Chlamydia trachomatis genital infections in tahiti

The rate ofChlamydia trachomatis infection was determined in three populations in Tahiti by means of a direct immunofluorescence test performed in specimens, tissue culture and detection of chlamydial antibody in serum specimens using a single-serotype indirect immunofluorescence test.Chlamydia trachomatis was recovered in 53 % of 53 bar girls, 24 % of 75 women attending a public maternity clinic for routine care, and 37 % of 71 men attending a sexually transmitted disease clinic with acute or subacute urethritis. The presence of chlamydial antibody in a high proportion of the groups studied confirmed the high frequency of chlamydial infections (62.3 %, 66.6 % and 83.1 % respectively).Neisseria gonorrhoeae infection was often associated with chlamydial infection in both bar girls and men with urethritis (11.4 % and 18.3 % respectively). With regard to clinical manifestations, 58.3 % (7/12) of bar girls and 23.2 % (10/43) women at the maternity clinic without clinical complaints were found to beChlamydia trachomatis-positive. The presence ofChlamydia trachomatis in these asymptomatic persons highlights their important role in spread of this organism in Tahiti. The findings indicate that routine testing forChlamydia trachomatis is warranted in patients attending the sexually transmitted disease and public maternity clinics in Tahiti.     

Evaluation of chlamydiazyme for the detection of genital infections caused by Chlamydia trachomatis.

Chlamydiazyme is a 4-h enzyme-linked immunoassay that detects an antigen of Chlamydia trachomatis directly in clinical specimens. This immunoassay was compared with cell culture for the diagnosis of chlamydial infections of the genital tract. The assay was evaluated at five clinics with a total of 1,277 cervical specimens of which 239 were culture positive. At three of these clinics where urethral samples were taken from males, 99 of 363 samples were culture positive. The sensitivity of the assay averaged 89.5% for detecting cervical infections and 78.8% for detecting male urethral infections. Specificity was 97.0% when samples from either males or females were tested. Some patients who were culture negative were infected with chlamydiae according to both Chlamydiazyme and a monoclonal antibody test that detected a chlamydial antigen distinct from the antigen detected by Chlamydiazyme. If the 15 females and 2 males who were positive by both immunoassays but culture negative were considered positive for chlamydial infection, the specificity of the assay was 98.4% in females and 97.7% in males. Chlamydiazyme is a simple and relatively rapid immunoassay that has sufficient sensitivity and specificity to supplant culture in the detection of genital chlamydial infections.     

Evaluation and comparison of tests to diagnose Chlamydia trachomatis genital infections

Infection with Chlamydia trachomatis results in intracytoplasmic inclusions and the generation of infectious elementary bodies (EBs). These can be detected by various procedures. Staining of epithelial cells with vital dyes was first used to detect inclusions, but is insensitive. Thus, Papanicolaou-stained cervical smears cannot be recommended. The advent of the ability to grow chlamydiae in cultured cells over 30 years ago had a major impact on chlamydial research and on detection. However, this procedure is probably <70% sensitive for cervical infection and less for urethral infection in men and is now practised infrequently following the advent of other, mostly less laborious and often equally, or more sensitive detection systems. Thus, staining a smear with a specific fluorescent monoclonal antibody to detect EBs is simple and the direct fluorescent antibody tests became a commercial proposition in the early to mid-1980s. Nevertheless, although highly sensitive and specific in competent hands, technical expertise is crucial and even the most experienced may be unable to read a large number of stained smears on slides quickly. In view of this, it is understandable that enzyme-linked immunosorbent assays (ELISAs) gained popularity from the mid-1980s onwards, for they are not very labour intensive and their reading is neither subjective nor tedious. Unfortunately, these aspects outweighed the fact that the ELISAs lack sensitivity, some being very insensitive. The situation has been rescued, however, by the advent in the early 1990s of methods that amplify chlamydial DNA, making it easily detectable by relatively simple procedures. The polymerase chain reaction is such a method and has high specificity and sensitivity, although commercial development has so far not met the high standard expected of it in terms of sensitivity. The ligase chain reaction does not invoke such criticism, and high values for both sensitivity and specificity may be expected, even on urine samples. This augers well for diagnosing an infected individual patient and for effective screening programmes. Antibody tests have no place in a screening programme and are of debatable value in diagnosis.      

A murine model for the study of Chlamydia trachomatis genital infections during pregnancy

Pregnant BALB/c mice were inoculated intravaginally on day 5 of gestation with the Chlamydia trachomatis mouse pneumonitis biovar. Animals that received 10(5), 10(6), or 10(7) inclusion-forming units (IFU) of C. trachomatis delivered prematurely on days 15 to 16 of gestation. A focal inflammatory infiltrate was observed in the wall of the uterus on the day 14 of gestation in animals inoculated with 10(5) IFU. In this group of mice, immunohistochemical analysis showed chlamydial inclusions in the endometrium and fetal membranes.     

