Detection of Chlamydia trachomatis in first-voided urine sediments from male urethritis by polymerase chain reaction]

Chlamydia trachomatis was detected from first-voided urine sediments of 97 male patients with urethritis by polymerase chain reaction (PCR). Since urine and urinary sediments only treated with proteinase K inhibited DNA amplification by PCR, DNA was further purified by phenol extraction and concentrated. Two oligonucleotides based on sequences within the major outer membrane gene from C. trachomatis serovar L2 were used as primers. A DNA fragment of 242 bp specific for C. trachomatis was amplified by PCR and detected by agarose gel electrophoresis. The DNA fragment was amplified by PCR in all specimens of urine sediments from 50 patients with Chlamydiazyme-positive urethral swab. In 38 specimens of urine sediments from 47 patients with Chlamydiazyme-negative urethral swab, PCR was negative. The overall coincidence rate between the PCR for detecting C. trachomatis in first-voided urine sediments and Clamydiazyme in urethral swab was 90.7% (88/97). Detection of C. trachomatis from first-voided urine sediments by PCR was considered to be noninvasive and useful for the diagnosis of male urethritis due to C. trachomatis.     

Detection of Neisseria gonorrhoeae from male patients with urethritis by polymerase chain reaction]

A polymerase chain reaction (PCR) procedure was developed for detection of Neisseria gonorrhoeae. Two oligonucleotides based on sequences within a 16S ribosomal RNA gene from N. gonorrhoeae were used as extension primers for the PCR. A single DNA fragment of 206 bp was amplified, when N. gonorrhoeae DNA was template for the PCR. No amplified product was detected in Chlamydia trachomatis DNA, Ureaplasma urealyticum DNA or other bacterial DNAs. The DNA fragment of 206 bp was detected on agarose gel electrophoresis, when DNA of greater than or equal to 6.5 N. gonorrhoeae per PCR was used as template DNA for the PCR. The culture and the PCR were carried out for detection of N. gonorrhoeae in 67 urethral swabs obtained from male patients with urethritis. In 27 of 28 specimens in which N. gonorrhoeae was isolated and identified by the culture, 206 bp DNA fragment was amplified by the PCR, but in one specimen no DNA fragment was detected. In 2 of 39 culture-negative specimens, 206 pb DNA fragment was detected and in the remaining specimens, PCR was negative for N. gonorrhoeae. The overall detection coincidence rate between the culture and the PCR was 95.5% (64/67). Thus, the PCR procedure developed in this study was sensitive and specific for detection of N. gonorrhoeae and could be applied for diagnosis of gonococcal urethritis.     

Study on Chlamydia trachomatis antigen detection in first-voided urine sediments

C. trachomatis antigen in first-voided urine sediments was detected by a new EIA kit using a monoclonal antibody, IDEIA CHLAMYDIA (IDEIA, Novo Nordisk), in males with urethritis and females with cervicitis. The result was compared with that by Chlamydiazyme (Abbott). 1. C. trachomatis antigen detection in male urethritis (285 cases) by the IDEIA test: The antigen detection rate was 37.9% (108/285) in urethral smears, and 33.7% (96/285) in first-voided urine sediments of the patients. The positive co-incidence rate between urethral smears and first-voided urine sediments was 82.4% (98/108). Thus, the detection of the antigen seems feasible in first-voided urine sediments. 2. Comparison of C. trachomatis antigen detection by the IDEIA and Chlamydiazyme tests: In 78 male cases with urethritis undergoing both tests, the rates of antigen detection from urethral smears and first-voided urine sediments were studied. The detection rate from urethral smears was 41.0% (32/78) for IDEIA, and 37.2% (29/78) for Chlamydiazyme. In first-voided urine sediments, the rate was 35.9% (28/78) for IDEIA and 24.4% (19/78) for Chlamydiazyme. In both specimens, the detection sensitivity was higher for IDEIA. 3. C. trachomatis antigen detection in chlamydial cervicitis (28 cases) by the IDEIA test: The antigen detection rate was 46.4% (13/28) in urethral smears and 60.7% (17/28) in first-voided urine sediments. The detection rate in first voided urine sediments was higher. Thus, in patients suspected of having chlamydial cervicitis, it seems necessary not only to search the antigen in cervical smears but also to study the first-voided urine sediments.(ABSTRACT TRUNCATED AT 250 WORDS)     

