The contribution of temperature, exposure intensity and visible light to the inhibitory effect of irradiation on acute chlamydial infection.

Water-filtered infrared A (wIRA) is radiation with a spectrum ranging from 780 to 1400 nm. Chlamydiaceae are obligate intracellular bacteria associated with various diseases in both animals and humans. A recent in vitro study demonstrated that wIRA combined with visible light (wIRA/VIS) has potential as a non-chemical method for the treatment of chlamydial infections without adversely affecting the cell viability. The aim of this study was to investigate the influence of various factors on the effect of wIRA/VIS on acute chlamydial infection, namely the impact of temperature, exposure intensity and infectious dose (multiplicity of infection) as well as the efficacy of the visible light component.We demonstrate that non-thermal effects contribute to the inhibition of acute chlamydial infection. Visible light enhances the inhibitory effect of wIRA on extracellular bacteria (elementary bodies or EBs).Moreover, the inhibitory effect of wIRA/VIS following treatment of EBs prior to infection correlated with increased irradiation intensity. The infectivity of mature chlamydial inclusions was significantly reduced upon wIRA/VIS exposure at all irradiation intensities investigated, suggesting the contribution of host cell factors to the anti-chlamydial effect of wIRA/VIS in the late stage of the developmental cycle. The effect of irradiation was not influenced by the infectious dose.

The chlamydial organism Simkania negevensis forms ER vacuole contact sites and inhibits ER-stress.

Most intracellular bacterial pathogens reside within membrane-surrounded host-derived vacuoles. Few of these bacteria exploit membranes from the host's endoplasmic reticulum (ER) to form a replicative vacuole. Here, we describe the formation of ER-vacuole contact sites as part of the replicative niche of the chlamydial organism Simkania negevensis. Formation of ER-vacuole contact sites is evolutionary conserved in the distantly related protozoan host Acanthamoeba castellanii. Simkania growth is accompanied by mitochondria associating with the Simkania-containing vacuole (SCV). Super-resolution microscopy as well as 3D reconstruction from electron micrographs of serial ultra-thin sections revealed a single vacuolar system forming extensive ER-SCV contact sites on the Simkania vacuolar surface. Simkania infection induced an ER-stress response, which was later downregulated. Induction of ER-stress with Thapsigargin or Tunicamycin was strongly inhibited in cells infected with Simkania. Inhibition of ER-stress was required for inclusion formation and efficient growth, demonstrating a role of ER-stress in the control of Simkania infection. Thus, Simkania forms extensive ER-SCV contact sites in host species evolutionary as diverse as human and amoeba. Moreover, Simkania is the first bacterial pathogen described to interfere with ER-stress induced signalling to promote infection.

Reactivation of chromosomally integrated human herpesvirus-6 by telomeric circle formation.

More than 95% of the human population is infected with human herpesvirus-6 (HHV-6) during early childhood and maintains latent HHV-6 genomes either in an extra-chromosomal form or as a chromosomally integrated HHV-6 (ciHHV-6). In addition, approximately 1% of humans are born with an inheritable form of ciHHV-6 integrated into the telomeres of chromosomes. Immunosuppression and stress conditions can reactivate latent HHV-6 replication, which is associated with clinical complications and even death. We have previously shown that Chlamydia trachomatis infection reactivates ciHHV-6 and induces the formation of extra-chromosomal viral DNA in ciHHV-6 cells. Here, we propose a model and provide experimental evidence for the mechanism of ciHHV-6 reactivation. Infection with Chlamydia induced a transient shortening of telomeric ends, which subsequently led to increased telomeric circle (t-circle) formation and incomplete reconstitution of circular viral genomes containing single viral direct repeat (DR). Correspondingly, short t-circles containing parts of the HHV-6 DR were detected in cells from individuals with genetically inherited ciHHV-6. Furthermore, telomere shortening induced in the absence of Chlamydia infection also caused circularization of ciHHV-6, supporting a t-circle based mechanism for ciHHV-6 reactivation.

Chlamydial infection in vitamin D receptor knockout mice is more intense and prolonged than in wild-type mice.

