Rectal Chlamydia trachomatis Infection: A Narrative Review of the State of the Science and Research Priorities.

ABSTRACT: Chlamydia trachomatis (CT) is the most commonly reported infection in the United States. Most chlamydial research to date has focused on urogenital infection, but a growing body of research has demonstrated that rectal chlamydia is a relatively common infection among clinic-attending men and women. We know that most rectal CT infections are asymptomatic, but the health implications of these infections, particularly for women, are unclear. In addition, there are key knowledge gaps related to the epidemiologic parameters of rectal chlamydia, the routes of acquisition, the duration of infection, and the clinical significance of a positive rectal CT test result. This lack of information has led to a blind spot in the potential role of rectal chlamydia in sustaining high levels of CT transmission in the United States. Furthermore, recent findings from animal models suggest that the immune response generated from gastrointestinal chlamydial infection can protect against urogenital infection; however, it remains to be determined whether rectal chlamydia similarly modulates anti-CT immunity in humans. This is a critical question in the context of ongoing efforts to develop a CT vaccine. In this narrative review, we summarize the state of the science for rectal chlamydia and discuss the key outstanding questions and research priorities in this neglected area of sexual health research.

Use of gene sequences as type for naming prokaryotes: Recommendations of the international committee on the taxonomy of chlamydiae.

The International Committee on Systematics of Prokaryotes (ICSP) discussed and rejected in 2020 a proposal to modify the International Code of Nomenclature of Prokaryotes to allow the use of gene sequences as type for naming prokaryotes. An alternative nomenclatural code, the Code of Nomenclature of Prokaryotes Described from Sequence Data (SeqCode), which considers genome sequences as type material for naming species, was published in 2022. Members of the ICSP subcommittee for the taxonomy of the phylum Chlamydiae (Chlamydiota) consider that the use of gene sequences as type would benefit the taxonomy of microorganisms that are difficult to culture such as the chlamydiae and other strictly intracellular bacteria. We recommend the registration of new names of uncultured prokaryotes in the SeqCode registry.

Anti-chlamydial antibiotic therapy for symptom improvement in peripheral artery disease: prospective evaluation of rifalazil effect on vascular symptoms of intermittent claudication and other endpoints in Chlamydia pneumoniae seropositive patients (PROVIDENCE-1).

BACKGROUND: A potentially strong association exists between Chlamydia pneumoniae and atherosclerosis, but the clinical benefits of antibiotic therapy have not been demonstrated. Preliminary studies of antibiotic therapy in peripheral artery disease have shown a decreased need for revascularization and improved walking ability. The objective of this phase-III trial was to assess the effect of a potent anti-Chlamydial agent, rifalazil, on peak walking time in patients with symptomatic peripheral artery disease. METHODS AND RESULTS: Patients with intermittent claudication secondary to peripheral artery disease who were seropositive for C pneumoniae were randomized to 25 mg rifalazil once weekly for 8 weeks or matching placebo. Two hundred ninety-seven patients were enrolled from 3 countries and were followed up for 1 year. The mean+/-SD ankle brachial index at baseline was 0.63+/-0.16. The primary end point, change from baseline in log peak walking time on a graded treadmill, was assessed 180 days after randomization. Secondary end points included changes in claudication onset time and quality of life, assessed with the Walking Impairment Questionnaire and the Short Form Medical Outcomes 36. No benefit of rifalazil therapy was found in the primary or any secondary end point among this cohort of patients with peripheral artery disease. The group treated with rifalazil improved their peak walking times by 23% (95% confidence interval, 15 to 31) from baseline to day 180, whereas the placebo group improved by 18% (95% confidence interval, 11 to 26; P=0.38). Peak walking time, claudication onset time, Walking Impairment Questionnaire, and Short Form Medical Outcomes 36 showed no treatment-by-time interaction during the 360-day study period. Thirty-two adjudicated cardiovascular events occurred, 16 in each treatment group. CONCLUSIONS: Rifalazil did not improve exercise performance or quality of life in patients with intermittent claudication. No safety concerns were identified. Given the very small effect size, it is unlikely that larger studies would demonstrate a symptomatic benefit of this therapy in peripheral artery disease.

