CHLAMYDIAL INFECTIONS

SUMMARY Chlamydiae are among the most successful bacterial pathogens, and there are few branches of medicine on which chlamydial infection and its sequelae do not impinge. Chlamydia trachomatis is responsible for many million cases of blindness, pelvic inflammatory disease, urethritis, epididymitis, infertility and ectopic pregnancy annually; it also causes lymphogranuloma venereum, reactive arthritis, ophthalmia neonatorum and infantile pneumonia. C. pneumoniae is among the most common causes of community-acquired pneumonia, and recent evidence suggests that it may play a part in the pathogenesis of coronary heart disease. C. psittaci is a highly prevalent zoonotic infection with a wide host range. It is of great economic importance, and causes sporadic but sometimes devastating disease in humans. Most chlamydial infections are subclinical, but even if the initial illness is mild there may be serious long-term sequelae. It is therefore important to identify and treat chlamydial infections in their early stages, but diagnosis usually depends on laboratory tests. Recent trials have shown that single doses of the long-acting macrolide azithromycin are effective in the treatment of genital and ocular C. trachomatis infection, but longer courses of antimicrobials remain the mainstay of treatment for C. pneumoniae and C. psittaci infections.     

IPAzyme Chlamydia and microimmunofluorescence tests for detecting antibodies against Chlamydia trachomatis in women undergoing laparoscopy

Superficial Chlamydia trachomatis infections are diagnosed by antigen detection whereas serological investigations are mainly performed to detect deep-seated infections. In this study the genus-specific IPAzyme and two species-specific microimmunofluorescence (MIF) tests were compared for detecting antibodies against C. trachomatis in women undergoing laparoscopy for diagnostic purposes or for ligation of the fallopian tubes. Microbiological findings were similar in both groups. C. trachomatis was detected in 4 of 38 women with ligation and in 6 of 61 women with diagnostic laparoscopy. Serum IgG antibody titres by IPAzyme were clearly positive in 24 out of 61 of the diagnostic group and in 12 out of 38 of the ligation group. However, 14 (39%) of the 36 IPAzyme-positive results were caused by antibodies against Chlamydia pneumoniae and/or Chlamydia psittaci, only 4 (11%) were caused by anti-C. trachomatis IgG and 8 (22%) were caused by both antibodies as tested by a conventional MIF; in 10 all MIF titres were lower than 1:32. A commercial MIF still containing too much genus-specific lipopolysaccharides was more sensitive, but also more crossreactive than the conventional MIF. IPAzyme IgA were only found in association with positive IPAzyme IgG results. We conclude that in case of presumed deep-seated C. trachomatis infection only sensitive species-specific assays for detecting C. trachomatis antibodies are helpful. IPAzyme and other genus-specific assays cannot be recommended to detect antibodies to C. trachomatis in the serum, because positive results are caused mainly by antibodies against respiratory chlamydiae.     

Vaccination against Chlamydial Genital Tract Infection after Immunization with Dendritic Cells Pulsed Ex Vivo with Nonviable Chlamydiae

Chlamydia trachomatis, an obligate intracellular bacterial pathogen of mucosal surfaces, is a major cause of preventable blindness and sexually transmitted diseases for which vaccines are badly needed. Despite considerable effort, antichlamydial vaccines have proven to be elusive using conventional immunization strategies. We report the use of murine bone marrow–derived dendritic cells (DC) pulsed ex vivo with killed chlamydiae as a novel approach to vaccination against chlamydial infection. Our results show that DC efficiently phagocytose chlamydiae, secrete IL-12 p40, and present chlamydial antigen(s) to infection sensitized CD4⁺ T cells. Mice immunized intravenously with chlamydial-pulsed DC produce protective immunity against chlamydial infection of the female genital tract equal to that obtained after infection with live organisms. Immunized mice shed ∼3 logs fewer infectious chlamydiae and are protected from genital tract inflammatory and obstructive disease. Protective immunity is correlated with a chlamydial-specific Th1-biased response that closely mimics the immune response produced after chlamydial infection. Thus, ex vivo antigen-pulsed DC represent a powerful tool for the study of protective immunity to chlamydial mucosal infection and for the identification of chlamydial protective antigens through reconstitution experiments. Moreover, these findings might impact the design of vaccine strategies against other medically important sexually transmitted diseases for which vaccines are sought but which have proven difficult to develop.     