Evaluation of Syva enzyme immunoassay for detection of Chlamydia trachomatis in genital specimens.

Detection of Chlamydia trachomatis infection was evaluated by culture and a new Syva enzyme immunoassay (EIA) in 1,012 patients at two Baltimore, Md., sexually transmitted disease clinics. The overall chlamydia prevalence determined by culture was 12%. For 506 fresh cervical and urethral specimens, the sensitivity of Syva EIA was 90% and its specificity was 94% compared with culture. Discordant Syva EIA results were further evaluated by staining the sediment in centrifuged culture transport media and Syva EIA transport tubes with a fluorescent monoclonal antibody to C. trachomatis to detect elementary bodies. Reanalysis of the data after use of this technique to resolve discordant results increased sensitivity and specificity to 92 and 96%, respectively. A subsample of 307 fresh cervical specimens was also tested in a three-way comparison using Abbott Chlamydiazyme, Syva EIA, and culture. In this sample, compared with culture, the sensitivity and specificity of Syva EIA were 87 and 95%, respectively, and for Chlamydiazyme they were 77 and 98%, respectively. Syva EIA is a 4-h, easy-to-perform enzyme-linked immunosorbent assay which has a high sensitivity with fresh genital specimens and offers an excellent alternative to culture.     

Genital Chlamydia trachomatis infections

Chlamydia trachomatis infections affect young, sexually active persons. Risk factors include multiple partners and failure to use condoms. The incidence of infection has increased in the past 10 years. Untreated C .  trachomatis infections are responsible for a large proportion of salpingitis, ectopic pregnancy, infertility and, to a lesser extent, epididymitis. Screening is a possible intervention to control the infection, which is often asymptomatic. The emergence of lymphogranuloma venereum proctitis in men who have sex with men, in Europe, and of a variant with a deletion in the cryptic plasmid, in Sweden, are new features of C .  trachomatis infections in the last years. A diagnosis is best made by using nucleic acid amplification tests, because they perform well and do not require invasive procedures for specimen collection. Single-dose therapy has been a significant development for treatment of an uncomplicated infection of the patient and his or her sexual partner.     

Cultural method for large-scale screening for Chlamydia trachomatis genital infection.

Idoxuridine-treated McCoy cells grown as monolayers in 96 well microplates provide a convenient method for the isolation of Chlamydia trachomatis. Staining of infected monolayers with periodic acid-Schiff reagent (PAS) allows easy recognition of C trachomatis inclusions without the need for dark-ground microscopy. By this method 384 clinical specimens can be examined concurrently. It is sufficiently sensitive to form the basis of a chlamydial culture service for patients attending Sexually Transmitted Diseases (STD) Clinics.     

Diagnosis of genital Chlamydia trachomatis infections in asymptomatic males by testing urine by PCR.

An enzyme-linked immunosorbent assay (EIA) (MikroTrak; Syva) was compared with PCR (Amplicor; Roche) for detection of Chlamydia trachomatis in first-void urine (FVU) from 184 men attending a skin and venereal disease clinic. The prevalence of C. trachomatis in the population studied was 18.5%. Discrepant results between Syva EIA and Roche PCR were retested by using major outer membrane protein primer-based PCR. After retesting, the sensitivity, the specificity, and the positive and negative predictive values for the Syva EIA were 85.3, 100, 100, and 77.5%, respectively, and those for the Roche PCR 100, 100, 100, and 100%, respectively. It was concluded that PCR provides a highly sensitive and specific noninvasive screening method for genital chlamydial infection in asymptomatic men.     

Evaluation of high-resolution typing methods for Chlamydia trachomatis in samples from heterosexual couples

We aimed to compare conventional ompA typing of Chlamydia trachomatis with multilocus sequence typing (MLST) and multilocus variable-number tandem-repeat (VNTR) analysis (MLVA). Previously used MLST and MLVA systems were compared to modified versions that used shorter target regions and nested PCR. Heterosexual couples were selected from among persons with urogenital C. trachomatis infections visiting the sexually transmitted infection outpatient clinic in Amsterdam, The Netherlands. We identified 30 couples with a total of 65 C. trachomatis-positive samples on which MLST and MLVA for eight target regions were performed. All regions were successfully sequenced in 52 samples, resulting in a complete profile for 18 couples and 12 individuals. Nine ompA genovars from D to K, with two variants of genovar G, were found. The numbers of sequence type and MLVA type profiles were 20 for MLST and 21 for MLVA, and a combination of MLST and MLVA yielded 28 profiles, with discriminatory indexes (D) ranging from 0.95 to 0.99. Partners in 17 couples shared identical profiles, while partners in 1 couple had completely different profiles. Three persons had infections at multiple anatomical locations, and within each of these three individuals, all profiles were identical. The discriminatory capacity of all MLST and MLVA methods is much higher than that of ompA genotyping (D = 0.78). No genotype variation was found within the samples of the same person or from heterosexual couples with a putative single transmission. This shows that the chlamydial genome in clinical specimens has an appropriate polymorphism to enable epidemiological cluster analysis using MLST and MLVA.     

