Chlamydia trachomatis infection in asymptomatic Chilean men and with urethritis. Usefulness of first catch urine samples

Many new antibiotics with improved pharmacodynamic and pharmacokinetic properties have been developed in the last 30 years. This development, however, went down in the last five years. With modern techniques of sequencing of the complete bacterial genum in order to find new targest, with combinatory chemistry and with the high throughput screening about one new drug per year can be developed in the future. The activity of antibiotics is diminished by the increasing number of resistant strains and by the increase of infections with naturally resistant microorganisms. The rational use of antibiotics can slow down this trend and perhaps reverse it. To reach this aim it is necessary to increase research activities in the field of pharmacodynamics in order to allow a more rational dosing. Better technology documentation and statistics in microbiological diagnostics could improve calculated chemotherapy. Furthermore we need more information about the epidemiology of resistant bacteria, at least the knowledge about receptors, mechanism of action and mechanism of resistance should be known to circumvent these obstacles in antimicrobial chemotherapy.     

Detection of Chlamydia trachomatis infection by PCR in first voided urines in men

The goal of this study was to evaluate whether the new commercially available PCR-based assay Amplicor C. trachomatis (Roche Molecular Systems) could improve the diagnosis of chlamydial urogenital infections in men, compared with cell culture of C. trachomatis considered as the reference method. A total of 466 men attending the STD clinic were tested by the Amplicor test in urine and by cell culture in urethra. The prevalence of C. trachomatis was 13.7% (64/466) by cell culture and 14.4% (67/466) by the Amplicor test. After resolution of the discrepant results, the sensitivity of culture was 91.4% in male urethral specimens. The resolved sensitivity of the PCR assay was 92.7% in male urine and the specificity was 99.5%. We concluded that this rapid PCR-based assay showed an improvement in quality for diagnosing C. trachomatis infections in men.     

Minimal inhibitory effect of male urine on detection of Chlamydia trachomatis by Roche Amplicor PCR

Congenital afibrinogenemia is a rare autosomal recessive disorder characterized by the complete absence of detectable fibrinogen. Uncontrolled bleeding after birth from the umbilical cord is common, and spontaneous intracerebral bleeding and splenic rupture can occur throughout life. Patients respond well to fibrinogen replacement therapy, either prophylactically or on demand. Because the half-life of infused fibrinogen is essentially normal, the genetic defect is assumed to be at the level of synthesis, but no responsible locus has been identified. Preliminary studies using Southern blotting suggested that no gross structural changes of the fibrinogen genes were present in patients. We report the identification of causative mutations in a nonconsanguineous Swiss family with congenital afibrinogenemia. The four affected male individuals (two brothers and their two first cousins) have homozygous deletions of approximately 11 kb of the fibrinogen alpha-chain gene (FGA). Haplotype data suggest that these deletions occurred separately, on three distinct ancestral chromosomes, implying that the FGA region of the fibrinogen locus is susceptible to deletion by a common mechanism. Furthermore, our results demonstrate that humans, like mice, may be born without the capacity to synthesize functional fibrinogen.     

Detection of Chlamydia trachomatis in first-void urine from men and women as an alternative to swabs

BACKGROUND: Ideally, an effective preventive strategy for the control of Chlamydia trachomatis infection should take into account the following attributes: rapid and simple specimen collection, low cost and noninvasive test processing. Therefore, we compared the performance profile of urine-based detection of C. trachomatis antigen in first-void urine with that of testing urethral and endocervical samples in men and women. MATERIAL AND METHODS: Urethral and endocervical samples and first-void urine from 285 men and 192 women attending the Sexually Transmitted Diseases Outpatient Clinic at the Medical University in Plovdiv, Bulgaria were tested using direct immunofluorescence assay (DFA) (MicroTrak, Syva, Palo Alto, CA, USA). RESULTS: Seventy (25%) of all men tested were positive for C. trachomatis antigen in either urethral or urine samples. 65 men (93%) had both a positive urethral and urine sample, three men (4%) had only a positive urethral sample and two (3%) had only a positive urine sample. Thirty-five women (18%) had C. trachomatis infection. Twenty-six women (74%) had both a positive endocervical and urethral sample, 6 (17%) had only a positive endocervical sample and 3 (8%) had only a positive urethral sample. All women with positive urethral samples tested positive on their urine samples. Two of the women with a negative urethral sample and a positive endocervical sample had a positive urine sample. CONCLUSIONS: These results show that using direct immunofluorescence assay on first-void urine samples is a reliable noninvasive method which can replace urethral swabs in the diagnosis of C. trachomatis infection in symptomatic men. Urine-based strategies are also an acceptable alternative for the diagnosis of C. trachomatis infection in symptomatic women when it is not possible to obtain an urogenital sample.     