Detection of Chlamydia trachomatis from urethral swab in male urethritis by polymerase chain reaction]

A polymerase chain reaction (PCR) with Chlamydia trachomatis-specific primers was applied for detection of C. trachomatis from urethral swab in male urethritis. The results were compared with those of culture method for detection of C. trachomatis. Of 18 clinical specimens tested in this study, inclusion bodies of C. trachomatis were detected in 11 specimens by the culture method. For PCR, sample DNA was prepared from transport medium in which urethral smear was suspended and two oligonucleotides based on sequences within the major outer membrane protein gene from C. trachomatis serovar L2 were used as extension primers. In 12 of the 18 specimens, 242bp DNA fragment was amplified by PCR and demonstrated to be the DNA fragment of C. trachomatis by Southern blot hybridization. No DNA of 242bp was amplified by PCR from five specimens in which any inclusion bodies of C. trachomatis were observed or from a specimen in which one inclusion body per cover slip was detected by culture method. C. trachomatis DNA of 242bp was amplified from all specimens in which 14 and more inclusion bodies per cover slip were detected by culture method. In two specimens concluded s negative by culture method, amplified C. trachomatis DNA were detected by PCR. Thus, the PCR would be a more simple and sensitive method for detection of C. trachomatis, compared with the culture method.     

Usefulness of real-time PCR in detecting Chlamydia trachomatis and Neisseria gonorrhoeae in endocervical swabs and first-voided urine specimens

We evaluated performance of Abbott RealTime CT/NG assay (real-time PCR, Abbott Japan) for detect Chlamydia trachomatis and Neisseria gonorrhoeae by real-time PCR in 88 female patients with cervicitis symptoms seen at gynecological clinics and 100 male patients with urethritis symptoms seen at urological or dermatology clinics in Kitakyushu, Japan. Endocervical swab and first-voided urine (FVU) specimens were then collected from women and FVU specimens from men. Detection rates of C. trachomatis and N. gonorrhoeae by real-time PCR in the 3 types of specimens were compared to those by ProbeTec ET assay (ProbeTec, BD Diagnostic System). The overall positive concordance between real-time PCR and ProbTec were 97.1% (66/68) for C. trachomatis and 100% (33/33) for N. gonorrhoeae, C. trachomatis detection yielded 3 discordant results in endocervical specimens and 1 discordant result in male FVU by real-time PCR and ProbTec. Three of 4 reexamined using Aptime Combo 2 Assay (Fuji Rebio Inc.) were positive for C. trachomatis. Endocervical swab and FVU specimen results for C. trachomatis were discordant in 3 cases in real-time PCR and 4 in ProbeTec. Subjects with 2 or more positive endocervical awab results in female or male FVU specimens were assumed to be "true positive" for C. trachomatis. The sensitivities of real-time PCR for detecting C. trachomatis was 94.4% in endocervical swabs, 77.8% in female FVU and 97.4% in the male FVU. The sensitivities for real-time PCR for detecting N. gonorrhoeae was 100% in all 3 specimentypes. Abbott RealTime CT/NG assay was useful for detecting C. trachomatis using endocervical swabs or male FVU specimens and for detecting N. gonorrhoeae using endocervical swabs and all FVU specimens.     