The association between serological features of chronic Chlamydia pneumoniae infection and markers of systemic inflammation and nutrition in COPD patients

Introduction: Chlamydia pneumoniae is an obligatory human pathogen involved in lower and upper airway infections, including pneumonia, bronchitis. Asymptomatic C. pneumoniae carriage is also relatively common. The association of C. pneumoniae infections with the chronic obstructive pulmonary disease (COPD) course is unclear.

Objectives: The aim of the study was to investigate the association between chronic C. pneumoniae infection and clinical features of COPD, markers of inflammation and metabolic dysfunction.

Patients and methods: The study included 59 patients with stable COPD who had no, or had?≥2 acute exacerbations during last year. The level of IgA and IgG antibody against C. pneumoniae, IL-6, IL-8, resistin, insulin, adiponectin and acyl ghrelin was measured in serum by enzyme-linked immunosorbent assay (ELISA).

Results: No differences in clinical and functional data were observed between COPD patients without serological features of C. pneumoniae infection and chronic C. pneumoniae infection. The level of anti C. pneumoniae IgA significantly correlated with IL-8, IL-6, resistin concentration in group of frequent exacerbators. IgG level correlated negatively with acetyl ghrelin and body mass index (BMI) in patients without frequent exacerbations, in contrast to frequent COPD exacerbation group where significant correlations between IgG level and BMI was demonstrated. Serum IL-6 correlated positively with resistin and insulin and negatively with adiponectin in group of patients with serological features of chronic C. pneumoniae infection only.

Conclusions: Our study showed that chronic C. pneumoniae infection does not influence the clinical course of COPD in the both study groups. Chronic C. pneumoniae infections might be associated with a distinct COPD phenotype that affects metabolic dysfunction.     

Management of bacterial infections in children with asthma

Respiratory viruses are the single most common causes of asthma exacerbations in children. Rhinovirus-induced wheezing is a risk factor for chronic asthma, but its mechanism has remained unknown. Human bocavirus is a common finding in wheezing children, but its role as a respiratory pathogen is still unclear. Mycoplasma pneumoniae may, like viruses, induce wheezing and asthma exacerbation. Chlamydia pneumoniae and, in recent studies, Chlamydia trachomatis, may not only induce asthma exacerbations but may also be involved in the pathogenesis of chronic asthma. Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis are often involved in respiratory infections associated with wheezing, but there is no evidence for their active role in asthma pathogenesis or exacerbation. This review summarizes current knowledge on the association between respiratory infections and asthma in children, with a special focus on the role of antibiotics in incipient asthma, asthma exacerbation and chronic stable asthma.     

In Vitro Activity of Nemonoxacin,a Novel Nonfluorinated Quinolone Antibiotic,against Chlamydia trachomatis and Chlamydia pneumoniae

The in vitro activities of nemonoxacin, levofloxacin, azithromycin, and doxycycline were tested against 10 isolates each of Chlamydia trachomatis and Chlamydia pneumoniae. The MICs at which 90% of the isolates of both C. trachomatis and C. pneumoniae were inhibited (MIC₉₀s) were 0.06 μg/ml (range, 0.03 to 0.13 μg/ml). The minimal bactericidal concentrations at which 90% of the isolates were killed by nemonoxacin (MBC₉₀s) were 0.06 μg/ml for C. trachomatis (range, 0.03 to 0.125 μg/ml) and 0.25 for C. pneumoniae (range, 0.015 to 0.5 μg/ml).     