Vitamin D hormone (1,25-dihydroxyvitamin D) is involved in innate immunity and induces host defense peptides in epithelial cells, suggesting its involvement in mucosal defense against infections. Chlamydia trachomatis is a major cause of bacterial sexually transmitted disease worldwide. We tested the hypothesis that the vitamin D endocrine system would attenuate chlamydial infection. Vitamin D receptor knock-out mice (VDR(-/-)) and wild-type mice (VDR(+/+)) were infected with 10(3) inclusion forming units of Chlamydia muridarum and cervical epithelial cells (HeLa cells) were infected with C. muridarum at multiplicity of infection 5:1 in the presence and absence of 1,25-dihydroxyvitamin D3. VDR(-/-) mice exhibited significantly higher bacterial loading than wild-type VDR(+/+) mice (P<0.01) and cleared the chlamydial infection in 39 days, compared with 18 days for VDR(+/+) mice. Monocytes and neutrophils were more numerous in the uterus and oviduct of VDR(-/-) mice than in VDR(+/+) mice (P<0.05) at d 45 after infection. Pre-treatment of HeLa cells with 10nM or 100nM 1,25-dihydroxyvitamin D3 decreased the infectivity of C. muridarum (P<0.001). Several differentially expressed protein spots were detected by proteomic analysis of chlamydial-infected HeLa cells pre-treated with 1,25-dihydroxyvitamin D3. Leukocyte elastase inhibitor (LEI), an anti-inflammatory protein, was up-regulated. Expression of LEI in the ovary and oviduct of infected VDR(+/+) mice was greater than that of infected VDR(-/-) mice. We conclude that the vitamin D endocrine system reduces the risk for prolonged chlamydial infections through regulation of several proteins and that LEI is involved in its anti-inflammatory activity.

Epitheliocystis in Atlantic salmon, Salmo salar L., farmed in fresh water in Ireland is associated with 'Candidatus Clavochlamydia salmonicola' infection.

Intracellular inclusions containing chlamydia-like organisms are frequently observed in the gill epithelial cells of Atlantic salmon, Salmo salar L., cultured in fresh water in Ireland. In this study, the causative agent was identified in four separate freshwater sites, using 16s rRNA sequencing, as 'Candidatus Clavochlamydia salmonicola'. Histopathology and real-time (RT) PCR were used to further assess infections. The prevalence of infection ranged from 75-100% between sites and infection intensity was highly variable. No significant lesions were associated with these infections. As a diagnostic tool, RT-PCR proved marginally more sensitive than histopathology. The fate of 'Candidatus Clavochlamydia salmonicola' in Atlantic salmon post-seawater transfer was investigated in a 12-week marine longitudinal study. Both RT-PCR and histopathological examination indicate that the organism disappears from the gills 4-6 weeks post-transfer.

Microscopic diagnosis of dysbacteriosis in stained vaginal smears in clinical practice.

Dysbacteriosis is a microscopical diagnosis. In women with dysbacteriosis, an overgrowth of coccoid bacteria and almost a complete absence of lactobacilli are observed in the (stained) vaginal smear. The aim of this study was to determine the accuracy of this microscopic diagnosis in clinical practice. The analysis concerned 342 consecutive cases in which the microscopy of the stained smears was performed by general practitioners trained in diagnosing dysbacteriosis. These smears were sent to the pathologist for confirmation of the microscopical diagnosis of the clinician. The cytological diagnoses of the pathologist, sometimes performed on restained slides when the quality of the staining was substandard, were considered as the "gold standard." In 92 of the 342 cases, dysbacteriosis was unequivocally established by the pathologist. Sensitivity and specificity of the microscopical diagnoses of the clinicians were 40% and 85%, respectively. There were 37 false-positive and 54 false-negative diagnoses of dysbacteriosis rendered by the clinicians. The most frequent reason for a false-negative diagnosis was an excess of lactobacilli in the smear. This study shows that even in stained smears it is difficult for clinicians to render a correct evaluation of the status of the vaginal flora.

Chlamydiales in guinea-pigs and their zoonotic potential.