Simultaneous detection of Neisseria gonorrhoeae and Chlamydia trachomatis by PCR in genitourinary specimens from men and women attending an STD clinic

Neisseria gonorrhoeae and Chlamydia trachomatis are the two most common bacterial sexually transmitted infections that manifest primarily as urethritis in males and endocervicitis in females, though the infection may be asymptomatic especially in women. Since complications may occur in untreated symptomatic and asymptomatic infected individuals, early diagnosis and treatment of infected individuals is required to prevent severe sequelae and spread of these diseases. Recently molecular amplification assays like Polymerase Chain Reaction (PCR) and Ligase Chain Reaction (LCR) have been found to be highly sensitive and specific methods for detection of N. gonorrhoeae and C. trachonmatis not only in urethral and cervical specimens but also in urine. The objective of this study was to screen male and female Sexually Transmitted Disease (STD) clinic attenders, with and without symptoms suggestive of urethritis and cervicitis for presence of N. gonorrhoeae and C. trachomatis using a multiplex PCR based assay, to compare its performance with culture for N. gonorrhoeae and Direct Fluorescent Antibody (DFA) staining for C. trachomatis and also to compare the efficacy of PCR test performed on urine and genital swab specimens collected from this high risk group. Genital specimens and urine was collected from STD clinic attenders. N. gonorrhoeae and C. trachomatis was detected in genital specimens by culture and DFA respectively. Multiplex PCR was used to detect N. gonorrhoeae and C. trachomatis infection in both genital and urine specimens. Among men with urethritis, N. gonorrhoeae was detected in 70% by culture and 77% by PCR, while C. trachomatis as detected in 7.5% by DFA and 17.5% by PCR. Among females with endocervicitis, N. gonorrhoeae was detected in 7.7% by culture and 30.7% by PCR, while C. trachomatis was detected in 7.7% by DFA and in 15.4% by PCR. None of the asymptomatic males were positive for N. gonorrhoeae and C. trachomatis by conventional methods, while 43.9% were positive for N. gonorrhoeae and 7.5% for C. trachomatis by PCR. Fifty per cent of asymptomatic women were positive for C. trachomatis by PCR alone. We encountered PCR positive but culture/DFA negative results and also PCR negative but culture/DFA positive results. In view of this a single PCR test cannot be used for diagnosis and treatment of N. gonorrhoeae and C. trachomatis infection unless confirmed by a second test.     

Comparison of polymerase chain reaction and IDEIA Chlamydia in detection of Chlamydia trachomatis from first-voided urine of male urethritis patients]

We have reported a method for detection of Chlamydia trachomatis by polymerase chain reaction (PCR) with two oligonucleotides based on sequences within the major outer membrane protein gene from C. trachomatis serovar L2. In the previous report, in addition to treatment of the mixture of first-voided urine (FVU) sediment and 1 ml of urine with proteinase K. DNA purification by phenol extraction was necessary for preparation of template DNA for PCR. In this study, FVU sediment was suspended in 1 ml of Chlamydiazyme dilution buffer and a part of the suspension was treated with proteinase K for DNA extraction. The DNA extraction solution could be used as template for PCR without purification of DNA by phenol extraction. One hundred FVU specimens obtained from male urethritis patients were examined with the two methods (PCR and IDEIA) for detection of C. trachomatis. In 33 of 100 specimens, the DNA fragments of C. trachomatis was amplified by the PCR and in 32 of 100, the chlamydial antigen was detected by IDEIA. The positive and negative coincidence rate of the PCR to IDEIA were 93.8% (30.32) and 95.6% (65/68) respectively, resulting in a high overall coincidence rate at 95%. Thus, the improved method with PCR using FVU as a specimen is proved to be a useful, non-invasive diagnostic tool for diagnosis of chlamydial urethritis.     

Can a two-glass urine test or leucocyte esterase test of first-void urine improve syndromic management of male urethritis in southern Thailand?