Is urine leukocyte esterase test a useful screening method to predict Chlamydia trachomatis infection in women?

We evaluated the use of the leukocyte esterase test (LET) on first-catch urine specimens from women as a screening test to predict infection with Chlamydia trachomatis. For diagnosis, we used Abbott's ligase chain reaction (LCR) on urine specimens and isolation by tissue culture (TC) on cervical brushes. Of 4,053 women attending sexually transmitted disease and family planning clinics, 4.3% (n = 174) were positive by TC and 5.9% (n = 239) were positive by LCR. When LET was compared to TC, the sensitivity, specificity, positive predictive value, and negative predictive value were 54.0, 67.0, 6.8, and 97.0%, respectively. The corresponding performance of LET versus LCR was 53.1, 67.3, 10.1, and 95.8%. Almost half of the laboratory-confirmed chlamydial infections were negative by LET. The low specificity probably reflects multiple causes of pyuria in women and results in a low positive predictive value. LET is neither sensitive nor specific as a predictor of chlamydial infection and cannot be recommended for use as a screening test for C. trachomatis with first-catch urine samples from females from low- or moderate-prevalence populations.     

Trend in Chlamydia trachomatis infection among pregnant women in the past ten years in Japan: significance of Chlamydia trachomatis seroprevalence

BACKGROUND AND OBJECTIVES: Chlamydia trachomatis infection is believed to be the most common bacterial sexually transmitted disease (STD) in industrialized countries. The objective of the current study was to assess the recent trend in the prevalence of C. trachomatis in Japan. GOAL OF THIS STUDY: To determine the trend in the seroprevalence for C. trachomatis among pregnant women in Nagasaki, Japan, during the past 10 years. STUDY DESIGN: The seroprevalence for C. trachomatis of 9,652 pregnant women of various ages screened in 1996 and 1997 was compared with those of 275 and 297 stocked samples from 1987 and 1992, respectively. Serum antibodies to C. trachomatis were detected by the enzyme immunoassay. Prospective samples of 33 seropositive cases were also analyzed to determine kinetics of the serum antibody titer. RESULTS: The seroprevalence has decreased in all age groups during the last 10 years. More than 70% of seropositive cases converted to be seronegative within 10 years. CONCLUSION: The prevalence of C. trachomatis has been decreasing among Japanese pregnant women.     

Performance of a commercial polymerase chain reaction test for endocervical Chlamydia trachomatis infection in a university hospital population

The rate of survival and causes of mortality in a cohort of 2103 psychiatric patients registered on a psychiatric case register and followed up for 7 years are compared with those of a general population sample (n = 2382) randomly extracted from the municipal census in Valencia (Spain). Using multivariate analysis by Cox regression, patients suffering organic psychoses and those diagnosed with drug abuse or dependency exhibited a greater risk of death than the general population for the total causes of death; no interaction was found between sociodemographic variables and psychiatric pathology. In terms of the causes of death, and controlling for the effect of age and sex, organic psychoses involved a greater risk of death due to cardiovascular and respiratory causes, and a greater risk of non-natural deaths than the general population. Schizophrenia and related conditions, the abuse of alcohol/ other drugs, and neurosis/personality disorders all presented a higher risk of death from liver disease. The major affective disorders involved a greater risk of death due to suicide or accidents. The study concludes with a discussion of the possible explanations of these results.     