Analytic performance and contamination control methods of a ligase chain reaction DNA amplification assay for detection of Chlamydia trachomatis in urogenital specimens

A ligase chain reaction (LCR) DNA amplification assay that targeted the cryptic plasmid of Chlamydia trachomatis was developed to detect C. trachomatis urogenital tract infection. The objectives of this study were to determine the cutoff and analytic performance of the LCR assay and to characterize the effectiveness of its postdetection contamination control method. The assay's cutoff was determined after receiver-operator characteristic (ROC) analysis of 4660 clinical data points. The assay detected one infectious unit per reaction of each of the 15 C. trachomatis serovars and did not cross-react with 13 Chlamydia pneumoniae strains, 13 Chlamydia psittaci strains, and 87 other bacteria, fungi, parasites, or viruses. In addition, the assay did not detect 77 processed urine specimens collected from patients with urinary tract infections caused by yeast or bacteria other than C. trachomatis. The assay was sufficiently precise to detect consistently two infectious units of C. trachomatis per reaction. False-positive assay results attributable to contamination with amplified product were minimized by the use of standard procedures as well as by a postdetection chemical inactivation method that could reduce the amount of amplified LCR product by a factor of ⩾10⁷.     

Low prevalence of Chlamydia pneumoniae infections during the Mycoplasma pneumoniae epidemic season: Results of nationwide surveillance in Japan

ObjectiveChlamydia pneumoniae and Mycoplasma pneumoniae are both common causes of atypical pneumonia. We conducted an annual national survey of Japanese children to screen them for C. pneumoniae infections during the M. pneumoniae epidemic season.MethodsNasopharyngeal swab specimens were collected from children aged 0–15 years with suspected acute lower respiratory tract infection due to atypical pathogens, at 85 medical facilities in Japan from June 2008 to March 2018. Specimens were tested for infection using real-time polymerase chain reaction assays.ResultsOf 5002 specimens tested, 1822 (36.5%) were positive for M. pneumoniae alone, 42 (0.8%) were positive for C. pneumoniae alone, and 20 (0.4%) were positive for both organisms. In children with C. pneumoniae infection, the median C. pneumoniae DNA copy number was higher in those with single infections than in those with M. pneumoniae coinfection (p = 0.08); however it did not differ significantly according to whether the children had received antibiotics prior to sample collection (p = 0.34).ConclusionsThe prevalence of C. pneumoniae infection was substantially lower than that of M. pneumoniae infection during the study period. The change in prevalence of C. pneumoniae was not influenced by that of M. pneumoniae. Children with single C. pneumoniae infection are likely to have had C. pneumoniae infection, while those with coinfection are likely to have been C. pneumoniae carriers.     

In Vitro Activity of AZD0914, a Novel DNA Gyrase Inhibitor,against Chlamydia trachomatis and Chlamydia pneumoniae

The in vitro activities of AZD0914, levofloxacin, azithromycin, and doxycycline against 10 isolates each of Chlamydia trachomatis and Chlamydia pneumoniae were tested. For AZD0914, the MIC₉₀s for C. trachomatis and C. pneumoniae were 0.25 μg/ml (range, 0.06 to 0.5 μg/ml) and 1 μg/ml (range, 0.25 to 1 μg/ml), respectively, and the minimal bactericidal concentrations at which 90% of the isolates were killed (MBC₉₀s) were 0.5 μg/ml for C. trachomatis (range, 0.125 to 1 μg/ml) and 2 μg/ml for C. pneumoniae (range, 0.5 to 2 μg/ml).     