Evaluation of enzyme immunoassay (Chlamydiazyme) for detecting Chlamydia trachomatis in genital tract specimens.

An enzyme immunoassay (Chlamydiazyme) for detecting Chlamydia trachomatis was evaluated on genital specimens from 96 men and 272 women attending a clinic for sexually transmitted diseases (STD clinic). Compared with a direct immunofluorescence test for chlamydial elementary bodies, the enzyme immunoassay had a sensitivity of 58% on specimens from men, a specificity of 90%, a positive predictive value of 93%, and a negative predictive value of 88%; the assay had a sensitivity of 67% on specimens from women, a specificity of 89%, a positive predictive value of 63% and a negative predictive value of 90%. Immunofluorescence provided the most stringent test for the performance of the enzyme immunoassay as values were improved a little when a cell culture procedure was used for comparison. Further evidence for the lack of sensitivity was the detection of elementary bodies, sometimes in large numbers, in the enzyme immunoassay buffer of 13 of 19 specimens that had given a negative enzyme immunoassay result and the finding in comparative titrations of four laboratory strains that the enzyme immunoassay was at least 100-fold less able to detect chlamydiae than either immunofluorescence or the cell culture procedure. Lack of specificity may be associated with the finding that the enzyme immunoassay antibody reacted with strains of Acinetobacter calcoaceticus, Escherichia coli, Gardnerella vaginalis, Neisseria gonorrhoeae and group B streptococci. The enzyme immunoassay was not considered to be sufficiently sensitive, specific, or reproducible for routine use.     

Comparison of multiple genital tract infections with Chlamydia trachomatis in different strains of female mice.

We have previously shown that female outbred CF-1 mice are susceptible to prolonged genital tract infection with the oculogenital serovars (D-K) of Chlamydia trachomatis, and that partial homotypic and heterotypic protection against reinfection is induced. To understand the possible role of inherent T-helper 1 (Th1)/Th2 polarity bias on both the course of infection and the level of acquired immunity induced by infection, 2 immunologically different and well-characterized inbred strains of mice, BALB/c and C57BL/6, were studied in this model. Groups of mice were inoculated intravaginally with C. trachomatis serovar D (Ct D) and monitored by culture to determine the duration of initial infection. Two months later, mice were reinfected, and monitored along with age- and condition-matched control groups. Plasma and vaginal secretions were collected for serologic analysis and specific delayed-type hypersensitivity was assessed by footpad swelling. Initial infection in C57BL/6 mice was comparable in duration to outbred CF-1 mice (median duration 42 versus 43.5 days), while BALB/c mice had a shorter median duration of initial infection (12 days). All strains had significantly shorter durations of infection following reinfection. BALB/c mice shed 4-10 times more inclusion-forming units (IFU) than both C57BL/6 and CF-1 mice on sample days during the first week of infection and all strains shed less IFU during reinfection. C57BL/6 and BALB/c mice had significantly lower anti-Ct D immunoglobulin G titers in both plasma and vaginal secretions than CF-1 mice following resolution of infection; the frequency of immunoglobulin A seropositive vaginal secretions was less in both inbred strains, being significantly less in the case of C57BL/6 mice. Qualitative analysis of the antigen specificity and isotype composition revealed differences among the mouse strains. All 3 strains had detectable levels of specific footpad swelling on day 14 of infection, whereas only BALB/c mice showed a significant response at 70 days post-infection. Significant differences between 2 strains of mice that differ in Th1/Th2 polarity bias were observed in: 1) the duration of infection; 2) the level of bacterial shedding during infection; and 3) the quantitative and qualitative cellular and humoral responses made in response to female genital tract infection with a human oculogenital isolate of C. trachomatis. In addition, a similar and significant level of partial acquired immunity to reinfection was observed in both strains, suggesting that inherent Th1/Th2 polarity bias present upon initial infection does not prevent the development of a protective immune response within the genital tract during infection with an oculogenital isolate of C. trachomatis.     