Amphiphysin IIm is required for survival of Chlamydia pneumoniae in macrophages.

Macrophages play a critical role in both innate and acquired immunity because of their unique ability to internalize, kill, and degrade bacterial pathogens through the process of phagocytosis. The adaptor protein, amphiphysin IIm, participates in phagocytosis and is transiently associated with early phagosomes. Certain pathogens, including Chlamydia pneumoniae, have evolved mechanisms to subvert macrophage phagosome maturation and, thus, are able to survive within these cells. We report here that, although amphiphysin IIm is usually only transiently associated with the phagosome, it is indefinitely retained on vacuoles containing C. pneumoniae. Under these wild-type conditions, C. pneumoniae do not elicit significant nitric oxide (NO) production and are not killed. Abrogation of amphiphysin IIm function results in C. pneumoniae-induced NO production and in the sterilization of the vacuole. The data suggest that C. pneumoniae retains amphiphysin IIm on the vacuole to survive within the macrophage.

High frequency of Chlamydia pneumoniae and risk factors in children with acute respiratory infection.

This study was performed as a contribution for a better understanding of Chlamydia pneumoniae frequency in children with respiratory infections. A total of 416 children were recruited from two clinical centers in Sao Luis, Brazil. Of these patients, 165 children had upper respiratory tract infections (URTI), 150 had community-acquired pneumonia (CAP), and 101 were asymptomatic volunteer children. Clinical and epidemiological data from the participants were recorded. Nasopharyngeal swab samples were collected to extract DNA. C. pneumoniae DNA positivity and copy numbers were obtained by an absolute quantitative real-time PCR method. RESULTS: Positivity for C. pneumoniae DNA was higher in samples from URTI children (38.2%) and from CAP children (18.0%) than in those from the control group (7.9%; p < 0.001). Moreover, C. pneumoniae DNA was denser in children with URTI than in asymptomatic children. Considering the cutoff, the highest value of C. pneumoniae DNA found in asymptomatic children of the 3.98 log10 copies/mL, 8.5% (14/165) of the children with URTI, and 3.3% (5/150) with CAP presented high copy numbers of C. pneumoniae DNA. CONCLUSION: Taken together, these results revealed a high frequency of C. pneumoniae in both children with URTI and CAP.

Chlamydia pneumoniae infection increases adherence of mouse macrophages to mouse endothelial cells in vitro and to aortas ex vivo.

Interactions between monocytes/macrophages and endothelial cells play an important role in the pathogenesis of atherosclerosis, and the adherence of monocytes to the arterial endothelium is one of the early events in atherogenesis. In the present study, peritoneal macrophages harvested from green fluorescent protein (GFP) transgenic mice were used to analyze how Chlamydia pneumoniae infection affects the adherence of GFP-macrophages to mouse endothelial cells in vitro and to the aorta from normolipidemic and hyperlipidemic mice ex vivo. In vitro studies showed that C. pneumoniae-infected GFP-macrophages adhered better than uninfected macrophages to endothelial cells and GFP-macrophages adhered better to infected than uninfected endothelial cells. The ex vivo studies showed that C. pneumoniae-infected macrophages adhered better than uninfected macrophages to aortas from both normolipidemic and hyperlipidemic C57BL/6J mice and apolipoprotein E (ApoE)-deficient mice. In contrast, adherence of C. pneumoniae-infected macrophages to the aortas of intercellular adhesion molecule 1 (ICAM-1) knockout mice was not enhanced, suggesting that ICAM-1 is crucial for activation of the adherence of C. pneumoniae-infected macrophages to the endothelium. In conclusion, the present study defined a homing mechanism by which C. pneumoniae promotes the adherence of mononuclear phagocytes to the endothelium at the site of atherosclerotic lesion formation to promote the progression of atherosclerosis.