The aim was to detect and characterize chlamydial infections in guinea-pigs (GP) with ocular disease, study their pathogenicity and zoonotic potential and to test for the presence of Acanthamoebae spp. in GP eyes and to investigate whether they could act as vectors for Chlamydia-like organisms. Overall 126 GP, of which 77 were symptomatic, were screened by clinical examination, cytology, gross pathology, histology, immunohistochemistry, polymerase chain reaction (PCR) and bacteriology. A new Chlamydiaceae-specific intergenic spacer rRNA gene PCR, designed to amplify this segment linking the 16S and 23S regions, was performed. DNA samples were also received from one owner including samples of his cat and rabbit. Guinea-pigs: 48 of 75 symptomatic, but only 11 of 48 asymptomatic GP were positive by PCR for Chlamydophila caviae guinea-pig inclusion conjunctivitis (GPIC) (P < 0.0001). Eighteen of 75 or 15/48, respectively, were positive for DNA from Chlamydia-like organisms. Acanthamoebae-DNA could be found in two GP, of which one was symptomatic. Owner, cat and rabbit: Samples of all three species were positive by PCR for C. caviae GPIC and the owner's one-day disposable contact lenses showed a positive PCR result for the Chlamydia-like organism Parachlamydia acanthamoebae. No Acanthamoebae-DNA could be detected. This study is the first to describe Chlamydia-like organisms in GP and to detect C. caviae GPIC in human, cat and rabbit. Therefore, C. caviae GPIC could pose a zoonotic potential. We believe that the finding of C. caviae GPIC in species other than GP is probably not unique.

Immuno-histochemical demonstration of the role of chlamydiaceae in renal, uterine and Salpingeal disease of the koala, and demonstration of chlamydiaceae in novel sites.

Numerous bacteria, including Chlamydophila pecorum and Chlamydophila pneumoniae, are known to occur in diseased sites in koalas. In the present study the significance of such organisms was investigated by demonstrating their distribution in situ, in tissues collected opportunistically from wild koalas. Chlamydiaceae were demonstrated in epithelial cells and macrophages in association with pyogranulomatous pyelonephritis (8/11 kidneys), focal interstitial nephritis (3/21), and active inflammation and fibrosis of the entire upper female reproductive tract (10/10). In one case of pyelonephritis, Gram-positive cocci were also demonstrated in association with Chlamydiaceae and, in another, haematogenous filamentous bacteria appeared to be the sole aetiological agent. Three cases of chlamydial metritis were also superficially co-infected by a mixture of other bacteria. Chlamydiaceae were also demonstrated in pulmonary alveolar macrophages and epithelial cells in association with pneumonitis, and in hepatic and splenic macrophages in one koala. The study illustrated the prominent role of Chlamydiaceae in renal disease and disease of the uterus, uterine tube and ovarian bursa, with implications for pathogenesis and therapy. In addition, macrophages appeared to be a potential site of latent persistence from which systemic spread of infection might occur.

Endometrial histopathology in patients with culture-proved upper genital tract infection and laparoscopically diagnosed acute salpingitis.

To define and quantitate histologic changes in the endometrium that best correlate with documented upper genital tract infection (UGTI) and laparoscopically diagnosed acute salpingitis, we studied endometrial biopsy specimens from 69 consecutive patients with clinically suspected acute pelvic inflammatory disease (PID) who underwent microbiological evaluation for UGTI and laparoscopic examination for acute salpingitis. Both UGTI and acute laparoscopically confirmed salpingitis were present in 37 patients (54%), UGTI without salpingitis in 1 (1%), salpingitis without UGTI in 11 (16%), and neither UGTI nor salpingitis in 20 (29%). Chlamydia trachomatis or Neisseria gonorrhoeae UGTI was found in 34 women, Escherichia coli in two patients, Peptococcus magnus in one woman, and with Streptococcus agalactiae in one woman. The following features were correlated both with UGTI and with salpingitis: presence of any neutrophils in the endometrial surface epithelium; neutrophils within gland lumens; dense subepithelial stromal lymphocytic infiltration; any stromal plasma cells; and germinal centers containing transformed lymphocytes. The simultaneous presence of five or more neutrophils per X 400 field in endometrial surface epithelium, together with one or more plasma cell per X 120 field in endometrial stroma, was the best predictor of UGTI plus salpingitis. This combination had a sensitivity of 92% and a specificity of 87% for predicting the diagnosis of both UGTI and laparoscopically confirmable acute salpingitis. Prospective studies are needed to assess the usefulness of these criteria.