Evaluation of two rapid tests for the diagnosis ofChlamydia trachomatis genital infections

The performance of two new enzyme immunoassays (EIA) for the detection ofChlamydia trachomatis in a practice setting was compared. A consecutive series of 207 female patients seen at an inner-city sexually transmitted disease clinic were tested by cell culture, the Kodak SureCell (SC) and Abbott TestPack Chlamydia (TP) EIAs. In addition 210 male patients, selected by physicians on the basis of the fact that multiple urethral samples could be obtained, were tested by cell culture and SC. The prevalence of infection was 19 % in the females and 12.5 % in males. The sensitivity, specificity, positive predictive value and negative predictive value for the SC and TP were 88 %, 95 %, 81 %, 97 % and 59 %, 99 %, 95 %, 91 %, respectively, in the female population. The sensitivity of the SC was significantly greater than that of the TP (p 0.002). The performance values of the SC in men (in the same order) were 64 %, 96 %, 71 % and 95 %, respectively. The SC in male patients and the TP in female patients had low sensitivity. The sensitivity of the SC in female patients was significantly higher than that of the TP. However, the SC yielded more false positive results. To determine the utility of these tests in a practice setting further studies are required.     

Multicenter evaluation of the AntigEnz Chlamydia enzyme immunoassay for diagnosis of Chlamydia trachomatis genital infection.

To evaluate the AntigEnz Chlamydia enzyme immunoassay (EIA) (Baxter/Bartels, Issaquah, Wash.), we studied 320 men and 1,209 women attending clinics for sexually transmitted diseases in Baltimore, San Francisco, and Seattle. At examination, two separate swabs were obtained from each patient, one for chlamydial culture and one for EIA. Cervical samples were collected from women, and urethral samples were collected from men. The prevalence of chlamydial infection by culture was 9% in Baltimore (n = 532), 11% in Seattle (n = 500), and 9% in San Francisco (n = 497). To resolve specimens with discrepant culture and EIA results, the EIA transport buffer was centrifuged and the resuspended pellet was stained by direct immunofluorescence to determine whether elementary bodies were present. Overall sensitivity of the AntigEnz Chlamydia assay compared with culture was 87% in men and 86% in women, and overall specificities were 94 and 97%, respectively. Differences between centers were seen, with sensitivities ranging from 76% among men and 79% among women in Seattle to 100% among men and 95% among women in Baltimore. With a true positive considered to be either a culture-positive or an EIA- and direct immunofluorescence-positive specimen, the revised sensitivity was 91% in men and 88% in women. Overall revised specificity was 99% in both men and women. We conclude that in this high-prevalence population, the sensitivity and specificity of this assay compare favorably with those of other noncultural antigen detection tests for the diagnosis of chlamydial genital infection.     

Simplified culture procedure for large-scale screening for Chlamydia trachomatis infections.

A method that uses a 48-well tissue culture cluster tray system for the isolation of Chlamydia trachomatis is described. The cluster tray system was as sensitive (100%) as and more time efficient than the conventional cover slip method, thereby being considerably cost saving. With both culture methods, the prevalence rates of genital carriage of C. trachomatis in women attending clinics for legal abortion and for cervical dysplasia were 5% (31 of 641 patients) and 2% (3 of 148 patients), respectively.     

Entry of genital Chlamydia trachomatis into polarized human epithelial cells.

To study the initial invasion process(es) of genital chlamydiae, a model system consisting of hormonally maintained primary cultures of human endometrial gland epithelial cells (HEGEC), grown in a polarized orientation on collagen-coated filters, was utilized. After Chlamydia trachomatis inoculation of the apical surface of polarized HEGEC, chlamydiae were readily visualized, by transmission electron microscopy, in coated pits and coated vesicles. This was true for HEGEC maintained in physiologic concentrations of estrogen (proliferative phase) and of estrogen plus progesterone (secretory phase), despite the finding that association of chlamydiae with secretory-phase HEGEC is significantly reduced (P = 0.025; A.S. Maslow, C.H. Davis, J. Choong, and P.B. Wyrick, Am. J. Obstet. Gynecol. 159:1006-1014, 1988). In contrast, chlamydiae were rarely observed in the clathrin-associated structures if the HEGEC were cultured on plastic surfaces. The same pattern of coated pit versus noncoated pit entry was reproducible in HeLa cells. The quantity of coated pits associated with isolated membrane sheets derived from HeLa cells, grown on poly-L-lysine-coated cover slips in medium containing the female hormones, was not significantly different as monitored by radiolabeling studies and by laser scanning microscopy. These data suggest that culture conditions which mimic in vivo cellular organization may enhance entry into coated pits for some obligate intracellular pathogens.