Identification and characterization of Chlamydia pneumoniae-specific proteins that activate tumor necrosis factor alpha production in RAW 264.7 murine macrophages.

Chlamydia pneumoniae is a common respiratory pathogen, which activates macrophages to induce inflammatory cytokines that may promote atherosclerosis. However, the antigens that induce macrophage activation have not been well defined. In the current study, three chlamydial proteins which are recognized during human infection, outer membrane protein 2 (OMP2) and two 53-kDa proteins (Cpn 0980 and Cpn 0809), were investigated to determine whether they activate macrophages and, if they do, what mechanism they use for this activation. It was shown that these three proteins could (i) induce expression of tumor necrosis factor alpha (TNF-alpha) and tissue factor and (ii) induce phosphorylation of p44/42 mitogen-activated protein kinases (MAPK) and activation of early growth response factor 1 (Egr-1). Control proteins, the N-terminal fragment of polymorphic membrane protein 8 and the thioredoxin portion of the fusion protein, had no effect on macrophages. Treatment of cells with a MEK1/2 inhibitor, U0126, dramatically reduced the phosphorylation of ERK, activation of Egr-1, and expression of TNF-alpha in macrophages treated with recombinant proteins. Toll-like receptors (TLRs) act as sensors for microbial antigens and can signal via the MAPK pathway. Chlamydial protein-induced expression of TNF-alpha was significantly reduced in macrophages lacking TLR2 or TLR4. These findings suggest that C. pneumoniae may activate macrophages through OMP2, Cpn 0980, and Cpn 0809 in addition to cHSP60 and that activation occurs via TLR2 or TLR4, Egr-1, and MAPK pathways.

Retinoic acid prevents Chlamydia pneumoniae-induced foam cell development in a mouse model of atherosclerosis.

Chlamydia pneumoniae, a common respiratory pathogen, has been associated with cardiovascular disease. C. pneumoniae infection accelerates atherosclerotic lesion development in hyperlipidemic animals. Retinoic acid, an anti-oxidant, inhibits infection of endothelial cells by C. pneumoniae. The present study demonstrated that retinoic acid suppresses the acceleration of foam cell lesion development induced by C. pneumoniae in hyperlipidemic C57BL/6J mice. Retinoic acid treatment had no effect on foam cell lesion development in uninfected animals. Lung infection and duration was decreased in treated mice, suggesting one mechanism by which retinoic acid reduces C. pneumoniae-accelerated foam cell lesion formation in hyperlipidemic mice.

Efficacy of benzoxazinorifamycins in a mouse model of Chlamydia pneumoniae lung infection.

The efficacy of rifalazil and other benzoxazinorifamycins was tested in a mouse model of lung infection against Chlamydia pneumoniae. Rifalazil and six related new chemical entities all showed efficacy after one dose per day for 3 days at either 3 or 1 mg/kg of body weight.

Punica granatum L. Leaf Extract Attenuates Lung Inflammation in Mice with Acute Lung Injury.

The hydroalcoholic extract of Punica granatum (pomegranate) leaves was previously demonstrated to be anti-inflammatory in a rat model of lipopolysaccharide- (LPS-) induced acute peritonitis. Here, we investigated the anti-inflammatory effects of the ethyl acetate fraction obtained from the pomegranate leaf hydroalcoholic extract (EAFPg) on the LPS-induced acute lung injury (ALI) mouse model. Male Swiss mice received either EAFPg at different doses or dexamethasone (per os) prior to LPS intranasal instillation. Vehicle-treated mice were used as controls. Animals were culled at 4 h after LPS challenge, and the bronchoalveolar lavage fluid (BALF) and lung samples were collected for analysis. EAFPg and kaempferol effects on NO and cytokine production by LPS-stimulated RAW 264.7 macrophages were also investigated. Pretreatment with EAFPg (100-300 mg/kg) markedly reduced cell accumulation (specially neutrophils) and collagen deposition in the lungs of ALI mice. The same animals presented with reduced lung and BALF TNF-α and IL-1ß expression in comparison with vehicle controls (p < 0.05). Additionally, incubation with either EAFPg or kaempferol (100 µg/ml) reduced NO production and cytokine gene expression in cultured LPS-treated RAW 264.7 macrophages. Overall, these results demonstrate that the prophylactic treatment with EAFPg attenuates acute lung inflammation. We suggest this fraction may be useful in treating ALI.

