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Chlamydia trachomatis ( CT ) genital tract infection is insidious, and patients often have no conscious symptoms.Delayed treatment after infection can lead to serious complications. Chlamydia muridarum ( CM ) genital tract infection in female mice can simulate CT genital tract infection in women, which is an ideal model to investigate the pathogenesis of CT . CM plasmid protein pGP3, chromosomal protein TC0237/TC0668, CM -specific CD8 + T cells, TNF-α, and IL-13 can induce genital tract inflammation, CD4 + T cells are responsible for CM clearance. However, tubal inflammation persists after genital tract CM is removed. Genital tract CM can spread spontaneously in vivo and colonize the gastrointestinal (GI) tract, but the GI tract CM cannot reverse spread to the genital tract. The survival time and number of CM transmitted from genital tract to GI tract are positively correlated with the long-term lesion of oviduct, while the CM inoculated directly into the GI tract has no pathogenicity in both the genital and GI tract. The double attack pattern of Chlamydia -induced genital tract inflammatory lesions is as follows: CM infection of oviduct epithelial cells initiates the process of oviduct repair as the first attack. After genital CM spreads to the GI tract, activated chlamydia-specific CD8 + T cells are recruited to the genital tract and secreted pro-fibrotic cytokines such as TNF-α and IL-13. This process is called the second attack which transform tubal repair initiated by the first attack into long-term tubal fibrosis/hydrosalpinx. Elucidating the pathogenic mechanism of Chlamydia infection can provide new ideas for the development of Chlamydia vaccine, which is expected to solve the problems of infertility caused by repeated CT infection in women.

While glycogen synthase A deficiency can reduce the growth and proliferation of Chlamydia muridarum, the effect of glycogen synthase A on the pathogenic process of C. muridarum remains unclear. To characterize the effect of glycogen synthase A deficiency on the pathogenicity of C. muridarum in the genital tract, BALB/c mice were intravaginally inoculated with wild-type, plasmid-free and glycogen synthase A-deficient C. muridarum, and the genital tract tissue was isolated to assess the severity of hydrosalpinx and the levels of oviduct dilatation at day 60 after infection. The glycogen storage capacity and in vitro infection ability of different C. muridarum strains were analyzed by periodic acid-Schiff staining and quantification of progeny elementary body(EB) formation. The tissue homogenate was used to determine the recovery of different C. muridarum strains. The results show that glycogen synthase A-deficient C. muridarum induced reduction of hydrosalpinx and attenuated the extent of oviduct dilatation in mice, and exhibited reduced growth and proliferation in the mouse lower genital tract. In addition, glycogen synthase A point mutations at different sites reduced the glycogen storage capacity and in vitro infectivity of C. muridarum to different degrees. Glycogen synthase A deficiency also reduced the host inflammatory reaction and ascending infection of C. muridarum., (© FEMS 2020.)

Sexually transmitted Chlamydia , which can cause fibrotic pathology in women's genital tracts, is also frequently detected in the gastrointestinal tract. However, the medical significance of the gastrointestinal Chlamydia remains unclear. A murine Chlamydia readily spreads from the mouse genital tract to the gastrointestinal tract while inducing oviduct fibrotic blockage or hydrosalpinx. We previously proposed a two-hit model in which the mouse gastrointestinal Chlamydia might induce the second hit to promote genital tract pathology, and we are now providing experimental evidence for testing the hypothesis. First, chlamydial mutants that are attenuated in inducing hydrosalpinx in the genital tract also reduce their colonization in the gastrointestinal tract, leading to a better correlation of chlamydial induction of hydrosalpinx with chlamydial colonization in the gastrointestinal tract than in the genital tract. Second, intragastric coinoculation with a wild-type Chlamydia rescued an attenuated Chlamydia mutant to induce hydrosalpinx, while the chlamydial mutant infection in the genital tract alone was unable to induce any significant hydrosalpinx. Finally, the coinoculated gastrointestinal Chlamydia failed to directly spread to the genital tract lumen, suggesting that gastrointestinal Chlamydia may promote genital pathology via an indirect mechanism. Thus, we have demonstrated a significant role of gastrointestinal Chlamydia in promoting pathology in the genital tract possibly via an indirect mechanism. This study provides a novel direction/dimension for further investigating chlamydial pathogenic mechanisms., (Copyright © 2020 American Society for Microbiology.)

Background and Aims: Atherosclerosis is a chronic inflammatory disease, and recent studies have shown that infection at remote sites can contribute to the progression of atherosclerosis in hyperlipidemic mouse models. In this report, we tested the hypothesis that genital Chlamydia infection could accelerate the onset and progression of atherosclerosis., Methods: Apolipoprotein E (Apoe -/- ) and LDL receptor knockout (Ldlr -/- ) mice on a high-fat diet were infected intra-vaginally with Chlamydia muridarum. Atherosclerotic lesions on the aortic sinuses and in the descending aorta were assessed at 8-weeks post-infection. Systemic, macrophage, and vascular site inflammatory responses were assessed and quantified., Results: Compared to the uninfected groups, infected Apoe -/- and Ldlr -/- mice developed significantly more atherosclerotic lesions in the aortic sinus and in the descending aorta. Increased lesions were associated with higher circulating levels of serum amyloid A-1, IL-1β, TNF-α, and increased VCAM-1 expression in the aortic sinus, suggesting an association with inflammatory responses observed during C. muridarum infection. Genital infection courses were similar in Apoe -/- , Ldlr -/- , and wild type mice. Further, Apoe -/- mice developed severe uterine pathology with increased dilatations. Apoe-deficiency also augmented cytokine/chemokine response in C. muridarum infected macrophages, suggesting that the difference in macrophage response could have contributed to the genital pathology in Apoe -/- mice., Conclusions: Overall, these studies demonstrate that genital Chlamydia infection exacerbates atherosclerotic lesions in hyperlipidemic mouse and suggest a novel role for Apoe in full recovery of uterine anatomy after chlamydial infection., (Copyright © 2019 Elsevier B.V. All rights reserved.)