Genotyping of Chlamydia trachomatis in urine specimens will facilitate large epidemiological studies

The serovars of Chlamydia trachomatis-positive urine specimens (n = 81; as detected by PCR and ligase chain reaction) were successfully analyzed in 94% of cases by omp1 PCR-based RFLP analysis. The use of urine specimens and this simple and sensitive typing method will greatly facilitate epidemiological studies of C. trachomatis serovar distribution in asymptomatic C. trachomatis infections in both females and males.     

Inhibition of PCR in genital and urine specimens submitted for Chlamydia trachomatis testing

We determined the frequency of PCR inhibition in genital and urine specimens submitted for Chlamydia trachomatis testing using the internal control DNA provided with the COBAS AMPLICOR C. trachomatis test and assessed methods to remove it. Inhibition occurred in 65 of 906 (7%) cervical swabs, 23 of 51 (45%) urethral swabs, and 2 of 175 (1.1%) urine samples. Overall, inhibition was eliminated in processed specimens after storage at 4 degrees C in 77 of 90 specimens (86%), freezing at -70 degrees C in 59 of 82 specimens (72%), storage at 4 degrees C followed by either 1:100 dilution in 37 of 43 specimens (86%) or 1:10 dilution in 42 of 47 specimens (89%), and phenol-chloroform extraction in 79 of 80 specimens (99%). No positive specimens were missed due to inhibition. We conclude that PCR inhibition is rare with urine specimens and infrequent with endocervical swabs but occurs frequently with urethral swabs. The frequency of PCR inhibition may be significantly reduced by methods which can be easily incorporated into the processing of specimens.     

Detection of Chlamydia trachomatis infections in women by Amplicor PCR: comparison of diagnostic performance with urine and cervical specimens

We used the Roche Amplicor PCR assay to compare urine and cervical swabs as sample material in the detection of Chlamydia trachomatis causing genital infections. The diagnostic performance of Amplicor PCR was compared with that of cell culture and the Gen-Probe PACE 2 assay with cervical specimens. If discrepant from other results, the specimens negative by PCR were diluted and reanalyzed to reveal PCR inhibitors. Of 666 patients, 39 (5.9%) were confirmed to have chlamydial infection. The respective sensitivity and specificity of Amplicor PCR were as follows: urine specimens, 82.0 and 99.7%; cervical specimens, 82.0 and 99.8%. Those for cell culture with cervical specimens were 84.6 and 100%. For the Gen-Probe PACE 2 assay, the sensitivity and specificity with cervical specimens were 79.5 and 100%, respectively. Without the effect of PCR inhibitors, the sensitivity of PCR with urine would have been 97.4%. Provided that the problems currently caused by inhibitors will be solved, the Amplicor PCR assay with urine specimens offers a tempting alternative for the diagnosis of C. trachomatis infection in women.     

Antibodies to two immunotypes of Chlamydia trachomatis in individuals with trachoma

In Tunisia (North Africa), trachoma remains a common eye disease. Most cases are caused by immunotype A of Chlamydia trachomatis. In a small proportion of cases (less than 10%), type-specific antibodies to both immunotype A and immunotype B exist in the serum of patients. These types do not crossreact, and occasionally sequential acquisition of antibodies has been demonstrated. Thus, individuals in one endemic area of trachoma may be infected with two immunotypes of C. trachomatis. The epidemiological and immunological implications of this new finding are briefly discussed.     

Evaluation of 2-SP transport medium for detection of Chlamydia trachomatis and Neisseria gonorrhoeae by two automated amplification systems and culture for chlamydia