The goal of this study was to determine whether a urine two-glass test or a leucocyte esterase (LE) test of first-void urine (FVU) improve the sensitivity or specificity of the World Health Organization (WHO) algorithm for the syndromic management of men with urethritis in southern Thailand. A secondary aim was to determine whether infection with Trichomonas vaginalis was sufficiently common to include treatment for it in a syndromic management protocol. One hundred and twenty-nine men with symptoms of urethritis seen at 2 STD clinics in Songkla Province, Thailand were enrolled. Symptoms and signs of each man were recorded and a urethral swab collected for microscopy and culture for Neisseria gonorrhoeae. A two-glass urine test and an LE test of an FVU specimen were performed. The FVU was tested by polymerase chain reaction (PCR) for N. gonorrhoeae, Chlamydia trachomatis and T. vaginalis. Dysuria was a symptom in 78% of men. A urethral discharge was a symptom in 68% but was evident on examination in 95% of the men. The prevalences of infection were 32.6% for N. gonorrhoeae, 23.3% for C. trachomatis, 1.6% for T. vaginalis and 51.9% for any infection. The sensitivities and specificities of urethral discharge on examination, two-glass test and LE test of FVU as indicators of infection with either or both of N. gonorrhoeae or C. trachomatis were 97% and 8%; 57% and 83%; and 59% and 78% respectively. Combinations of urethral discharge on examination and one of the other indicators were more specific but much less sensitive than the presence of discharge alone. Culture for N. gonorrhoeae was found to be only 43% sensitive compared with an expanded gold standard involving a PCR test. Our analysis demonstrates that neither the two-glass test nor the LE test of FVU were useful in improving on the WHO algorithm for management of men with urethritis. T. vaginalis was not common enough to include in a first-line syndromic management protocol for male urethritis. We recommend that, in southern Thailand, men with symptoms of urethritis in whom a urethral discharge is present on examination be offered immediate treatment for both N. gonorrhoeae and C. trachomatis as per the WHO algorithm.     

Comparison of polymerase chain reaction and enzyme immunoassay (Chlamydiazyme) in detection of Chlamydia trachomatis from male urethritis]

A method using polymerase chain reaction (PCR) was compared to an enzyme immunoassay (Chlamydiazyme) for detection of Chlamydia trachomatis by testing a reference strain and clinical specimens. Two oligonucleotides based on sequences within the major outer membrane protein gene from C. trachomatis serovar L2 were used as primers for the PCR. A DNA fragment of 242 bp specific for C. trachomatis was amplified by the PCR, when DNA of greater than or equal to 10(2) C. trachomatis was used as template for the PCR. A chlamydial antigen was detected by Chlamydiazyme, when greater than or equal to 2.6 x 10(3) C. trachomatis were applied for the enzyme immunoassay. The PCR method was 26 times more sensitive than Chlamydiazyme in detection of C. trachomatis. The PCR method and Chlamydiazyme were carried out to examine 74 urethral swabs obtained from male patients with urethritis for detection of C. trachomatis. In 45 of 74 specimens, the DNA fragment of C. trachomatis was amplified by the PCR, and in 41 of 74, the chlamydial antigen was detected by Chlamydiazyme. The detection rate of the PCR method (60.8%) was higher than that of Chlamydiazyme (55.4%). The positive coincidence rate of the PCR method to Chlamydiazyme was 100% (41/41) and negative coincidence rate was 87.9% (29/33). The overall coincidence rate between the two methods was high (94.6%). Thus, the PCR method was more sensitive than Chlamydiazyme for detection of C. trachomatis and specific for diagnosis of chlamydial urethritis.     

Observations on the microbiology of urethritis in black South African men

The occurrence of Neisseria gonorrhoeae, Chlamydia trachomatis and Mycoplasma genitalium was determined by molecular techniques in urine specimens from 182 black South African men who had symptoms and/or overt signs of urethritis. Eighty-six (47.3%) of these men were infected with N. gonorrhoeae. There were 185 men without overt evidence of urethritis, 16 (8.6%) of whom were also infected with N. gonorrhoeae. Of the 96 men who had non-gonococcal urethritis, 14 (14.6%) were infected with C. trachomatis, 16 (16.7%) with M. genitalium and only one with both microorganisms. In comparison, 15 (8.9%) of 169 men without overt urethritis and without N. gonorrhoeae were infected with C. trachomatis and 15 (8.9%) with M. genitalium, proportions that were about half the size of those in the group with overt urethritis.     

Molecular testing (strand displacement assay) for identification of urethral gonorrhoea in men: can it replace culture as the gold standard?