Clinical efficacy of clarithromycin against uterine cervical and pharyngeal <Emphasis Type="Italic">Chlamydia trachomatis</Emphasis> and the sensitivity of polymerase chain reaction to detect <Emphasis Type="Italic">C. trachomatis</Emphasis> at various time points after treatment

The detection and eradication of pharyngeal Chlamydia trachomatis in patients with chlamydial uterine cervicitis (commercial sex workers and others) were investigated. Pharyngeal C. trachomatis was detected in 75.0% of the commercial sex workers and in 21.9% of the other subjects. All the pharyngeal C. trachomatis-positive patients had a past history of orogenital contact. Chlamydial infection was treated with clarithromycin for 7 or 14 days. The presence of C. trachomatis was determined by polymerase chain reaction (PCR) on days 8, 15, and 22 after completion of the treatment. In the 7-day treatment group, the eradication rate of pharyngeal C. trachomatis was 53.3%, 56.7%, and 60.0% on days 8, 15, and 22, respectively, after completion of the treatment, while the eradication rate of cervical C. trachomatis was 83.3%, 96.7%, and 100% on days 8, 15, and 22, respectively. The eradication rate of pharyngeal C. trachomatis in the 7-day treatment was significantly lower than that of cervical C. trachomatis, while there was no significant difference in the 14-day treatment. The eradication rate of pharyngeal C. trachomatis in the 14-day treatment was significantly higher than that in the 7-day treatment. Since the DNA of dead organisms may be detected because of high PCR sensitivity, appropriate therapeutic judgment by PCR could be done around day 22 after completion of the treatment.     

Induction of the Chlamydia muridarum Stress/Persistence Response Increases Azithromycin Treatment Failure in a Murine Model of Infection

Viable but noninfectious (stressed/persistent) chlamydiae are more resistant to azithromycin (AZM) in culture than are organisms in the normal developmental cycle. Chlamydia muridarum-infected mice were exposed to amoxicillin to induce the organisms to enter the persistent/stressed state and subsequently treated with AZM. AZM treatment failure was observed in 22% of persistently infected mice, with an average of 321,667 inclusion-forming units (IFU) shed after AZM treatment. Productively infected mice had a 9% rate of AZM treatment failure and shed an average of 12,083 IFU. These data suggest that stressed chlamydiae are more resistant to frontline antichlamydial drugs in vivo.     

Investigation of pneumonia-causing pathogenic organisms in children and the usefulness of tebipenem pivoxil for their treatment

We investigated the usefulness of the novel oral carbapenem antibiotic tebipenem pivoxil (TBPM-PI) for treating bacterial pneumonia in children. Sputum and nasopharyngeal swabs were collected simultaneously, and causative organisms were identified by conventional bacterial culture together with exhaustive bacterial and viral identification by real-time polymerase chain reaction (PCR). The subjects were eight patients diagnosed with mild or moderate pneumonia at Sotobo Children’s Clinic Outpatient Department between October 2006 and June 2007. TBPM-PI was administered at the recommended clinical dose of 4 mg/kg b.i.d. to five patients and at a high dose of 6 mg/kg b.i.d. to three patients. Sputum was collected from all patients, and 11 strains were detected from washed sputum culture. Causative organisms were mainly Streptococcus pneumoniae (3 strains) and Haemophilus influenzae (6 strains), and nasopharyngeal swabs showed the same organisms as coughed-up sputum. Real-time PCR for individual viruses and Mycoplasma pneumoniae identified four cases of only bacterial infection, one case of M. pneumoniae coinfection, two cases of viral coinfection, and one case of both viral and M. pneumoniae coinfection. The clinical results indicated efficacy in all patients, and causative organisms were 100% eliminated. In the four patients with only bacterial infection, the average fever of 38.9°C at the start of treatment normalized the following day, showing excellent efficacy. No clinically problematic adverse events occurred, and compliance was good. We consider that these cases provide valuable insights into the identity of pathogenic organisms of pneumonia in children and the possible role of TBPM-PI in outpatient treatment.     

Underdiagnosis of Chlamydia trachomatis and Chlamydia psittaci revealed by introduction of respiratory multiplex PCR assay with Chlamydiaceae family primers

We describe unanticipated detection of respiratory infection with Chlamydia trachomatis and Chlamydia psittaci after introduction of respiratory multiplex polymerase chain reaction assay that includes Chlamydiaceae family primers. We detected cases of pediatric C. trachomatis and of adult C. psittaci infection in patients with previously unrecognized risk factors. Directed testing for C. trachomatis and C. psittaci based on clinical features and risk factors alone is likely to miss the majority of infected cases.     