AIMS: To assess the performance of 2-sucrose-phosphate based transport medium (2-SP) for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae by an automated commercial polymerase chain reaction (PCR) and ligase chain reaction (LCR) compared to centrifugation culture on McCoy cells for C trachomatis. Second, to compare both amplification systems for initial diagnostic testing of a low prevalence population for sexually transmitted diseases. METHODS: Four hundred and eighty one consecutive urogenital and conjunctival specimens were examined. All tests were performed on the same specimen collected with a dacron swab and transported in 2-SP medium. Samples that were positive by culture or by both PCR and LCR were considered to be true positives. RESULTS: The prevalences of C trachomatis and of N gonorrhoeae were 2.7% and 0.4%, respectively. PCR had a resolved sensitivity and specificity of 100% and 99.8%, respectively, for C trachomatis, and 100% and 98.9%, respectively, for N gonorrhoeae. LCR was 100% sensitive and specific for both pathogens. The resolved sensitivity of the shell vial assay was 85%. No culture positive sample would have been missed by PCR or LCR. The inhibition rate for PCR was 4.8%. CONCLUSIONS: 2-SP medium proved to be suitable for both PCR and LCR. It is not limited to any one test manufacturer and allows the performance of amplification techniques and viral and chlamydia culture from the same specimen. The LCR was more reliable than PCR on initial testing. However, hands on time is longer, and no amplification control is available for LCR.     

Comparison of serological tests for the detection of antibodies against Chlamydia trachomatis and Chlamydia pneumoniae in rheumatological patients

In cases of reactive arthritis, a suspected Chlamydia trachomatis infection is often detected by serological methods. However, mostly tests with genus-specific antigens are used, neglecting the fact that antibodies against Chlamydia pneumoniae are highly prevalent in the adult population. Therefore we tested sera of 129 patients with various rheumatological disorders and of 18 healthy persons in parallel with a genus-specific test (IPAZYME) and with the species-specific microimmunofluorescence test for C. trachomatis and C. pneumoniae antibodies. The data showed that 55% of the 64 IPA-positive results were caused by antibodies (IgG) against Chlamydia pneumoniae, only 6% by anti-Chlamydia trachomatis IgG and 20% by both specificities. For IgA antibodies, the percentages were 44%, 12.5% and 12.5% respectively. In 12 IPA-positive cases, the MIF showed no reaction. 58% of all 147 sera tested with MIF had IgG antibodies against C. pneumoniae, 5% had anti-C. trachomatis IgG and 8% IgG against both species. The percentages for IgA were 29%, 2% and 2%, respectively. IgM positivity in MIF disappeared after absorption with rheumatoid factor absorbent. No significant differences were found between the various groups of patients. The data suggest that due to the high prevalence of anti-C. pneumoniae antibody, genus-species tests cannot be used as screening tests for the serological diagnosis of C. trachomatis infections.     

Improvement of cervical Chlamydia detection in asymptomatic family planning attendees by using Amplicor Chlamydia trachomatis assay

We evaluated the Amplicor C. trachomatis PCR assay (Roche Molecular Systems, N.J.) for the diagnosis of cervical infection in asymptomatic women attending a family planning clinic, aged between 18 and 25 years. Culture onto McCoy cells with fluorescent monoclonal staining was the reference system. Cervical specimens from 485 women were tested. The prevalence of C. trachomatis was 10.5% by culture and 11% by Amplicor. No specimen was positive by culture and negative by PCR. Three PCR-positive, culture-negative specimens were positive by MOMP-PCR and a second plasmid-based PCR. The resolved sensitivity of PCR and culture were 100% and 94.5%, respectively. Specificities for both were 100%, positive and negative predictive values for culture were 100% and 99.3%. Total test efficiency was 99.4%. The Amplicor C. trachomatis assay gave very clear results, quite above or below the cut-off value, and showed high sensitivity and specificity, improved ease of handling and represented a good alternative to culture for large scale diagnosis of asymptomatic C. trachomatis infection.     

