamydia spp. are major causes of important human diseases, but dissecting the host-pathogen interactions has been hampered by the lack of bacterial genetics and the difficulty in carrying out forward genetic screens in mammalian hosts. RNA interference (RNAi)-based methodologies for gene inactivation can now be easily carried out in genetically tractable model hosts, such as Drosophila melanogaster, and offer a new approach to identifying host genes required for pathogenesis. We tested whether Chlamydia trachomatis infection of D. melanogaster S2 cells recapitulated critical aspects of mammalian cell infections. As in mammalian cells, C. trachomatis entry was greatly reduced by heparin and cytochalasin D. Inclusions were formed in S2 cells, acquired Golgi-derived sphingolipids, and avoided phagolysosomal fusion. Elementary body (EB) to reticulate body (RB) differentiation was observed, however, no RB to EB development or host cell killing was observed. RNAi-mediated inactivation of Rac, a Rho GTPase recently shown to be required for C. trachomatis entry in mammalian cells, inhibits C. trachomatis infection in S2 cells. We conclude that Drosophila S2 cells faithfully mimic early events in Chlamydia host cell interactions and provides a bona fide system to systematically dissect host functions important in the pathogenesis of obligate intracellular pathogens.

Chlamydia trachomatis uses host cell dynein to traffic to the microtubule-organizing center in a p50 dynamitin-independent process.

Chlamydiae are pathogenic obligate intracellular bacteria with a biphasic developmental cycle that involves cell types adapted for extracellular survival (elementary bodies, EBs) and intracellular multiplication (reticulate bodies, RBs). The intracellular development of chlamydiae occurs entirely within a membrane-bound vacuole termed an inclusion. Within 2 hours after entry into host cells, Chlamydia trachomatis EBs are trafficked to the perinuclear region of the host cell and remain in close proximity to the Golgi apparatus, where they begin to fuse with a subset of host vesicles containing sphingomyelin. Here, we provide evidence that chlamydial migration from the cell periphery to the peri-Golgi region resembles host cell vesicular trafficking. Chlamydiae move towards the minus end of microtubules and aggregate at the microtubule-organizing center (MTOC). In mammalian cells the most important minus-end-directed microtubule motor is cytoplasmic dynein. Microinjection of antibodies to a subunit of cytoplasmic dynein inhibited movement of chlamydiae to the MTOC, whereas microinjection of antibodies to the plus-directed microtubule motor, kinesin, had no effect. Surprisingly, overexpression of the protein p50 dynamitin, a subunit of the dynactin complex that links vesicular cargo to the dynein motor in minus directed vesicle trafficking, did not abrogate chlamydial migration even though host vesicle transport was inhibited. Nascent chlamydial inclusions did, however, colocalize with the p150(Glued) dynactin subunit, which suggests that p150(Glued) may be required for dynein activation or processivity but that the cargo-binding activity of dynactin, supplied by p50 dynamitin subunits and possibly other subunits, is not. Because chlamydial transcription and translation were required for this intracellular trafficking, chlamydial proteins modifying the cytoplasmic face of the inclusion membrane are probable candidates for proteins fulfilling this function.

Head-to-head multicenter comparison of DNA probe and nucleic acid amplification tests for Chlamydia trachomatis infection in women performed with an improved reference standard.

Few evaluations of tests for Chlamydia trachomatis have compared nucleic acid amplification tests (NAATs) with diagnostic tests other than those by culture. In a five-city study of 3,551 women, we compared the results of commercial ligase chain reaction (LCR) and PCR tests performed on cervical swabs and urine with the results of PACE 2 tests performed on cervical swabs, using independent reference standards that included both cervical swabs and urethral swab-urine specimens. Using cervical culture as a standard, the sensitivities of PACE 2, LCR, and PCR tests with cervical specimens were 78.1, 96.9, and 89.9%, respectively, and the specificities were 99.3, 97.5, and 98.2%, respectively. Using either cervical swab or urine LCR-positive tests as the standard decreased sensitivities to 60.8% for PACE 2 and to 75.8 and 74.9% for PCR with cervical swabs and urine, respectively. Specificities increased to 99.7% for PACE 2 and to 99.7 and 99.4% for PCR with cervical swabs and urine, respectively. Sensitivities with a cervical swab-urine PCR standard were 61.9% for PACE 2 and 85.5 and 80.8% for LCR with cervical swabs and urine, respectively. Specificities were 99.6% for PACE 2 and 99.0 and 98.9% for LCR with cervical swabs and urine, respectively. Cervical swab versus urine differences were significant only for PCR specificities (P = 0.034). Overall, LCR sensitivity exceeded that of PCR, and sensitivities obtained with cervical swabs exceeded those obtained with urine specimens by small amounts. These data have substantiated, using a large multicenter sample and a patient standard, that LCR and PCR tests performed on endocervical swabs and urine are superior to PACE 2 tests for screening C. trachomatis infections in women. In our study, NAATs improved the detection of infected women by 17 to 38% compared to PACE 2.

Phthirus pubis as a predictor for chlamydia infections in adolescents.

