Cross-reaction of clinical isolates of bacteria and yeasts with the chlamydiazyme test for chlamydial antigen, before and after use of a blocking reagent.

More than 1,000 clinical isolates of bacteria and yeasts were identified, subcultured, and tested at 10(8) colony-forming units per milliliter with the Chlamydiazyme assay to determine the variety of microorganisms which could cause false-positive results for Chlamydial antigen. False-positive Chlamydiazyme results were obtained from 27 of 465 (5.8%) gram-negative, aerobic, or facultatively anaerobic bacterial isolates (including 8 of 39 [20.5%] Neisseria gonorrhoeae and 8 of 149 [5.4%] Escherichia coli isolates) and also from 2 of 46 (4.3%) Bacteroides species isolates. No false-positive results were obtained either from 373 gram-positive bacterial isolates or from 153 yeasts. Results from 21 of 29 (72.4%) isolates that cross-reacted with Chlamydiazyme antibodies were repeatedly positive, but all 21 were confirmed as false-positive results using a blocking antibody. When an initial Chlamydiazyme result is positive, repeating the test, with and without use of the blocking antibody, appears to be effective in identifying those results (more likely from poorly collected endocervical specimens) that are falsely positive, even in the presence of high concentrations of cross-reacting bacteria. Microscopic determination of endocervical specimen adequacy also may help to minimize false-positive (and false-negative) Chlamydiazyme results.

Impact of endocervical specimen quality on apparent prevalence of Chlamydia trachomatis infections diagnosed using an enzyme-linked immunosorbent assay method.

Duplicate endocervical swabs were collected from 1824 patients for detection of Chlamydia trachomatis. Specimen pairs were combined into 400 microL of 0.9% saline solution. After vortexing, a 40-microL sample was smeared and stained with Papanicolaou's method for detection of endocervical and/or metaplastic (E-M) cells. The remaining specimen was tested for C trachomatis antigen with the use of an enzyme-linked immunosorbent assay (ELISA) procedure (Chlamydiazyme, Abbott Laboratories, North Chicago, Ill). Chlamydia trachomatis antigen was detected and confirmed (with the use of a blocking antibody [Abbott Laboratories]) in only 16 (1.7%) of 918 specimens that lacked detectable E-M cells, but it was detected significantly more frequently not only in 88 (13.3%) of 661 specimens that contained detectable E-M cells but also in 32 (13.1%) of 245 specimens that contained too many red blood cells to analyze microscopically. Of the initially positive ELISA results, none of 37 were falsely positive from specimens that contained 11 or more E-M cells, but significantly more (six [27.3%] of 22) were falsely positive from specimens that lacked detectable E-M cells. Variations in specimen quality had a significant impact on the incidence of both true-positive and false-positive ELISA results and could significantly influence understanding of the prevalence of chlamydial infections in women.

Efficacy of duplicate genital specimens and repeated testing for confirming positive results for chlamydiazyme detection of Chlamydia trachomatis antigen.

In an attempt to increase Chlamydiazyme (Abbott Laboratories) detection of Chlamydia trachomatis antigen and to establish the reproducibility of positive results, we carried out an investigation into the usefulness of testing duplicate specimens, of more aggressive endocervical specimen collection by using cytobrushes instead of swabs, and of the repeated testing of both specimens from patients with one or two positive results. Duplicate endocervical (female) and urethral (male) specimens, including one swab and one cytobrush specimen from 1,331 nonpregnant women, were collected from symptomatic and asymptomatic patients. Specimens were transported and tested for C. trachomatis antigen as specified by the manufacturer. Tests on all specimens from patients with positive results were repeated. Antigen was initially detected in one or both specimens from 210 (10.7%) of 1,968 patients, and repetition of the tests confirmed its presence in 198 (10.1%) of the patients, including all 183 patients in whom it was initially detected in both specimens. Initial results from at least 8 of the 12 patients with irreproducible antigen detection were most probably falsely positive. Results from 21 (10.6%) of the 198 patients for whom antigen detection was confirmed were repeatedly positive on only one specimen (9 [4.5%] on the second of the two specimens collected). Of 115 women from whom one swab and one cytobrush sample were taken and who had repeatedly positive results, antigen was detected in 7 (6.1%) only on the swab sample and in 4 (3.5%) only on the cytobrush sample. Use of the cytobrush does not appear justified with the Chlamydiazyme assay, and collection of duplicate specimens provided only a modest increase in detection of C. trachomatis. However, repeated testing of specimens when results from only one of two specimens are positive appears to be of clinical value.