Inoculation of Chlamydia pneumoniae or Chlamydia trachomatis with ligands that inhibit attachment to host cells reduces infectivity in the mouse model of lung infection: implication for anti-adhesive therapy.

Previous studies have shown that the chlamydial glycan contains a high-mannose oligosaccharide, which mediates attachment and infectivity of the organism. Removal of the glycan decreases infectivity in vitro and in vivo. The present study demonstrates that simultaneous inoculation of chlamydial organisms and a ligand that prevents glycan binding reduces lung burden in infected animals.

Chlamydia pneumoniae--an infectious risk factor for atherosclerosis?

Cardiovascular disease, of which atherosclerosis is an important component, is the leading cause of death in the western world. Although there are well-defined risk factors for atherosclerosis, these factors do not account for all incidences of the disease. Because atherosclerotic processes are typified by chronic inflammatory responses, which are similar to those that are elicited by chronic infection, the role of infection in promoting or accelerating atherosclerosis has received renewed attention. This review focuses on the accumulating evidence that chronic infection with Chlamydia pneumoniae, a ubiquitous human respiratory pathogen, might contribute to atherosclerotic lesion progression.

Chlamydia pneumoniae induces tissue factor expression in mouse macrophages via activation of Egr-1 and the MEK-ERK1/2 pathway.

Recent studies have suggested that infection with Chlamydia pneumoniae (C pneumoniae) may contribute to the instability of atherosclerotic plaques and thrombosis and is associated with acute coronary events. Tissue factor (TF), a potent prothrombotic molecule, is expressed by macrophages and other cell types within atherosclerotic lesions and plays an essential role in thrombus formation after plaque rupture. Therefore the effects of C pneumoniae on induction of TF expression in macrophages were investigated. Infection of RAW mouse macrophages with C pneumoniae induced a time-dependent increase in procoagulant activity, expression of TF protein, and TF mRNA. C pneumoniae infection stimulated increased binding of nuclear proteins to the consensus DNA sequence for Egr-1, a key response element within the TF promoter, and increased the expression of Egr-1 protein. Transient transfections of RAW cells with mutated TF promoter constructs showed that the Egr-1 binding region is an important transcriptional regulator of C pneumoniae-induced TF expression. Furthermore, C pneumoniae-stimulated phosphorylation of ERK1/2 and Elk-1 and pharmacological inhibition of mitogen-activated protein kinase activity reduced the expression of TF and Egr-1. Antibody and polymyxin B blocking of the Toll-like receptor 4 (TLR4) partially reduced the C pneumoniae-induced expression of TF and Egr-1. In conclusion, the C pneumoniae-induced increase in TF expression in macrophages is mediated in part by Egr-1, signaling through TLR4, and activation of the MEK-ERK1/2 pathway.

Effects of Murine Norovirus on Chlamydia pneumoniae-Accelerated Atherosclerosis in ApoE(-/-) Mice.