A real-time quantitative polymerase chain reaction assay for the detection of Chlamydia in the mouse genital tract model

Chlamydia trachomatis is a human pathogen that infects genital tracts in women. Disease control may be achieved through development of an efficacious vaccine. A mouse genital tract model serves as a tool for evaluation of vaccine candidates. Currently, assessment of infection in mice is performed by enumeration of inclusion-forming units (IFUs) through microscopic counting of fluorescently stained bacteria. We have developed a highly sensitive real-time quantitative polymerase chain reaction (RT-qPCR) assay for enumeration of Chlamydia from mouse genital tracts to increase assay sensitivity, remove subjectivity, and improve sample throughput. The qPCR assay uses a 16S ribosomal gene sequence that is conserved across Chlamydia species and serovars, resulting in detection of multiple serovars of C. trachomatis, as well as Chlamydia muridarum and Chlamydia pneumoniae. The PCR assay provided results similar to IFU enumeration (94% agreement between the 2 assays) and is highly sensitive and specific with less inherent subjectivity than traditional enumeration methods.     

Microbiologic Efficacy of Azithromycin and Susceptibilities to Azithromycin of Isolates of Chlamydia pneumoniae from Adults and Children with Community-Acquired Pneumonia

Chlamydia pneumoniae was eradicated from the nasopharynges of 26 of 33 (78.8%) evaluable children and adults with community-acquired pneumonia who were treated with azithromycin. We tested 55 isolates of C. pneumoniae obtained from 46 of these patients against azithromycin. The MIC at which 90% of the isolates were inhibited and the minimal chlamydiacidal concentration at which 90% of strains tested were killed of azithromycin for these isolates were both 0.5 μg/ml. Seven patients remained culture positive after treatment. The MICs of azithromycin for isolates from two patients increased fourfold after therapy. However, all the patients with persistent infection improved clinically. Further studies of treatment of C. pneumoniae infection, utilizing culture, are needed both to assess efficacy and to monitor for the possible development of antibiotic resistance.     

Impact of capsaicin,an active component of chili pepper,on pathogenic chlamydial growth (Chlamydia trachomatis and Chlamydia pneumoniae) in immortal human epithelial HeLa cells

Chlamydia trachomatis is the leading cause of sexually transmitted infections worldwide. Capsaicin, a component of chili pepper, which can stimulate actin remodeling via capsaicin receptor TRPV1 (transient receptor potential vanilloid 1) and anti-inflammatory effects via PPARγ (peroxisome proliferator-activated receptor-γ) and LXRα (liver X receptor α), is a potential candidate to control chlamydial growth in host cells. We examined whether capsaicin could inhibit C. trachomatis growth in immortal human epithelial HeLa cells. Inclusion forming unit and quantitative PCR assays showed that capsaicin significantly inhibited bacterial growth in cells in a dose-dependent manner, even in the presence of cycloheximide, a eukaryotic protein synthesis inhibitor. Confocal microscopic and transmission electron microscopic observations revealed an obvious decrease in bacterial numbers to inclusions bodies formed in the cells. Although capsaicin can stimulate the apoptosis of cells, no increase in cleaved PARP (poly (ADP-ribose) polymerase), an apoptotic indicator, was observed at a working concentration. All of the drugs tested (capsazepine, a TRPV1 antagonist; 5CPPSS-50, an LXRα inhibitor; and T0070907, a PPARγ inhibitor) had no effect on chlamydial inhibition in the presence of capsaicin. In addition, we also confirmed that capsaicin inhibited Chlamydia pneumoniae growth, indicating a phenomena not specific to C. trachomatis. Thus, we conclude that capsaicin can block chlamydial growth without the requirement of host cell protein synthesis, but by another, yet to be defined, mechanism.     