It has been shown that caspase-1, but not its upstream activator, ASC, contributes to oviduct pathology during mouse genital Chlamydia muridarum infection. We hypothesized that this dichotomy is due to the inadvertent absence of caspase-11 in previously used caspase-1-deficient mice. To address this, we studied the independent contributions of caspase-1 and -11 during genital Chlamydia infection. Our results show that caspase-11 deficiency was sufficient to recapitulate the effect of the combined absence of both caspase-1 and caspase-11 on oviduct pathology. Further, mice that were deficient for both caspase-1 and -11 but that expressed caspase-11 as a transgene (essentially, caspase-1-deficient mice) had no significant difference in oviduct pathology from control mice. Caspase-11-deficient mice showed reduced dilation in both the oviducts and uterus. To determine the mechanism by which caspase-11-deficient mice developed reduced pathology, the chlamydial burden and immune cell infiltration were determined in the oviducts. In the caspase-11-deficient mice, we observed increased chlamydial burdens in the upper genital tract, which correlated with increased CD4 T cell recruitment, suggesting a contribution of caspase-11 in infection control. Additionally, there were significantly fewer neutrophils in the oviducts of caspase-11-deficient mice, supporting the observed decrease in the incidence of oviduct pathology. Therefore, caspase-11 activation contributes to pathogen control and oviduct disease independently of caspase-1 activation., (Copyright © 2019 American Society for Microbiology.)

Chlamydiosis is the most documented and serious disease of koalas, characterized by ocular, urinary, and reproductive lesions. Since little attention has been paid to the pathological effects of this infection in the male reproductive system, we aimed to determine the incidence and severity of reproductive pathology associated with chlamydial infection in male koalas submitted to koala hospitals in southeast Queensland. The entire reproductive tract from 62 sexually mature male koalas not suitable for rehabilitation was evaluated and 677 tissue samples were collected for histology, immunohistochemistry (IHC), and real-time polymerase chain reaction (qPCR). Lymphoplasmacytic inflammation was observed in 178 of 677 (26.3%) tissue samples from the upper and lower reproductive tract, mainly in the prostatic, penile, and membranous urethra. IHC was positive for the chlamydial antigen in 19 of 451 normal samples (4.2%) and 46 of 178 samples with inflammation (25.8%), located within the cytoplasm of epithelial cells of the epididymis, vas deferens, prostate, bulbourethral glands, and the prostatic membranous and penile urethra. Chlamydia pecorum was detected via qPCR in 319 of 451 normal samples (70.7%) and 159 of 178 samples with inflammation (89.3%), with the highest incidence in the penile urethra, prostate, membranous urethra, and bulbourethral glands. This study suggests that Chlamydia infection in the male reproductive tract is more widespread than originally thought. Furthermore, the male reproductive tract might be a reservoir for persistent chlamydial infections in koalas, with important implications for prophylactic strategies and epidemiology.

Problem: Chlamydia trachomatis infections are often associated with acute syndromes including cervicitis, urethritis, and endometritis, which can lead to chronic sequelae such as pelvic inflammatory disease (PID), chronic pelvic pain, ectopic pregnancy, and tubal infertility. As epithelial cells are the primary cell type productively infected during genital tract Chlamydia infections, we investigated whether Chlamydia has any impact on the integrity of the host epithelial barrier as a possible mechanism to facilitate the dissemination of infection, and examined whether TLR3 function modulates its impact., Method of Study: We used wild-type and TLR3-deficient murine oviduct epithelial (OE) cells to ascertain whether C. muridarum infection had any effect on the epithelial barrier integrity of these cells as measured by transepithelial resistance (TER) and cell permeability assays. We next assessed whether infection impacted the transcription and protein function of the cellular tight-junction (TJ) genes for claudins1-4, ZO-1, JAM1 and occludin via quantitative real-time PCR (qPCR) and western blot., Results: qPCR, immunoblotting, transwell permeability assays, and TER studies show that Chlamydia compromises cellular TJ function throughout infection in murine OE cells and that TLR3 deficiency significantly exacerbates this effect., Conclusion: Our data show that TLR3 plays a role in modulating epithelial barrier function during Chlamydia infection of epithelial cells lining the genital tract. These findings propose a role for TLR3 signaling in maintaining the integrity of epithelial barrier function during genital tract Chlamydia infection, a function that we hypothesize is important in helping limit the chlamydial spread and subsequent genital tract pathology., Competing Interests: The authors have declared that no competing interests exist.

Background: Infection with plasmid-free Chlamydia trachomatis and Chlamydia muridarum fails to induce severe pathology, however, the mechanisms underlying this phenotype are unclear., Objectives: To elucidate the mechanisms of chlamydial plasmid-mediated pathology in mouse oviducts., Materials & Methods: BALB/c mice were intravaginally infected with either plasmid-competent or plasmid-free C. muridarum strains. To compare the survival and ascending infection of these strains, vaginal swabs and genital tract tissues were collected and cultured with HeLa cells to monitor the recovery of live organisms. In addition, Chlamydia strains were intrabursally inoculated into the oviducts of mice to assess pathogenicity. Cytokine levels in the vaginal swabs collected from both the plasmid-competent and plasmid-free C. muridarum-infected mice were detected using Bio-Plex Pro Mouse Cytokine, Chemokine, and Growth Factor Assays., Results: The plasmid-competent C. muridarum strain induced hydrosalpinx formation in mouse oviducts following intravaginal inoculation, however, this was not the case for the plasmid-free C. muridarum strain. The lack of hydrosalpinges in response to the plasmid-free C. muridarum strain correlated with its significantly reduced ability to survive and disseminate to the upper genital tract. Furthermore, the plasmid-free C. muridarum failed to induce hydrosalpinx formation in mice, even when the strain was intrabursally injected into oviducts. A comparison of the cytokine levels in mouse vaginal secretions showed that the plasmid-free C. muridarum strain induced less IL-15, LIF, MIP-2, IL-1α, IL-1β, TNF-α, and RANTES., Conclusion: C. muridarum plasmid contributes to oviduct pathology by promoting bacterial survival and ascending infection, and triggering host inflammatory responses.

Chlamydia is a major bacterial pathogen in humans and animals globally. Yet 80% of infections never progress to clinical disease. Decades of research have generated an interconnected network linking pathogen, host, and environmental factors to disease expression, but the relative importance of these and whether they account for disease progression remains unknown. To address this, we used structural equation modeling to evaluate putative factors likely to contribute to urogenital and ocular chlamydial disease in the koala (Phascolarctos cinereus). These factors include Chlamydia detection, load, and ompA genotype; urogenital and ocular microbiomes; host sex, age, weight, body condition; breading season, time of year; location; retrovirus co-infection; and major histocompatibility complex class II (MHCII) alleles. We show different microbiological processes underpin disease progression at urogenital and ocular sites. From each category of factors, urogenital disease was most strongly predicted by chlamydial PCR detection and load, koala body condition and environmental location. In contrast, ocular disease was most strongly predicted by phylum-level Chlamydiae microbiome proportions, sampling during breeding season and co-infection with koala retrovirus subtype B. Host MHCII alleles also contributed predictive power to both disease models. Our results also show considerable uncertainty remains, suggesting major causal mechanisms are yet to be discovered.