The objective was to evaluate the performance of Becton Dickinson's BD Probe Tec(TM) (BDPT) strand displacement amplification (SDA) test for the detection of Neisseria gonorrhoeae on urethral specimens from men with urethritis compared with conventional culture and to show that SDA improves the diagnostic yield of gonorrhoea infections (GC). Anonymized retrospective testing of stored urethral swab samples from men attending genitourinary services in East London was performed using SDA. The prevalence of GC culture positive infections in this sample was 20/152 (13%). The sensitivity, specificity, positive predictive value and negative predictive value for the BDPT-SDA system compared with culture were 100%, 95%, 77% and 100%, respectively. In this study population, the BDPT-SDA assay was a highly sensitive and specific test for the diagnosis of N. gonorrhoeae from urethral swabs in men. Therefore, SDA can be used to complement culture in the diagnosis of N. gonorrhoeae infection. No ethics committee approval was obtained as all samples were anonymized.     

Bacteriological and epidemiological study on Neisseria gonorrhoeae isolated from the pharyngeal specimens of male and female patients with gonorrhea

Neisseria gonorrhoeae were isolated from pharyngeal specimens of male and female patients and also from urethral and cervical discharges of male and female patients, respectively, suspected of having gonococcal infections in a urologic clinic in Kawasaki City. Microbiological and epidemiological studies were performed in 127 male and 41 female patients. The specimens were streaked onto the modified Thayer-Martin Selective Agar and the plates were incubated at 35 degrees C for 48 h under an atmosphere of 10% CO2. In 127 male patients, N. gonorrhoeae were detected in 117 (92.1%) of the urethral specimens. In these patients, N. gonorrhoeae were detected in pharyngeal specimens from 14 (11.0%) patients, but the pathogen was also detected in urethral specimens from these patients without exception. In 41 female patients. N. gonorrhoeae were detected in 20 (48.8%) of the 41 cervical discharges. When the pharyngeal specimens were tested, N. gonorrhoeae were detected in 14 (34.1%) of the 41 specimens. N. gonorrhoeae was simultaneously detected only in pharyngeal and cervical specimens from 11 of the 41 female patients and the pathogen was detected only in pharyngeal specimens from other 3 patients. There were no marked differences in antimicrobial susceptibilities between N. gonorrhoeae isolates from pharyngeal specimens and those from urethral or cervical discharges in all the patients tested. The PFGE patterns of 50 gonococcal isolates (25 pairs) from 25 patients (14 males and 11 females) in whom N. gonorrhoeae were simultaneously detected from pharyngeal and urethral or cervical specimens were analyzed. In 24 of 25 patients. N. gonorrhoeae isolated from the pharyngeal and urethral or cervical specimens in the same patients showed the same PFGE patterns. However, the 25 pairs showed the different PFGE patterns. From these results it is clarified that N. gonorrhoeae are detected in the pharyngeal specimens from considerable numbers of patients with gonorrhea, and there is a possibility that the pathogens prevailing among the patients differ in genetic sources.     

呼吸道感染腺病毒基因分型研究

目的人腺病毒是引起急性呼吸道感染的常见病原体,采用PCR法对人腺病毒进行分型鉴别可为人腺病毒感染的防治提供可靠依据。方法采集急性呼吸道感染患者咽拭子标本,提取DNA,采用PCR法对其进行基因分型分析。结果以500bp作为DNA分子质量标准对待测标本PCR产物进行1.2%琼脂糖凝胶电泳,46份标本中21份鉴定为腺病毒通用型(扩增片段300bp),1、2、3、4、7、21型分别有3、8、1、15、2、2份(扩增片段分别为396、393、316、305、305和507bp)。结论本院急性呼吸道感染患者感染的腺病毒以通用基因型、基因4型和基因2型为主。及时检测流行腺病毒型别及其发展规律,可为呼吸道感染类疾病的临床治疗以及防控提供指导。     

Diagnosis of gonorrhoea using a genetic transformation test at a venereal disease clinic in Thailand