Efficacy of an RNA detection test kit in the diagnosis of genital chlamydial infection

 A nucleic acid amplification method based on DNA detection, the current standard method for the diagnosis of genital infection by Chlamydia trachomatis, has been shown to potentially yield false-positive results after treatment in the clinical setting. RNA detection methods are more appropriate because viable organisms have multiple RNA copies that are surely detected by the method. In this study, we evaluated the efficacy of a new RNA detection test kit, the VIDAS PROBE CT test, in the diagnosis of genital chlamydial infection. For comparison, the standard DNA detection method, Amplicor STD-I, was also used in the study. First voided-urine samples and urethral smears from male patients with urethritis, and first voided-urine samples and cervical smears from female patients with cervicitis served as samples for the detection of C. trachomatis. Of the 60 first voided-urine samples from male patients, 21 were positive and 39 negative with the VIDAS PROBE CT test. Amplicor STD-I achieved exactly the same result. In female patients with cervicitis, the two test kits produced the same result, with 2 positive cervical smears and 38 negative. These results suggest that the VIDAS PROBE CT test is as efficient as Amplicor STD-I in the detection of C. trachomatis. While studies including a greater number of patients will be needed for revealing the unique advantages of the new RNA detection test kit, VIDAS PROBE CT, we concluded from the current study that the test may be clinically useful in the diagnosis of genital chlamydial infection. Received: September 24, 2002 / Accepted: October 29, 2002     

Comparison of complement fixation and microimmunofluorescence tests in respiratory infections caused by chlamydia and an evaluation of the serum amyloid a protein in chlamydial infections

We retrospectively analyzed serum samples for antibodies to chlamydia species using the micro-immunofluorescence (MIF) test from 72 patients previously tested by the clamydia complement fixation (CF) test, which is used to detect pulmonary psittacosis. Nineteen patients were positive for chlamydia with the CF test. Of these, 9 patients (47.4%) were also positive forC. psittaci infection, and 7 patients (36.8%) were positive forC. pneumoniae infection by the MIF test. Five (9.4%) and (11.3%) of the 53 CF-negative patients were positive for psittacosis andC. pneumoniae infection by the MIF test, respectively. Our results indicate that the serum of patients suspected of pulmonary psittacosis should be examined by the MIF test as well as CF in order to distinguish between infections caused byC. psittaci, C. pneumoniae andC. trachomatis, and to improve the sensitivity of serodiagnosis. Serum samples of all 7 patients positive by both CF and the MIF test forC. pneumoniae infection showed a diagnostic IgG antibody change toC. pneumoniae without IgM antibody changes, which suggested a primary infection. These results suggest that the diagnostic CF antibody response occasionally appears in reinfection, especially inC. pneumoniae infection. There was a positive correlation (r=0.858) between C-reactive protein and serum amyloid protein A, a sensitive acutephase serum reactant, over a wide range of concentrations in 19 patients with chlamydia respiratory infections.     

Chlamydia pneumoniae and its role in chronic obstructive pulmonary disease

The diverging of T-helper (Th) cells into predominantly Thl and Th2 subsets on the basis of their cytokine profiles has decisively improved our understanding of the pathogenesis of many chronic infectious diseases. Recent data suggest that the presence of interferon-γ and the subsequent suppression of interleukin-4 production leads to a Thl-type reponse that is required for the resolution of infections caused by intracellular pathogens. The ability of the macrophages to respond aggressively during early antigen contact seems to be one crucial factor in the development of an appropriate Th-cell response. Several host-related factors can affect macrophage function and the polarization of T-cell responses, ie the shift from a Thl response to a Th2 one, and thus dramatically deteriorate the resolution of infections caused by intracellular agents such as Chlamydia pneumoniae. Chronic C. pneumoniae infection has been associated with several common chronic diseases, quite recently with chronic obstructive pulmonary disease. Chronic C. pneumoniae infection may amplify smoking-associated inflammation in the bronchi and may be a contributory factor in the development of irreversible pathological changes.