Chlamydia trachomatis urogenital serovars primarily replicate in epithelial cells lining the reproductive tract. Epithelial cells recognize Chlamydia through cell surface and cytosolic receptors, and/or endosomal innate receptors such as Toll-like receptors (TLRs). Activation of these receptors triggers both innate and adaptive immune mechanisms that are required for chlamydial clearance, but are also responsible for the immunopathology in the reproductive tract. We previously demonstrated that Chlamydia muridarum (Cm) induces IFN-β in oviduct epithelial cells (OE) in a TLR3-dependent manner, and that the synthesis of several cytokines and chemokines are diminished in Cm-challenged OE derived from TLR3-/- 129S1 mice. Furthermore, our in vitro studies showed that Cm replication in TLR3-/- OE is more efficient than in wild-type OE. Because TLR3 modulates the release inflammatory mediators involved in host defense during Cm infection, we hypothesized that TLR3 plays a protective role against Cm-induced genital tract pathology in congenic C57BL/6N mice. Using the Cm mouse model for human Chlamydia genital tract infections, we demonstrated that TLR3-/- mice had increased Cm shedding during early and mid-stage genital infection. In early stage infection, TLR3-/- mice showed a diminished synthesis of IFN-β, IL-1β, and IL-6, but enhanced production of IL-10, TNF-α, and IFN-γ. In mid-stage infection, TLR3-/- mice exhibited significantly enhanced lymphocytic endometritis and salpingitis than wild-type mice. These lymphocytes were predominantly scattered along the endometrial stroma and the associated smooth muscle, and the lamina propria supporting the oviducts. Surprisingly, our data show that CD4+ T-cells are significantly enhanced in the genital tract TLR3-/- mice during mid-stage Chlamydial infection. In late-stage infections, both mouse strains developed hydrosalpinx; however, the extent of hydrosalpinx was more severe in TLR3-/- mice. Together, these data suggest that TLR3 promotes the clearance of Cm during early and mid-stages of genital tract infection, and that loss of TLR3 is detrimental in the development hydrosalpinx.

Ascension to the oviduct is necessary for Chlamydia to induce tubal infertility. Using the Chlamydia muridarum induction of hydrosalpinx mouse model, we have demonstrated a significant role of the uterotubal junction in preventing chlamydial ascending infection. First, delivery of C. muridarum to either side of the uterotubal junction resulted in significant reduction in live organisms from the tissues on the opposite sides. However, the recovery yields remained similar among different sections of the uterine horn. These observations suggest that the uterotubal junction may function as a barrier between the uterine horn and oviduct. Second, deficiency in innate immunity signaling pathways mediated by either MyD88 or STING significantly compromised the uterotubal junction barrier function, permitting C. muridarum to spread freely between uterine horn and oviduct. Finally, transcervical inoculation of C. muridarum led to significantly higher incidence of bilateral hydrosalpinges in the STING-deficient mice while the same inoculation mainly induced unilateral hydrosalpinx in the wild type mice, suggesting that the STING pathway-dependent uterotubal junction plays a significant role in preventing tubal pathology. Thus, we have demonstrated for the first time that the uterotubal junction is a functional barrier for preventing tubal infection by a sexually transmitted agent, providing the first in vivo evidence for detecting chlamydial infection by the STING pathway.

Genital infection by Chlamydia trachomatis is the most common bacterial sexually transmitted disease worldwide. It causes serious reproductive health complications, including pelvic inflammatory disease and infertility. Stress is implicated as a risk factor for various infections; however, its effect on chlamydia genital infection is unknown. We previously showed that repeated exposure of mice to cold water results in increased severity of chlamydia genital infection. In this study, cold water-induced stress resulted in (i) elevated levels of norepinephrine (NE) and epinephrine in the spleen and genital tract of stressed mice; (ii) elevated IL-1β, TNF-α, IL-6 and nitric oxide production in macrophage-rich peritoneal cells of mice; (iii) supplement of NE in vitro exerts an immunosuppressive effect on splenic T-cell production of cytokines; (iv) decreased C. muridarum shedding in the genital tract of β1Adr/β2Adr receptor KO mice; and (v) a higher rate of infertility in infected mice. These results suggest that cold water stress induces the production of catecholamines, which may play a critical role in the modulation of the immune system leading to increased intensity of C. muridarum genital infection., (© FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Chlamydia trachomatis is the most prevalent sexually transmitted bacterial infection. However, whether Chlamydia trachomatis has a negative impact on sperm quality and male fertility is still controversial. Herein, we report the effects on sperm quality of the in vitro exposure of spermatozoa to Chlamydia trachomatis, and also the effects of male genital infection on male fertility using an animal model. Human and mouse sperm were obtained from healthy donors and cauda epididimys from C57BL/6 mice, respectively. Highly motile human or mouse spermatozoa were in vitro exposed to C. trachomatis (serovar E or LGV) or C. muridarum, respectively. Then, sperm quality parameters were analyzed. Moreover, male fertility of Chlamydia muridarum infected male C57BL/6 mice was assessed. Human or murine sperm in vitro exposed to increasing bacterial concentrations or soluble factors from C. trachomatis or C. muridarum, respectively, did not show differences in sperm motility and viability, apoptosis, mitochondrial membrane potential, DNA fragmentation, ROS production and lipid peroxidation levels, when compared with control sperm (p > 0.05). Moreover, no differences in fertility parameters (potency, fecundity, fertility index, pre- and post-implantation loss) were observed between control and infected males. In conclusion, our results indicate that Chlamydia spp. neither directly exerts deleterious effects on spermatozoa nor impairs male fertility.

To respond to the situation of high prevalence and need for effective treatment for sexually transmitted infections (STIs) in low-resource settings, syndromic diagnostic approach was recommended by the World Health Organization and was adopted by National AIDS Control Organization at the primary health centre level. A retrospective study was undertaken in symptomatic patients attending an STI clinic to validate the syndromic approach for genitourinary discharge syndrome. For aetiological diagnosis, culture and/or polymerase chain reaction was used. An infective aetiology could be established in only 20% (106 of 530) patients. The present data call for an early appraisal and review of the diagnostic policy by national authorities on syndromic case management.