A genetic transformation test (GTT), a technique in which gonococcal DNA is detected in clinical specimens, was used to search for Neisseria gonorrhoeae infections in 37 men and 159 women at the Venereal Disease clinic in Cholburi, Thailand. Swabs were collected in duplicate from cervical specimens from 159 women and from urethral specimens from 37 men. One of each specimen was cultured on Thayer-Martin media while the other was mailed to the United States at room temperature for the GTT which involved a delay of 10 to 14 days. With the urethral specimens N. gonorrhoeae was identified in 84% (31/37) of specimens and there was 100% concordance between the results of the GTT and culturing specimens directly on Thayer-Martin media. With cervical specimens N. gonorrhoeae was isolated from 26% (41/159) by the standard culture technique and 19% (13/159) by the GTT. Seventy-six percent of the culture positive specimens were positive with the GTT and two specimens from which N. gonorrhoeae were not isolated were positive in the GTT. The GTT technique enables physicians to send swab collected from patient with suspected gonorrhoea without any special transport media to a central laboratory for laboratory diagnosis of gonorrhoeal infections. This technique which uses reagents which are available in most bacteriology laboratories, should facilitate surveillance of gonorrhoea especially when specimens are collected in clinics where bacteriology laboratory facilities are not available.     

聚合酶链反应检测首次排空尿诊断男性泌尿生殖道解脲支原体感染的研究

目的 评估聚合酶链反应(PCR)检测首次排空尿(FVU)诊断男性泌尿生殖道解脲支原体(UU)感染的实用性。方法 对172例男性非淋菌性尿道炎患者的FVU进行PCR检测和培养，作为对比同时取尿道拭子做PCR及培养，以拭子培养为金标准判断各种方法的检测效果。结果 FVU-PCR法和FVU培养法的敏感性分别为100．0%和98．3％，特异性分别为95．6％和98．2％，阳性预测值(PPV)分别为92．1％和96．6％，阴性预测值(NPV)分别为100．0%和99．1％，总一致性分别为97．1％和98．3％。结论 PCR检测FvU法是具高度敏感性和特异性的非创伤性男性泌尿生殖道解脲支原体实验室诊断方法，FVU可代替拭子标本进行PCR以诊断男性泌尿生殖道UU感染。     

The infection rate of Chlamydia trachomatis in young adult men without urogenital symptoms--screening test using urine sediments

The detection of C. trachomatis antigen in first-voided urine sediments has recently been achieved by means of IDEIA CHLAMYDIA (IDEIA, Novo Nordisk), an EIA kit using monoclonal antibodies. Therefore, this kit was used as a screening test to examine the infection rate of C. trachomatis in young adult men without symptoms. The titers of serum IgA and IgG antibodies to C. trachomatis were also determined. 1. Antigen detection from first-voided urine sediments of young adult men without urogenital symptoms (141 cases): The detection rate by IDEIA was 5.0% (7/141). Three of the 7 cases which were positive for antigen in first-voided urine sediments were reaffirmed as having asymptomatic C. trachomatis urethritis, since they were also revealed to have C. trachomatis in the urethra. 2. The positive rates of serum antibodies: The titers of serum antibody were determined in 128 cases out of the 141 cases. The positive rates of IgA and IgG were 6.3% (8/128) and 35.9% (46/128) respectively. The positive rates of IgA and IgG antibodies were significantly higher in cases with positive antigen in first-voided urine sediments than in those which were negative. These results indicate that this kit is useful for antigen detection. 3. The screening test revealed asymptomatic C. trachomatis infections in 5% of young adult men, suggesting the extensive spread of the infection. The screening test using first-voided urine sediments will be useful in public health.     

Leucocyte esterase testing of first-voided urine and urethral and cervical smears to identify Mycoplasma genitalium-infected men and women

Leucocyte esterase (LE) in first-voided urine (FVU) and presence of leucocytes in urethral and cervical smears were evaluated to identify Mycoplasma genitalium infection in 416 men and 417 women attending Department of Genitourinary Medicine. M. genitalium was diagnosed in FVU specimens by realtime polymerase chain reaction. The prevalence of M. genitalium was 6.5% in women and 6.7% in men. In total, 88.5% (23/26) of M. genitalium-infected men were identified by a combination of urethral smear and the LE test. In women, the combination of urethral and/or cervical smears and/or a positive LE test identified 91.3% (21/23) of M. genitalium-infected patients. Organism load in FVU correlated significantly with presence of urethritis (> or =4 leucocytes per high-power field) in men. A combination of LE testing of urine and urethral and/or cervical smears can be used as screening tests to select patients for specific M. genitalium testing. By this strategy, about 10% of infected individuals will remain undetected.     