BACKGROUND: Providers not skilled in the evaluation of sexually transmitted infections (STIs) may treat pubic lice infestation without considering other organisms. This study compared the rate of chlamydia and gonorrhea infections in adolescents with and without pubic lice. GOAL: The goals of the study were to compare the rate of chlamydia and gonorrhea infections between adolescents infested or not infested with pubic lice and to evaluate lice infestation as a predictor for concurrent chlamydia or gonorrhea infection. STUDY DESIGN: A retrospective chart review of sexually active adolescents at a juvenile detention center in the Midwest between July 1998 and June 2000 was conducted. The index group was 62 adolescents with pubic lice screened for concurrent STIs. The control group included 201 randomly selected adolescents without pubic lice who underwent STI screening. RESULTS: In the entire study population (263 subjects), there were 60 cases of chlamydia (23% of all subjects) and 29 cases of gonorrhea (11%). Neisseria gonorrhoeae infection was present in 18% of index subjects and 9% of controls. Chlamydia trachomatis infection was noted in 39% of index subjects and 18% of controls. Pubic lice infestation predicted C trachomatis infection (odds ratio = 3.31). CONCLUSIONS: Pubic lice infestation is predictive of a concurrent C trachomatis infection in this population. Adolescents infested with pubic lice should be screened for other STIs, including chlamydia and gonorrhea.

Chlamydia trachomatis: probable cause of prostatitis.

Seventy-eight men with symptoms of chronic or subacute prostatitis were enrolled. Investigations for the presence of Chlamydia trachomatis in urethral swabs were carried out. The expressed prostatic secretions were additionally examined for Mycoplasma hominis, Ureaplasma urealyticum, Gardnerella vaginalis, other gram-negative and gram-positive bacteria, Trichomonas vaginalis, yeast-like fungi and leucocyte count. Furthermore, all patients were evaluated for the presence of serum anti-chlamydial IgG antibodies. Signs of inflammation on the basis of the count of leucocytes per hpf in the prostatic secretions were detected in 42 patients (group I). Prostatodynia was found in the remaining 36 men (group II). In group I, chlamydial antigen was detected in the urethra and expressed prostatic secretions (EPS) in 6 (14.3%) and 9 (21.4%) patients, respectively. No evidence of current chlamydial infection was found in group II. The presence of serum anti-chlamydial IgG antibodies was demonstrated in 13/42 (30.9%) patients with prostatitis and in 3/36 (8.3%) patients with prostatodynia (P < 0.01). The results suggest that chlamydia may be one of the causative agents of chronic prostatitis.

Complicaciones nasofaríngeas de la conjutivitis folicular por chalmydia trachomatis. Una causa de la falta de respuesta al tratamiento / Nasopharingeal complications secundary to Clamydia trachomatis follicular conjunctivitis. A cause of therapeutic failure

Se realizó un estudio retrospectivo en 6 409 casos consecutivos en un lapso de enero de 1992 a febrero de 1997, se seleccionaron 183 frotis nasales y 271 faríngeos provenientes de 396 pacientes. Chlamydia trachomatis fue el agente etiológico en el 67.80 por ciento de los frotis nasales y 8.11 por ciento de las muestras faríngeas. La infección nasal es casi siempre secundaria a la infección ocular y a su vez es un factor de diseminación y de falta de respuesta adecuada al tratamiento, por lo que es indispensable identificar el problema y dar tratamiento simultáneo

Seroprevalencia de infección por Chlamydia pneumoniae en población sana de Santiago de Chile / Seroprevalence of chlamydia pneumoniae infection in healthy population from Santiago, Chile

La evidencia de exposición previa a Chlamydia pneumoniae fue evaluada en 353 sujetos sanos de Santiago de Chile. Se efectuó determinación sérica de anticuerpos (IgG) específicos anti-Chlamydia pneumoniae con antígenos de la cepa AR-39 proporcionados por la Washington Research Foundation, con técnica de inmunofluorescencia indirecta. Títulos iguales o mayores de 1:16 fueron considerados positivos. 134 sueros fueron positivos para Chlamydia penumoniae, con título máximo de 1:256. Concluimos que en esta muestra la seroprevalencia fue de 38 por ciento y que en un título igual o mayor de 1:512 sugiere el diagnóstico de infección aguda por Chlamydia pneumoniae

The interaction of Chlamydia trachomatis with host cells: ultrastructural studies of the mechanism of release of a biovar II strain from HeLa 229 cells.

Serotypes of Chlamydia trachomatis classified as biovar II strains (immunotypes A, Ba, and B-K) are currently recognized as important human pathogens that produce disease characterized by a rather complex pathogenesis. We have studied some morphological phenomena in the interaction of C. trachomatis (strain UW3/Cx, serotype D) with HeLa 229 cells to define the mechanisms of release of these obligate intracellular parasites. Fluorescent-antibody staining of unfixed HeLa cells infected with chlamydiae suggested that this biotype of C. trachomatis can exit cells without concomitant death of the host cell. The mechanisms by which chlamydiae were released from cells were studied by scanning and transmission electron microscopy. Ultrastructural observations indicated that the chlamydial inclusion was segregated from host cytoplasm and transported to the host cell surface by a process similar to exocytosis. These observations of interactions between C. trachomatis and the host cell in vitro may be relevant for understanding the complex pathogenesis these organisms produce in vivo, specifically their ability to produce asymptomatic or latent infections.

Chlamydia trachomatis: an important sexually transmitted disease in adolescents and young adults.

Chlamydia trachomatis is being recognized as an important sexually transmitted disease in adolescents and young adults. This report reviews the recent literature regarding the many clinical entities encompassed by this organism; this includes urethritis and cervicitis as well as epididymitis, salpingitis, peritonitis, perihepatitis, urethral syndrome, Reiter syndrome, arthritis, endocarditis, and others. It is emphasized that many aspects of chlamydial infections parallel those of gonorrhea, including incidence, transmission, carrier state, reservoir, complications, (local and systemic), and others. A paragonococcal spectrum of sexual chlamydial disorders is discussed as well as effective antibiotic therapy. This microbiological agent must always be considered if venereal disease is suspected by the clinician in teenagers or adults. Mixed infections with Chlamydia trachomatis and Neisseria gonorrhoeae are common in both males and females. It may be preferable to treat gonorrhea with tetracycline to cover for this possibility.