Chlamydia trachomatis is the leading cause of bacterial sexually transmitted infections (STIs) and preventable blindness. Untreated, asymptomatic infection as well as frequent re-infection are common and may drive pelvic inflammatory disease, ectopic pregnancy, and infertility. In vivo models of chlamydial infection continue to be instrumental in progress toward a vaccine and further elucidating the pathogenesis of this intracellular bacterium, however significant gaps in our understanding remain. Chlamydial host cell exit occurs via two mechanisms, lysis and extrusion, although the latter has yet to be reported in vivo and its biological role is unclear. The objective of this study was to investigate whether chlamydial extrusions are shed in vivo following infection with multiple strains of Chlamydia . We utilized an established C3H/HeJ murine cervicovaginal infection model with C. trachomatis serovars D and L2 and the Chlamydia muridarum strain MoPn to monitor the (i) time course of infection and mode of host cell exit, (ii) mucosal and systemic immune response to infection, and (iii) gross and histopathology following clearance of active infection. The key finding herein is the first identification of chlamydial extrusions shed from host cells in an in vivo model. Extrusions, a recently appreciated mode of host cell exit and potential means of dissemination, had been previously observed solely in vitro . The results of this study demonstrate that chlamydial extrusions exist in vivo and thus warrant further investigation to determine their role in chlamydial pathogenesis.

Genital tract infections with ureaplasma urealyticum (UU) and chlamydia trachomatis (CT) are the most frequent sexually-transmitted disease worldwide. UU and CT infections are considered to be the leading cause for infertility and adverse pregnancy outcomes. However, little is known about the specific effect of cervical UU and CT infections on the etiology of female infertility, as well as the pregnancy outcomes of the patients undergoing in vitro fertilization/intracytoplasmic sperm injection-embryo transfer (IVF/ICSI-ET). In order to find the association between cervical UU and/or CT infection and pregnancy outcomes, we conducted a retrospective case-control study on the patients undergoing IVF/ICSI-ET with cervical UU and/or CT infection. A total of 2208 patients who received IVF/ICSI-ET were enrolled in this study. Data on the general conditions, pregnancy history and clinical pregnant outcomes were analyzed in terms of the cervical UU and CT detection. Our results revealed that cervical UU and CT infections were the risk factors for ectopic pregnancy and tubal factor-induced infertility. Moreover, the pregnancy rate, abortion rate, ectopic pregnancy rate and premature birth rate in patients with UU and/or CT infections showed no significant difference when compared with the control group. We recommend that cervical UU and CT detection should be an optional item for infertility patients and clinical UU detection should differentiate the subtypes of cervical UU. Positive cervical UU and CT infections should not be taken as strict contraindications for IVF/ICSI-ET.

The Chlamydia muridarum induction of the upper genital tract pathology in mice has been used to investigate the mechanisms of chlamydial pathogenesis. We report that the NCF1 (neutrophil cytosolic factor1)-encoded p47phox (phagocyte oxidase), an essential subunit of nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase, contributes significantly to C. muridarum induction of hydrosalpinx. Mice lacking p47phox (p47phox-deficient) were no longer able to develop significant hydrosalpinx following an intravaginal infection with C. muridarum. However, there was no significant difference in uterine horn dilation (as a result of the endometrial glandular duct dilation) between the p47phox-deficient and -sufficient mice. Thus, the role of NADPH oxidase in chlamydial pathogenesis is restricted to the oviduct tissue rather than the entire upper genital tract. Interestingly, both the p47phox-deficient and -sufficient mice displayed similar levels of chlamydial live organism shedding from the lower genital tract, suggesting that the NADPH oxidase is not required for the mouse control of chlamydial infection in the lower genital tract. Furthermore, the p47phox deficiency did not affect the infectious organism burden in the upper genital tract tissues, indicating that the NADPH-oxidase activity is not necessary for the mouse prevention of chlamydial ascension from the lower to upper genital tracts. However, the p47phox-defieicnt mice displayed a significantly reduced chronic inflammatory infiltration in the oviduct but not uterine tissues, supporting the finding that the NADPH oxidase activity is required for chlamydial induction of dilation in the oviduct but not the endometrial glandular duct. Thus, we have demonstrated a significant role of the host NADPH oxidase in promoting chronic inflammatory pathology in the oviduct following chlamydial infection., (Copyright © 2015 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.)

Chlamydia trachomatis urogenital serovars are intracellular bacteria that parasitize human reproductive tract epithelium. As the principal cell type supporting bacterial replication, epithelial cells are central to Chlamydia immunobiology initially as sentries and innate defenders, and subsequently as collaborators in adaptive immunity-mediated bacterial clearance. In asymptomatic individuals who do not seek medical care a decisive struggle between C. trachomatis and host defenses occurs at the epithelial interface. For this study, we modeled the immunobiology of epithelial cells and macrophages lining healthy genital mucosa and inflamed/infected mucosa during the transition from innate to adaptive immunity. Upper reproductive tract epithelial cell line responses were compared to bone marrow-derived macrophages utilizing gene expression microarray technology. Those comparisons showed minor differences in the intrinsic innate defenses of macrophages and epithelial cells. Major lineage-specific differences in immunobiology relate to epithelial collaboration with adaptive immunity including an epithelial requirement for inflammatory cytokines to express MHC class II molecules, and a paucity and imbalance between costimulatory and coinhibitory ligands on epithelial cells that potentially limits sterilizing immunity (replication termination) to Chlamydia-specific T cells activated with limited or unconventional second signals., (© FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Rhesus macaques were studied to directly address the potential for plasmid-deficient Chlamydia trachomatis to serve as a live attenuated vaccine in the genital tract. Five repeated cervical inoculations of rhesus macaques with wild-type serovar D strain D/UW-3/Cx or a plasmid-deficient derivative of this strain, CTD153, resulted in infections with similar kinetics and induced comparable levels of protective immunity. After all animals received five challenges with D/UW-3/Cx, levels of inflammation observed grossly and histologically were similar between the groups. Animals in both groups developed evidence of oviduct dilatation; however, reduced oviduct dilatation was observed for "controllers," i.e., animals without detectable chlamydial DNA in the fimbriae at weeks 5 and 12. Grouping animals into "ascenders" and "controllers" revealed that elevated early T cell responses were associated with protection, whereas higher antibody responses were associated with ascension. Protected animals shared common major histocompatibility complex (MHC) alleles. Overall, genetic differences of individual animals, rather than the presence or absence of the chlamydial plasmid in the primary infecting strain, appeared to play a role in determining the outcome of infection., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