Chlamydia pneumoniae (Cpn), a common respiratory pathogen of humans, is associated with human cardiovascular disease and the acceleration of atherosclerosis in hyperlipidemic animal models. Our laboratory has demonstrated that murine norovirus (MNV), a prevalent infection of laboratory mice, can unpredictably alter atherosclerosis in hyperlipidemic Ldlr(-/-) and ApoE(-/-) mice. Given that MNV has a tropism for macrophages and may exacerbate atherogenesis, we investigated whether coinfection with MNV and Cpn might alter macrophage phenotypes in vitro and atherosclerosis in ApoE(-/-) mice. In the presence of oxidized low-density lipoprotein, coinfection of ApoE(-/-) bone marrow-derived macrophages (BMDM) with MNV and Cpn resulted in significant increases in gene expression of IL6, MCP1, iNOS, and TNFα compared with Cpn-monoinfected BMDM. On the basis of these findings, we hypothesized that concurrent MNV-Cpn infection might increase plaque lesion size in vivo. As expected, Cpn monoinfection of ApoE(-/-) mice increased mean plaque size by 62% compared with that in uninfected mice. However, MNV did not significantly alter plaque lesion size in MNV-Cpn-coinfected mice compared with Cpn-monoinfected mice. There were no differences in aortic cytokines locally at the site of plaque development or in peritoneal macrophages at 1 wk after infection in MNV-Cpn-coinfected mice compared with Cpn-monoinfected mice. MNV was not detected in the aortic tissue of MNV-infected mice at 1 or 8 wk after infection regardless of Cpn status. These data suggest that MNV infection does not appreciably alter plaque development in Cpn-accelerated atherosclerosis in ApoE(-/-) mice.

Foam cell formation inhibits growth of Chlamydia pneumoniae but does not attenuate Chlamydia pneumoniae-induced secretion of proinflammatory cytokines.

BACKGROUND: It has not yet been determined whether lipid-loaded macrophages (foam cells), a major cellular component of atherosclerotic lesions, have the capacity to support growth of Chlamydia pneumoniae and be activated to secrete proinflammatory cytokines in response to C pneumoniae infection. METHODS AND RESULTS: Lipid loading of RAW 264.7 cells and mouse peritoneal macrophages with either oxidized or acetylated LDL significantly inhibits the growth of C pneumoniae. Modified forms of LDL are not directly toxic to C pneumoniae and do not inhibit either the initial binding or internalization of C pneumoniae by macrophages. Lipid loading does not reduce infection of macrophages with Chlamydia trachomatis. Treatment of lipid-loaded macrophages with live, heat-killed, or UV-inactivated C pneumoniae stimulates secretion of cytokines. C pneumoniae also induces expression of the mRNA for tumor necrosis factor-alpha in foam cells despite inhibition of nuclear factor-kappaB binding to DNA by prior treatment with oxidized LDL. CONCLUSIONS: Foam cell formation is not conducive to growth of C pneumoniae but does not inhibit the C pneumoniae-induced secretion of proinflammatory cytokines.

Nitric oxide synthase plays a role in Chlamydia pneumoniae-induced atherosclerosis.

OBJECTIVE: Chlamydia pneumoniae infection has been associated with atherosclerosis, although the mechanisms by which C. pneumoniae contribute to atherogenesis remain unclear. Altered production of nitric oxide, a known bactericidal and anti-inflammatory agent, represents one possible mechanistic link. To examine this issue, a diet-induced, hyperlipidemic mouse model of early atherosclerosis was used. METHODS: A series of intranasal inoculations of C. pneumoniae strain AR-39 were administered to mice lacking endothelial or inducible nitric oxide synthase and to normal controls. After 18 weeks on an atherogenic diet, atherosclerotic lesion area in the aortic sinus was measured using computer-assisted morphometry. RESULTS: In the absence of C. pneumoniae infection, diet-fed eNOS(-/-) mice developed enlarged fatty streak lesions of borderline significance in comparison to uninfected, wild-type mice, while the lesion area in uninfected, diet-fed iNOS(-/-) mice did not differ significantly from lesion area in wild-type animals. In contrast, lesion area in infected eNOS(-/-) mice increased slightly, but not significantly in comparison to uninfected eNOS(-/-) mice. Lesion area in the infected iNOS(-/-) mice was significantly enlarged when compared to both uninfected iNOS(-/-) mice as well as to infected wild-type mice. CONCLUSIONS: These data suggest that production of nitric oxide by eNOS protects against development of fatty streak lesions in uninfected hyperlipidemic mice, but does not offer additional protection in infected hyperlipidemic mice, while iNOS may play a protective role, thus limiting chlamydial exacerbation of fatty streak lesions.

Chronic inhibition of cyclooxygenase-2 does not alter plaque composition in a mouse model of advanced unstable atherosclerosis.