Direct fluorescent-antibody confirmation of chlamydial antigen below the detection threshold of the chlamydiazyme enzyme-linked immunosorbent assay.

Of 4,000 endocervical specimens tested with the Chlamydiazyme enzyme-linked immunosorbent assay (Abbott Laboratories), 233 (5.8%) gave positive results (A492 above the cutoff), which were confirmed with a blocking reagent (Abbott). An additional 34 specimens (14.6%) with chlamydial antigen were detected and confirmed with the direct fluorescent-antibody test (Syva) from among those 66 Chlamydiazyme-negative specimens which had A492s that ranged from 0.030 to the cutoff and that could be blocked by > or = 50% with the blocking reagent.

Effect of endocervical specimen quality on detection of Chlamydia trachomatis and on the incidence of false-positive results with the Chlamydiazyme method.

Duplicate endocervical swabs were collected from 1,675 patients to assess the effects of variations in specimen quality on Chlamydiazyme (Abbott Laboratories) detection of Chlamydia trachomatis and the incidence of false-positive results. One swab (at random) from each patient was tested for C. trachomatis antigen by using the standard Chlamydiazyme procedure. A 200-microliter volume of 0.9% saline was added to the other swab from each patient. After vortexing, 20 microliters was smeared on a slide for Papanicolaou (Pap) staining and the remaining specimen was then tested with the Chlamydiazyme assay. The Chlamydiazyme result was positive for 170 (10.1%) and 165 (9.8%) of the stained and unstained duplicate specimens, respectively (no significant difference). Pap stains on smears from 1,536 (91.7%) of the patients were analyzed, and endocervical and/or metaplastic (E-M) cells were detected in 789 (51.4%) smears. Of these 1,536 stained and analyzed specimens, 150 (9.8%) were Chlamydiazyme positive but only 132 (88.0%) of the positive results were confirmed by repeating the test and using a monoclonal blocking antibody (Abbott). Confirmed Chlamydiazyme-positive results were obtained from only 34 (4.6%) of 747 specimens lacking E-M cells but from 98 (12.4%) of 789 specimens containing the cells (P less than 0.001). Of the 150 initially Chlamydiazyme-positive results obtained with Pap-stained, analyzed specimens, 12 (26.1%) of 46 were falsely positive from specimens lacking E-M cells but only 6 (5.8%) of 104 were falsely positive from specimens containing E-M cells (P less than 0.01). C. trachomatis antigen was detected significantly more frequently and false-positive results were significantly less common from specimens in which E-M cells were detected.

Comparison of cytobrushes with swabs for recovery of endocervical cells and for Chlamydiazyme detection of Chlamydia trachomatis.

Endocervical swab and cytobrush specimens from 1,301 symptomatic women were microscopically analyzed for adequacy and tested using Chlamydiazyme (CZ) (Abbott Laboratories). When the swab specimen was collected first, blocking antibody-confirmed CZ-positive results were obtained from 48 (8.0%) of 599 patients, 42 (87.5%) from swabs and 46 (95.8%) from cytobrushes (not significant). When the swab specimen was collected second, confirmed CZ-positive results were obtained from 46 (6.6%) of 702 patients, 44 (95.6%) from swabs and 41 (89.1%) from cytobrushes (not significant). Cytobrushes offered no significant advantage over swabs for CZ detection of Chlamydia trachomatis.

Diff-Quik stain as a simplified alternative to Papanicolaou stain for determination of quality of endocervical specimens submitted for PCR detection of Chlamydia trachomatis.

The simple, rapid, two-step Diff-Quik stain procedure (Baxter Diagnostics) was compared with the Papanicolaou stain for microscopic determination of endocervical specimen quality. Results from 230 (98.7%) of 233 specimens stained by both methods indicated agreement between the two staining methods for detection of the endocervical cells or erythrocytes indicating specimen adequacy. By using the Amplicor Chlamydia trachomatis Test (Roche Diagnostic Systems) to detect C.trachomatis and the Diff-Quik stain to assess specimen adequacy, PCR-positive results were obtained from 147 (9.1%) of 1,615 microscopically adequate specimens but from only 13 (2.2%) of the 583 inadequate specimens (P < 0.001).