The hallmark of chlamydial infection is the development of upper genital pathology in the form of hydrosalpinx and oviduct and/or tubal dilatation. Although molecular events leading to genital tissue presentation and cellular architectural remodelling are unclear, early-stage host immune responses are believed to contribute to these long-term sequelae. Recently, we reported the contribution of selected infection-associated microRNAs (miRs) in the generation of host immunity at early-stage infection (day 6 after intravaginal Chlamydia muridarum challenge in C57BL/6 mice). In this report, we describe the contribution of an infection-associated microRNA, i.e. miR-214, to host immunity. Chlamydia muridarum infection in the C57BL/6 mouse genital tract significantly down-regulated miR-214 while up-regulating intracellular adhesion molecule 1 (ICAM1) gene expression. These in vivo observations were confirmed by establishing direct regulation of ICAM-1 by miR-214 in ex vivo genital cell cultures in the presence of miR-214 mimic and inhibitor. Because, ICAM-1 contributes to recruitment of neutrophils following infection, we also demonstrated that alteration of ICAM1 by miR-214 in interleukin-17A-deficient (IL-17A(-/-) ) mice correlated with reduction of neutrophils infiltrating genital tissue at day 6 after challenge. Additionally, these early-stage events resulted in significantly decreased genital pathology in IL-17A(-/-) mice compared with C57BL/6 mice. This report provides evidence for early-stage regulation of ICAM1 by microRNAs, resulting in reduction of genital pathology associated with chlamydial infection., (© 2015 John Wiley & Sons Ltd.)

Background: We demonstrated previously that tumor necrosis factor α (TNF-α)-producing Chlamydia-specific CD8(+) T cells cause oviduct pathological sequelae., Methods: In the current study, we used wild-type C57BL/6J (WT) mice with a deficiency in genes encoding TNF receptor superfamily member 1a (TNFR1; TNFR1 knockout [KO] mice), TNF receptor superfamily member 1b (TNFR2; TNFR2 KO mice), and both TNFR1 and TNFR2 (TNFR1/2 double KO [DKO] mice) and mix-match adoptive transfers of CD8(+) T cells to study chlamydial pathogenesis., Results: TNFR1 KO, TNFR2 KO, and TNFR1/2 DKO mice displayed comparable clearance of primary or secondary genital Chlamydia muridarum infection but significantly reduced oviduct pathology, compared with WT animals. The Chlamydia-specific total cellular cytokine response in splenic and draining lymph nodes and the antibody response in serum were comparable between the WT and KO animals. However, CD8(+) T cells from TNFR2 KO mice displayed significantly reduced activation (CD11a expression and cytokine production), compared with TNFR1 KO or WT animals. Repletion of TNFR2 KO mice with WT CD8(+) T cells but not with TNFR2 KO CD8(+) T cells and repletion of TNFR1 KO mice with either WT or TNFR1 KO CD8(+) T cells restored oviduct pathology to WT levels in both KO groups., Conclusions: Collectively, these results demonstrate that TNFR2-bearing CD8(+) T cells and TNFR1-bearing non-CD8(+) T cells contribute significantly to oviduct pathology following genital chlamydial infection., (© The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Background: The role of organism load in Chlamydia trachomatis infection is not well understood. We conducted a systematic review to investigate the epidemiology of C. trachomatis organism load in human genital chlamydia infection., Methods: Embase, PubMed, and Medline databases were searched for literature published through August 2014. English-language publications that quantified load in humans were eligible. Participant characteristics and laboratory data were extracted., Results: A total of 737 records were identified, and 29 publications involving 40 883 participants were included. In women, load was highest for cervical swabs and lowest for urine specimens. In men, load was highest for rectal swabs and similar for urethral swabs and urine specimens. Evidence of any association between load and age, serovar, risk of transmission, hormone levels, and concurrent sexually transmitted infections was inconsistent. Eight of 9 culture-based studies found an association between load and signs and symptoms, in contrast with only 3 of 8 nucleic acid amplification test (NAAT)-based studies (P = .03)., Conclusion: Chlamydia organism load varies by specimen type and site of sampling, and viable chlamydia organism load may be a more important indicator of severity of infection than total load measured by NAAT., (© The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Although modern Chlamydia muridarum has been passaged for decades, there are no reports on the consequences of serial passage with strong selection pressure on its fitness. In order to explore the potential for Pasteurian selection to induce genomic and phenotypic perturbations to C. muridarum, a starter population was passaged in cultured cells for 28 generations without standard infection assistance. The resultant population, designated CMG28, displays markedly reduced in vitro dependence on centrifugation for infection and low incidence and severity of upper genital tract pathology following intravaginal inoculation into mice compared to the parental C. muridarum population, CMG0. Deep sequencing of CMG0 and CMG28 revealed novel protein variants in the hypothetical genes TC0237 (Q117E) and TC0668 (G322R). In vitro attachment assays of isogenic plaque clone pairs with mutations in either TC0237 and TC0668 or only TC0237 reveal that TC0237(Q117E) is solely responsible for enhanced adherence to host cells. Paradoxically, double mutants, but not TC0237(Q117E) single mutants, display severely attenuated in vivo pathogenicity. These findings implicate TC0237 and TC0668 as novel genetic factors involved in chlamydial attachment and pathogenicity, respectively, and show that serial passage under selection pressure remains an effective tool for studying Chlamydia pathogenicity., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

A mouse model for Chlamydia suis genital infection was developed. Ninety-nine mice were randomly divided into three groups and intravaginally inoculated with chlamydia: 45 mice (group 1) received C. suis purified elementary bodies (EBs), 27 (group 2) were inoculated with C. trachomatis genotype E EBs and 27 mice (group 3) with C. trachomatis genotype F EBs. Additionally, 10 mice were used as a negative control. At seven days post-infection (dpi) secretory anti-C. suis IgA were recovered from vaginal swabs of all C. suis inoculated mice. Chlamydia suis was isolated from 93, 84, 71 and 33% vaginal swabs at 3, 5, 7 and 12 dpi. Chlamydia trachomatis genotype E and F were isolated from 100% vaginal swabs up to 7 dpi and from 61 and 72%, respectively, at 12 dpi. Viable C. suis and C. trachomatis organisms were isolated from uterus and tubes up to 16 and 28 dpi, respectively. The results of the present study show the susceptibility of mice to intravaginal inoculation with C. suis. A more rapid course and resolution of C. suis infection, in comparison to C. trachomatis, was highlighted. The mouse model could be useful for comparative investigations involving C. suis and C. trachomatis species., (© FEMS 2014. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The aim of the present study was to reveal the characteristic features of genital Chlamydia suis infection and re-infection in female pigs by studying the immune response, pathological changes, replication of chlamydial bacteria in the genital tract and excretion of viable bacteria. Pigs were intravaginally infected and re-infected with C. suis strain S45, the type strain of this species. We demonstrated that S45 is pathogenic for the female urogenital tract. Chlamydia replication occurred throughout the urogenital tract, causing inflammation and pathology. Furthermore, genital infection elicited both cellular and humoral immune responses. Compared to the primo-infection of pigs with C. suis, re-infection was characterized by less severe macroscopic lesions and less chlamydial elementary bodies and inclusions in the urogenital tract. This indicates the development of a certain level of protection following the initial infection. Protective immunity against re-infection coincided with higher Chlamydia-specific IgG and IgA antibody titers in sera and vaginal secretions, higher proliferative responses of peripheral blood mononuclear cells (PBMC), higher percentages of blood B lymphocytes, monocytes and CD8⁺ T cells and upregulated production of IFN-γ and IL-10 by PBMC.