OBJECTIVE: Inflammation contributes to atherosclerotic plaque initiation and progression. Recent studies suggest that anti-inflammatory drugs such as cyclooxygenase-2 (Cox-2) inhibitors have anti-atherogenic effects. The current study was designed to investigate whether administration of a Cox-2 inhibitor to older apolipoprotein E deficient (apo E-/-) mice with established lesions alters the composition and increases the stability of the lesions. METHODS AND RESULTS: The Cox-2 inhibitor Celecoxib was administered in chow to 26-week-old, male, apo E-/- mice exhibiting advanced, unstable atherosclerotic lesions within the innominate/brachiocephalic artery. Mice administered Celecoxib had no significant changes in serum cholesterol or the average cross sectional area of atherosclerotic lesion in the innominate artery after 15 weeks of treatment in comparison to non-treated control mice. Histological analyses of sections of the innominate artery demonstrated no significant changes in the frequency of markers of advanced and unstable atherosclerotic plaques, including intra-plaque hemorrhage, vascular calcification, thinning of the fibrous cap, size of the necrotic core and macrophage content. There were also no significant differences in the content of Cox-2 within the lesions. Quantitative real time polymerase chain reaction with mRNA isolated from the aorta of each mouse revealed no significant changes in the expression of tissue factor and inducible nitric oxide synthase. However, mRNA levels for MCP-1 were increased fivefold following 15 weeks of treatment with Celecoxib in comparison to non-treated control mice. CONCLUSIONS: These data suggest that Celecoxib has no effect on the composition of advanced atherosclerotic lesions in older apo E-/- mice.

Infection and Atherosclerosis Development.

Atherosclerosis is a chronic disease hallmarked by chronic inflammation, endothelial dysfunction and lipid accumulation in the vasculature. Although lipid modification and deposition are thought to be a major source of the continuous inflammatory stimulus, a large body of evidence suggests that infectious agents may contribute to atherosclerotic processes. This could occur by either direct effects through infection of vascular cells and/or through indirect effects by induction of cytokine and acute phase reactant proteins by infection at other sites. Multiple bacterial and viral pathogens have been associated with atherosclerosis by seroepidemiological studies, identification of the infectious agent in human atherosclerotic tissue, and experimental studies demonstrating an acceleration of atherosclerosis following infection in animal models of atherosclerosis. This review will focus on those infectious agents for which biological plausibility has been demonstrated in animal models and on the challenges of proving a role of infection in human atherosclerotic disease.

Lesion progression and plaque composition are not altered in older apoE-/- mice lacking tumor necrosis factor-alpha receptor p55.

BACKGROUND: Inflammatory processes are an integral component of the initiation, progression, and destabilization of atherosclerotic lesions. Tumor necrosis factor-alpha (TNF-alpha) is considered a primary mediator of inflammatory processes. METHODS AND RESULTS: The role of TNF-alpha in plaque progression and plaque destabilization was investigated in the innominate arteries of older TNF-alpha receptor p55 deficient mice that were generated on a hyperlipidemic apolipoprotein E deficient background (p55-/- apoE-/-). There were no significant differences in levels of circulating cytokines, plaque progression, plaque composition or features of plaque destabilization in p55-/- apoE-/- compared to wild type (p55+/+ apoE-/-) mice. CONCLUSIONS: Progression and destabilization of advanced atherosclerotic lesions does not seem to be mediated via the TNF-alpha receptor p55.

Chlamydial infections of the cardiovascular system.

This paper presents a review on cardiovascular diseases which can be caused by chlamydial infection with the emphasis in the recent development in association between Chlamydia pneumoniae and cardiovascular disease. The review includes seroepidemiologic observations; the discovery of C. pneumoniae in atheromatous plaques; in vivo studies using animal models indicating that C. pneumoniae is a co-risk factor of hyperlipidemia for atherosclerosis; in vitro studies demonstrating putative mechanisms by which C. pneumoniae could contribute to the immunopathology of atherosclerosis; and early promising antibiotic intervention studies.