Comparison of first-voided urine specimens with endocervical swab specimens for enzyme-linked immunosorbent assay detection of Chlamydia trachomatis in women.

OBJECTIVE: To compare first-voided urine specimens with paired endocervical swab specimens from women to determine the role of urine in complementing or replacing swab specimens for the detection of the chlamydial antigen. DESIGN: For 18 months, both endocervical swab specimens (the criterion standard) and urine specimens were tested for the chlamydial antigen, using an enzyme-linked immunosorbent assay (Chlamydiazyme, Abbott Laboratories, North Chicago, Ill). Positive results were confirmed using a blocking reagent (Abbott Laboratories) and/or a direct fluorescent antibody test (Micro-Trak, Syva, Palo Alto, Calif). A low level of chlamydial antigen (below the enzyme-linked immunosorbent assay threshold recommended by the manufacturer) was also looked for and, when found, was confirmed by the direct fluorescent antibody test. SETTING: Prenatal and family practice clinics in a 500-bed community hospital. PATIENTS: Specimens were collected from 489 random asymptomatic or symptomatic women. MAIN OUTCOME MEASURE: The detection of the chlamydial antigen from endocervical swab specimens was compared with the detection from first-voided urine specimens. RESULTS: Acceptable swab and urine specimens were obtained from 300 (61.3%) of the patients. The antigen of Chlamydia trachomatis was confirmed in 20 (6.7%) of the 300 women. Of the infected patients, the antigen was detected in both swab and urine specimens for nine patients (45%), only in the swab specimens for eight (40%), and only in the urine specimens for three (15%). Testing urine in addition to endocervical swab specimens allowed for the detection of 18% more chlamydial infections, whereas confirming the presence of the antigen below the enzyme-linked immunosorbent assay cutoff resulted in the detection of 54% more infections. CONCLUSIONS: Collecting multiple specimens and testing for low levels of chlamydial antigen may significantly improve the detection of chlamydial infections in women. First-voided urine may be an appropriate complementary specimen to endocervical swab specimens, but urine by itself does not allow for the adequate detection of the chlamydial antigen in women.

Efficacy of an enzyme-linked immunosorbent assay for detection of urinary tract immunoglobulins for diagnosis of urinary tract infections.

Results of the Uristat test (Shield Diagnostics Ltd.), a novel enzyme-linked immunosorbent assay (ELISA) for detection of urine antibodies to seven common bacterial pathogens, were compared with results of urine culture, urinalysis, and clinical history to determine the usefulness of Uristat in the diagnosis of urinary tract infections (UTIs). Midstream, catheterized, and indwelling catheter urine specimens sent to the laboratory for culture were included in the study. Quantitative cultures were performed on both 5% sheep blood agar and eosin-methylene blue agar. Uristat ELISAs were performed according to the manufacturer's instructions. By using a Bacillus subtilis bioassay technique, antibacterial activity was detected in the urine of 236 (22.2%) of 1,061 patients. Probable, possible, or asymptomatic UTIs were diagnosed for 258 (24.3%) of the 1,061 patients. Of those infections, 219 (84.9%) were caused by bacterial species whose antibodies were detectable by Uristat. Uristat's sensitivity and specificity were 76.7 and 56.0%, respectively. Uristat's predictive values of positive and negative results were 31.2 and 90.2%, respectively. Further development of the Uristat test is necessary before it can be of assistance in the diagnosis of UTIs.

Improved PCR detection of Chlamydia trachomatis by using an altered method of specimen transport and high-quality endocervical specimens.

Duplicate endocervical swabs were collected for detection of Chlamydia trachomatis by PCR (Roche Diagnostics). One swab was swirled in Specimen Transport Medium (Roche) for PCR testing and discarded. A saline aliquot from the other specimen, sent as a dry swab to the laboratory, was Papanicolaou stained to determine specimen adequacy, and the remainder was PCR tested. Significantly more (24%) PCR-positive results (118 versus 95; P < 0.001) were obtained with the dry specimens than with the swirled specimens when first tested. In addition, PCR-positive results were obtained with 107 (10.6%) of 1,007 microscopically adequate specimens but with only 3 (0.9%) of 341 inadequate specimens (P < 0.001).