Chlamydial pathogenicity in the upper genital tract relies on chlamydial ascending from the lower genital tract. To monitor chlamydial ascension, we engineered a luciferase-expressing C. muridarum. In cells infected with the luciferase-expressing C. muridarum, luciferase gene expression and enzymatic activity (measured as bioluminescence intensity) correlated well along the infection course, suggesting that bioluminescence can be used for monitoring chlamydial replication. Following an intravaginal inoculation with the luciferase-expressing C. muridarum, 8 of 10 mice displayed bioluminescence signal in the lower with 4 also in the upper genital tracts on day 3 after infection. By day 7, all 10 mice developed bioluminescence signal in the upper genital tracts. The bioluminescence signal was maintained in the upper genital tract in 6 and 2 mice by days 14 and 21, respectively. The bioluminescence signal was no longer detectable in any of the mice by day 28. The whole body imaging approach also revealed an unexpected airway infection following the intravaginal inoculation. Although the concomitant airway infection was transient and did not significantly alter the genital tract infection time courses, caution should be taken during data interpretation. The above observations have demonstrated that C. muridarum can not only achieve rapid ascending infection in the genital tract but also cause airway infection following a genital tract inoculation. These findings have laid a foundation for further optimizing the C. muridarum intravaginal infection murine model for understanding chlamydial pathogenic mechanisms.

Chlamydia trachomatis is the most common sexually transmitted bacterial infection worldwide. The impact of this pathogen on human reproduction has intensified research efforts to better understand chlamydial infection and pathogenesis. Whilst there are animal models available that mimic many aspects of human chlamydial infection, the mouse is regarded as the most practical and widely used of the models. Studies in mice have greatly contributed to our understanding of the host-pathogen interaction and provided an excellent medium for evaluating vaccines. Here we explore the advantages and disadvantages of all animal models of chlamydial genital tract infection, with a focus on the murine model and what we have learnt from it so far.

Objective: Chlamydia trachomatis infections are a significant cause of reproductive tract pathology. Protective and pathological immune mediators must be differentiated to design a safe and effective vaccine., Methods: Wild-type mice and mice deficient in IL-22 and IL-23 were infected intravaginally with Chlamydia muridarum, and their course of infection and oviduct pathology were compared. Local genital tract and draining lymph node immune responses were also examined in IL-23-deficient mice., Results: IL-22- and IL-23-deficient mice exhibited normal susceptibility to infection and oviduct pathology. IL-23 was required for the development of a Chlamydia-specific Th17 response in the lymph nodes and for production of IL-22 and IL-17 in the genital tract. However, influx of Th1 and innate immune cells was not compromised in the absence of IL-23., Conclusion: IL-22 and IL-23 play either redundant or minimal roles in the pathogenesis of Chlamydia infection in the mouse model. Induction of Th17-associated cytokines by a Chlamydia vaccine should be avoided as these responses are not central to resolution of infection and have pathologic potential., (© 2013 John Wiley & Sons Ltd.)

Background/purpose(s): Genital infection by Chlamydia trachomatis (CT) is the most common bacterial sexually transmitted disease worldwide. The infection can cause serious reproductive health complications including pelvic inflammatory disease and infertility. Stress is implicated as a risk factor for various infections; however, its effect on Chlamydia genital infection and complications are unknown., Methods: We investigated the effect of cold-stress on resistance to Chlamydia genital infection, stress hormone production, and the functions of immune cells in a mouse model. Mice were infected intravaginally with CT after a 24-day cold-stress application. The course of infection was monitored by cervicovaginal swabbing for isolation of live Chlamydia in tissue culture. The production of stress hormones and cytokines in genital tracts, spleen or blood were assessed., Results: Exposure of mice to 24-day stress resulted in: (a) increased susceptibility to Chlamydia genital infection and greater intensity of infection, (b) increased plasma or tissue noradrenaline and adrenaline levels, and (c) decreased mRNA and protein levels of major cytokines and chemokines in the spleen and genital tract., Conclusion: These results suggest that cold-induced stress induces the production of catecholamines, which may play a critical role in the modulation of the immune system leading to increased susceptibility and greater intensity of Chlamydia genital infection that could promote the development of complications., (Copyright © 2012. Published by Elsevier B.V.)

B cells can contribute to acquired immunity against intracellular bacteria, but do not usually participate in primary clearance. Here, we examined the endogenous CD4 T cell response to genital infection with Chlamydia muridarum using MHC class-II tetramers. Chlamydia-specific CD4 T cells expanded rapidly and persisted as a stable memory pool for several months after infection. While most lymph node Chlamydia-specific CD4 T cells expressed T-bet, a small percentage co-expressed Foxp3, and RORγt-expressing T cells were enriched within the reproductive tract. Local Chlamydia-specific CD4 T cell priming was markedly reduced in mice lacking B cells, and bacteria were able to disseminate to the peritoneal cavity, initiating a cellular infiltrate and ascites. However, bacterial dissemination also coincided with elevated systemic Chlamydia-specific CD4 T cell responses and resolution of primary infection. Together, these data reveal heterogeneity in pathogen-specific CD4 T cell responses within the genital tract and an unexpected requirement for B cells in regulating local T cell activation and bacterial dissemination during genital infection.