Comparison of performances of two commercially available tests, a PCR assay and a ligase chain reaction test, in detection of urogenital Chlamydia trachomatis infection

The diagnostic performance of a PCR test (Roche Cobas Amplicor CT/NG Test) and that of a ligase chain reaction (LCR) test (Abbott LCx Chlamydia trachomatis assay) were compared by using endocervical and urethral swab specimen culture as a reference test. First-void urine (FVU) and endocervical and urethral swab specimens were collected from 1,015 unselected patients attending a sexually transmitted disease clinic and a clinic for adolescents in Helsinki, Finland. Chlamydia trachomatis was cultured from samples from the endocervix or urethra. PCR was performed with fresh and frozen urine and the culture transport medium. LCR was performed with fresh and frozen urine and LCx swab transport medium. Diagnostic consistency and diagnostic accuracy were statistically tested. The test results were identical for 984 patients (97%). Discrepant results were observed for 31 patients. Overall, LCR and PCR showed excellent kappa coefficients of consistency for both swab and FVU specimens (0.93 and 0.95, respectively). Sixty-one patients (6%) were culture positive. Testing of FVU by LCR or PCR increased the overall positivity rates to 7.0 and 7.7%, respectively. While PCR of FVU detected the greatest number of C. trachomatis infections (sensitivity, 96.1%), for some PCR-positive FVU specimens the results could not be confirmed (specificity, 99.6%). PCR and LCR were more sensitive than culture (sensitivities, 92 and 93% versus 79% for culture) in the diagnosis of genital C. trachomatis infection. In conclusion, both tests can be recommended for use in the clinical laboratory and for the screening of asymptomatic C. trachomatis infections.     

Chlamydia trachomatis utilizes the host cell microtubule network during early events of infection

The host cell cytoskeleton is known to play a vital role in the life cycles of several pathogenic intracellular microorganisms by providing the basis for a successful invasion and by promoting movement of the pathogen once inside the host cell cytoplasm. McCoy cells infected with Chlamydia trachomatis serovars E or L2 revealed, by indirect immunofluorescence microscopy, collocation of microtubules and Chlamydia-containing vesicles during the process of migration from the host cell surface to a perinuclear location. The vast majority of microtubule-associated Chlamydia vesicles also collocated with tyrosine-phosphorylated McCoy cell proteins. After migration, the Chlamydia-containing vesicles were positioned exactly at the centre of the microtubule network, indicating a microtubule-dependent mode of chlamydial redistribution. Inhibition of host cell dynein, a microtubule-dependent motor protein known to be involved in directed vesicle transport along microtubules, was observed to have a pronounced effect on C. trachomatis infectivity. Furthermore, dynein was found to collocate with perinuclear aggregates of C. trachomatis E and L2 but not C. pneumoniae VR-1310, indicating a marked difference in the cytoskeletal requirements for C. trachomatis and C. pneumoniae during early infection events. In support of this view, C. pneumoniae VR-1310 was shown to induce much less tyrosine phosphorylation of HeLa cell proteins during uptake than that seen for C. trachomatis.     

Selective screening for Chlamydia trachomatis infection in a primary care population of women

The potency of food chemicals to induce cell aging was evaluated in human diploid fibroblast cells HAIN-55 having a finite replicative potential by using in vitro aging markers, i.e., decreases of maximum proliferative potential (lifespan) of cells, saturation density in monolayer culture (SD), plating efficiency (PE) and mitotic index (MI), and an increase of cells with polyploid karyotypes. By treatment twice with low concentration of genotoxic chemicals aflatoxin B1, allylisothiocyanate or trans-cinnamaldehyde (severe clastogenic flavoring agent; Kasamaki et al., 1982), lifespan (expressed by the number of cumulative cell population doubling (CPD)) of the treated cells was reduced by 8-12 CPDs accompanied by change of the other aging markers. By successive treatment (29 or 25 times) with non-genotoxic chemical aspartame (N-L-aspartyl-L-phenylalanine) or L-canavanine (structural analogue of L-arginine), lifespan of the treated cells was also slightly shortened (by 2-6 CPDs) compared with the untreated control cells. In the process of cell aging, Mitochondrial activity (MTT activity) decreased almost in parallel with the decrease of SD and MI. On the basis of these results, a variety of genotoxic and non-genotoxic chemicals were examined by using MTT activity as the aging marker for their effects on the aging of HAIN-55 cells and bovine artery endothelial cells which also had a finite replicative potential. The results showed that seven genotoxic and nine non-genotoxic chemicals promoted cell aging.