Evaluation of the new nucleic acid amplification system for direct detection of Chlamydia trachomatis and Neisseria gonorrhoeae in women

The performance of a real-time DNA amplification assay, BD ProbeTec ET System (BDPT, BD Diagnostic Systems), to detect Chlamydia trachomatis and Neisseria gonorrhoeae on endocervical and oropharyngeal samples was evaluated. After obtaining informed consent, 364 endocervical, 363 urine and 247 oropharyngeal specimens were collected from 307 cases. The overall agreement rate of the BDPT and Amplicor (AMP, Roche) assays for the detection of C. trachomatis and N. gonorrhoeae in endocervical samples was 99.2% (361/364) for C. trachomatis and 99.5% (362/364) for N. gonorrhoeae. Assay of oropharyngeal swabs by the BDPT yielded 21 C. trachomatis positives, and 19 of them were C. trachomatis negative by the DNA probe assay (Gen-Probe PACE). The AMP assay showed that 16/19 (84.2%) of the BDPT +/DNA probe - samples were positive. The BDPT also yielded 21 N. gonorrhoeae positives, 15 of which were negative with the DNA probe. Additional testing showed that all 15 BDPT +/DNA probe - samples were positive by the established nested PCR method. Our data suggest that the performance of the BDPT is comparable to that of AMP for detection of C. trachomatis and N. gonorrhoeae in endocervical swab samples and that it may be a useful method for detecting of C. trachomatis and N. gonorrhoeae in oropharyngeal samples clinically.     

Evaluation of a DNA-hybridization method for detection of African and Asian strains of Neisseria gonorrhoeae in men with urethritis

The 2.6-megadalton (MDa) cryptic plasmid and the 4.4-MDa beta-lactamase plasmid of Neisseria gonorrhoeae were radiolabeled with [32P] nucleotides and used as probes for direct detection of gonococci and beta-lactamase plasmids in urethral exudates from men with urethritis. The sensitivity and specificity of the DNA probes were compared with culture isolation of N. gonorrhoeae and biochemical tests of gonococcal isolates for beta-lactamase production. Of 216 urethral specimens, 180 were positive for N. gonorrhoeae by DNA probe and culture, 27 were negative by both tests, and 9 gave discordant results. Compared with culture and with the chromogenic cephalosporin assay, the sensitivity and the specificity of the DNA probe was 99% and 93% and that of the beta-lactamase probe assay was 91% and 96%, respectively. Electrophoresis of plasmids isolated from 90 gonococcal cultures showed that all contained the 2.6-MDa plasmid, 29 possessed a 3.2-MDa plasmid, 18 a 4.4-MDa beta-lactamase plasmid, and 11 had a 24.5-MDa conjugal plasmid. We conclude that the sensitivity of our DNA probes was comparable to that of culture for diagnosis of gonorrhea and to conventional tests for detection of beta-lactamase.     

DNA hybridization technique for the detection of Neisseria gonorrhoeae in men with urethritis

A technique to detect Neisseria gonorrhoeae directly in clinical specimens was developed using a modified DNA-hybridization method. It uses the gonococcal cryptic plasmid as the radiolabeled probe, can detect as few as 100 colony-forming units of N gonorrhoeae or as little as 0.1 pg of purified gonococcal plasmid DNA, and is highly specific. This technique for differentiating between gonococcal and nongonococcal urethritis was evaluated in men with symptomatic urethritis in Seattle. Sixty-three (89%) of 71 who had cultures positive for N gonorrhoeae were also positive by DNA hybridization, and all 42 whose cultures were negative were also negative by DNA hybridization. Five of six isolates from patients who were positive by culture but negative by hybridization lacked the gonococcal cryptic plasmid and belonged to a unique auxo-type which requires proline, citrulline, and uracil for growth.