IL-17 is believed to be important for protection against extracellular pathogens, where clearance is dependent on neutrophil recruitment and local activation of epithelial cell defences. However, the role of IL-17 in protection against intracellular pathogens such as Chlamydia is less clear. We have compared (i) the course of natural genital tract C. muridarum infection, (ii) the development of oviduct pathology and (iii) the development of vaccine-induced immunity against infection in wild type (WT) BALB/c and IL-17 knockout mice (IL-17-/-) to determine if IL-17-mediated immunity is implicated in the development of infection-induced pathology and/or protection. Both the magnitude and duration of genital infection was significantly reduced in IL-17-/- mice compared to BALB/c. Similarly, hydrosalpinx was also greatly reduced in IL-17-/- mice and this correlated with reduced neutrophil and macrophage infiltration of oviduct tissues. Matrix metalloproteinase (MMP) 9 and MMP2 were increased in WT oviducts compared to IL-17-/- animals at day 7 post-infection. In contrast, oviducts from IL-17-/- mice contained higher MMP9 and MMP2 at day 21. Infection also elicited higher levels of Chlamydia-neutralizing antibody in serum of IL-17-/- mice than WT mice. Following intranasal immunization with C. muridarumMajor Outer Membrane Protein (MOMP) and cholera toxin plus CpG adjuvants, significantly higher levels of chlamydial MOMP-specific IgG and IgA were found in serum and vaginal washes of IL-17-/- mice. T cell proliferation and IFNγ production by splenocytes was greater in WT animals following in vitro re-stimulation, however vaccination was only effective at reducing infection in WT, not IL-17-/- mice. Intranasal or transcutaneous immunization protected WT but not IL-17-/- mice against hydrosalpinx development. Our data show that in the absence of IL-17, the severity of C. muridarum genital infection and associated oviduct pathology are significantly attenuated, however neither infection or pathology can be reduced further by vaccination protocols that effectively protect WT mice.

Aim: The study tries to highlight the implication of CT infection in male infertility., Material and Methods: There were 857 male patients enrolled in the study. The male cases have been investigated in the Gynecology and Urology Department of Ghencea Medical Center, Bucharest. All the selected cases have been done a spermogram test and for the diagnosis of the Chlamydia Trachomatis infection, the ELISA test was used. The test detects the presence in the serum of the anti-chlamydia specific antibodies of type IgA, IgM, IgG (BAG-Chlamydia-AIA)., Results and Discussions: From the total of the investigated subjects, only 233 cases had a modified spermogram. According to the specialty literature, the human factor is involved in 35% of the infertility cases, the male one in 30%, in 20% both factors are involved and in 15% of the cases, there is no incriminating cause after complete investigations. Accordingly, the study supports the literature data.

Chlamydia trachomatis is a Gram-negative obligate intracellular bacterial pathogen. It is the leading cause of bacterial sexually transmitted disease in the world, with more than 100 million new cases of genital tract infections with C. trachomatis occurring each year. Animal models are indispensable for the study of C. trachomatis infections and the development and evaluation of candidate vaccines. In this paper, the most commonly used animal models to study female genital tract infections with C. trachomatis will be reviewed, namely, the mouse, guinea pig, and nonhuman primate models. Additionally, we will focus on the more recently developed pig model.

Intravaginal infection with C. muridarum in mice often results in hydrosalpinx similar to that found in women urogenitally infected with C. trachomatis, making the C. muridarum lower genital tract infection murine model suitable for studying C. trachomatis pathogenesis. To our surprise, DBA1/j mice were highly resistant to hydrosalpinx following an intravaginal infection with C. muridarum although these mice were as susceptible to lower genital tract infection as other mouse strains. A significantly lower level of C. muridarum organisms was recovered from the oviduct of DBA1/j mice, correlating the resistance to hydrosalpinx with reduced ascension of C. muridarum to the oviduct. The DBA1/j resistance to hydrosalpinx was effectively overcome by intracervical inoculation with C. muridarum. The intracervically inoculated DBA1/j mice developed severe hydrosalpinx with the highest levels of live C. muridarum organisms recovered from uterine tissue on day 3 and oviduct tissue on day 7 post inoculation while in intravaginally inoculated DBA1/j mice, the peak of live organism recovery from uterine tissue was delayed to day 7 with no rise in the amount of live organisms recovered from the oviduct. These observations have not only validated the correlation between hydrosalpinx and live organism invasion in the oviduct but also demonstrated that the intracervical inoculation, by promoting rapid chlamydial replication in the uterine epithelial cells and ascension to the oviduct of DBA1/j mice, may be used for further understanding chlamydial pathogenic mechanisms. The above findings also suggest that strategies aimed at reducing tubal infection may be most effective in blocking tubal pathology.

We previously reported that 5 Chlamydia muridarum antigens reacted with antisera from >90% mice urogenitally infected with C. muridarum and they are TC0660 (ABC transporter or ArtJ), TC0727 (outer membrane complex protein B or OmcB), TC0828 (macrophage infectivity potentiator or MIP), TC0726 (inclusion membrane protein or Inc) & TC0268 (hypothetical protein or HP). The orthologs of these antigens in Chlamydia trachomatis were also highly reactive with antisera from women urogenitally infected with C. trachomatis. In the current study, we evaluated these C. muridarum antigens for their ability to induce protection against a C. muridarum intravaginal challenge infection in mice. We found that only MIP induced the most pronounced protection against C. muridarum infection. The protection correlated well with robust C. muridarum MIP-specific antibody and Th1-dominant T cell responses. The MIP-immunized mice displayed significantly reduced live organism shedding from the lower genital tract and highly attenuated inflammatory pathologies in the upper genital tissues. These results demonstrate that MIP, an immunodominant antigen identified by both human and mouse antisera, may be considered a component of a multi-subunit chlamydial vaccine for inducing protective immunity., (Copyright © 2013 Institut Pasteur. All rights reserved.)

Chlamydia trachomatis is the most common bacterial sexually transmitted disease worldwide and leads to serious pathological sequelae in the upper genital tract (UGT) including pelvic inflammatory disease, ectopic pregnancy, and infertility. Several components of the host immune responses have been shown to contribute to the UGT pathology following genital chlamydial infection. We have shown recently that CD8(+) T cells induce the chlamydial UGT pathology via the production of TNF-α. However, those studies did not determine whether the pathology is mediated by bystander or antigen-specific CD8(+) T cells. In this study, we compared chlamydial clearance and UGT pathology in OT-1 transgenic mice and the corresponding C57BL/6J wild-type mice following primary intravaginal Chlamydia muridarum infection. All CD8(+) T cells in the OT-1 mice respond only to the Ova 257-264 peptide and are incapable of responding to other antigenic epitopes including those of Chlamydia. OT-1 mice displayed vaginal chlamydial clearance comparable to the wild-type animals. However, both oviduct and uterine horn pathology were minimal in the OT-1 mice compared with the high degree of pathology observed in the wild-type animals. These results strongly suggest that Chlamydia-specific, not bystander, CD8(+) T cells mediate the UGT pathological sequelae following genital chlamydial infection., (© 2013 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.)

We have identified a CD8⁺CXCR5⁺ T cell that prevents the development of oviduct dilation following C. muridarum genital infection. Phenotypic studies show that CD8⁺CXCR5⁺ cells express markers of T regulatory cells (FoxP3, CD25, and GITR) but do not express a necessary component of cytotoxic cells (perforin). Cxcr5⁻/⁻ mice have significantly lower numbers of CD8⁺ cells and lack the CD8⁺CXCR5⁺ population while the total number of CD4⁺ cells is equivalent between mouse strains. The transfer of CD8⁺ splenocytes from WT mice reduces the oviduct dilation seen in Cxcr5⁻/⁻ mice following C. muridarum infection. Future studies will investigate the mechanism by which this cell type regulates genital tract pathology.

Urogenital infection with Chlamydia trachomatis in some women can lead to upper genital tract pathologies, such as hydrosalpinx, potentially affecting fertility. In the current study, 27 of 40 mice intravaginally infected with Chlamydia muridarum developed visible hydrosalpinges in the oviduct while the remaining 13 did not, although all infected mice displayed similar infection time courses. Antisera from the 40 mice recognized 130 out of 257 C. muridarum proteins as antigens and 17 as immunodominant antigens. Importantly, the 27 mice with hydrosalpinges preferentially recognized two C. muridarum proteins (TC0582 and TC0912, designated pathology-associated antigens) while the 13 mice with no hydrosalpinx preferentially recognized 10 proteins (TC0047, TC0117, TC0190, TC0197, TC0257, TC0279, TC0326, TC0630, TC0689, and TC0816, designated nonpathology antigens). The preferential recognition was validated by absorption and independently confirmed in Western blots. The C. trachomatis homolog of TC0912 is encoded by a highly polymorphic gene that is associated with ocular pathogenesis. A fragment of TC0912 was found to improve the differentiation of hydrosalpinx from nonhydrosalpinx mice. TC0582 is a highly conserved ATP synthase, and it may contribute to chlamydial pathogenesis via mechanisms similar to those hypothesized for the highly conserved HSP60. Thus, we have identified chlamydial antigens and epitopes that are associated with either susceptibility or resistance to upper genital tract pathology, which will help us to further understand chlamydial pathogenesis and to develop anti-Chlamydia subunit vaccines.

Loss of the conserved "cryptic" plasmid from C. trachomatis and C. muridarum is pleiotropic, resulting in reduced innate inflammatory activation via TLR2, glycogen accumulation and infectivity. The more genetically distant C. caviae GPIC is a natural pathogen of guinea pigs and induces upper genital tract pathology when inoculated intravaginally, modeling human disease. To examine the contribution of pCpGP1 to C. caviae pathogenesis, a cured derivative of GPIC, strain CC13, was derived and evaluated in vitro and in vivo. Transcriptional profiling of CC13 revealed only partial conservation of previously identified plasmid-responsive chromosomal loci (PRCL) in C. caviae. However, 2-deoxyglucose (2DG) treatment of GPIC and CC13 resulted in reduced transcription of all identified PRCL, including glgA, indicating the presence of a plasmid-independent glucose response in this species. In contrast to plasmid-cured C. muridarum and C. trachomatis, plasmid-cured C. caviae strain CC13 signaled via TLR2 in vitro and elicited cytokine production in vivo similar to wild-type C. caviae. Furthermore, inflammatory pathology induced by infection of guinea pigs with CC13 was similar to that induced by GPIC, although we observed more rapid resolution of CC13 infection in estrogen-treated guinea pigs. These data indicate that either the plasmid is not involved in expression or regulation of virulence in C. caviae or that redundant effectors prevent these phenotypic changes from being observed in C. caviae plasmid-cured strains.

Background: Regulation of immune responses is critical for controlling inflammation and disruption of this process can lead to tissue damage. We reported that CXCL13 was induced in fallopian tube tissue following C. trachomatis infection. Here, we examined the influence of the CXCL13-CXCR5 axis in chlamydial genital infection., Methodology and Principal Findings: Disruption of the CXCL13-CXCR5 axis by injecting anti-CXCL13 Ab to BALB/c mice or using Cxcr5-/- mice increased chronic inflammation in the upper genital tract (UGT; uterine horns and oviducts) after Chlamydia muridarum genital infection (GT). Further studies in Cxcr5-/- mice showed an elevation in bacterial burden in the GT and increased numbers of neutrophils, activated DCs and activated NKT cells early after infection. After resolution, we noted increased fibrosis and the accumulation of a variety of T cells subsets (CD4-IFNγ, CD4-IL-17, CD4-IL-10 & CD8-TNFα) in the oviducts. NKT cell depletion in vitro reduced IL-17α and various cytokines and chemokines, suggesting that activated NKT cells modulate neutrophils and DCs through cytokine/chemokine secretion. Further, chlamydial glycolipids directly activated two distinct types of NKT cell hybridomas in a cell-free CD1d presentation assay and genital infection of Cd1d-/- mice showed reduced oviduct inflammation compared to WT mice. CXCR5 involvement in pathology was also noted using single-nucleotide polymorphism analysis in C. trachomatis infected women attending a sub-fertility clinic. Women who developed tubal pathology after a C. trachomatis infection had a decrease in the frequency of CXCR5 SNP +10950 T>C (rs3922)., Conclusions/significance: These experiments indicate that disruption of the CXCL13-CXCR5 axis permits increased activation of NKT cells by type I and type II glycolipids of Chlamydia muridarum and results in UGT pathology potentially through increased numbers of neutrophils and T cell subsets associated with UGT pathology. In addition, CXCR5 appears to contribute to inter-individual differences in human tubal pathology following C. trachomatis infection.

Background: Although Tim-3 & PD-L1 signaling pathways play important roles in negatively regulating immune responses, their roles in chlamydial infection have not been evaluated., Methods: Neutralization antibodies targeting Tim-3 and PD-L1 were used to treat mice. Following an intravaginal infection with C. muridarum organisms, mice with or without the dual antibody treatment were compared for live chlamydial organism shedding from the lower genital tract and inflammatory pathology in the upper genital tract., Results: Mice treated with anti-Tim-3 and anti-PD-L1 antibodies displayed a time course of live organism shedding similar to that of mice treated with equivalent amounts of isotype-matched IgG molecules. The combined antibody blocking failed to alter either the lower genital tract cytokine or systemic humoral and cellular adaptive responses to C. muridarum infection. However, the antibody blocking significantly enhanced C. muridarum-induced pathologies in the upper genital tract, including more significant hydrosalpinx and inflammatory infiltration in uterine horn and oviduct tissues., Conclusions: The Tim-3 and PD-L1-mediated signaling can significantly reduce pathologies in the upper genital tract without suppressing immunity against chlamydial infection, suggesting that Tim-3 and PD-L1-mediated negative regulation may be manipulated to attenuate tubal pathologies in women persistently infected with C. trachomatis organisms.