BD ProbeTec ET Research Articles

Incidence of Chlamydia trachomatis infections in the provinces of Novara and VCO: one year of observations

Chlamydia trachomatis infections represent the most common sexually transmitted diseases.The lack of growth of these microorganisms on common media and the roundabout research by immunofluorescence, has meant that, until now, these special-lived intracellular microorganisms have played a role probably underestimated in cases of infections affecting the sexual organs.The introduction of molecular biology has made possible their research in a much more simple, reliable and standardized methods. The study was carried out on endocervical samples, male urethral swabs and urine samples of men and women, determining the C. trachomatis DNA amplified using the BD ProbeTec ET system. This system uses technology SDA (Strand Displacement Amplification - Elongationcrowding of the blanks). The study was conducted from June 2008 to june 2009. Symptomatic and asyntptomatic patients residents in the provinces of Novara and the Verbano-Cusio-Ossola were considered. In this period 784 determinations were made in molecular biology. Chlamydia trachomatis was found in 26 cases equal to a percentage of 3.32%. While considering these preliminary data, however, we can estimate the rate of detection of C. trachomatis as a good starting point, because we have the impression that improved procedures particularly relate to the techniques for sampling and for storage and transport of the sample, can undoubtedly lead to much higher percentages.

The Swedish new variant of Chlamydia trachomatis (nvCT) remains undetected by many European laboratories as revealed in the recent PCR/NAT ring trial organised by INSTAND e.V., Germany

The May 2009 round of INSTAND's ring trial "Chlamydia trachomatis detection PCR/NAT" included a sample with high amount of the Swedish new variant of C. trachomatis (nvCT). A spectrum of at least 12 different commercial diagnostic nucleic acid amplification tests (NAATs) and many different in house NAATs were applied by the 128 participating laboratories which reported 152 results. Approximately 80% of the results correctly reported the presence of C. trachomatis in the nvCT specimen. The nvCT sample was mainly missed, as expected, by participants using the Roche COBAS Amplicor CT/NG (15.5% of reported results) but also by several participants using in house NAATs. The trend towards using nvCT-detecting NAATs is obvious and in addition to the new dual-target NAATs from Roche and Abbott, and BD ProbeTec ET, also a number of new CE mark-certified commercial tests from smaller diagnostic companies as well as many different in house NAATs were used. Laboratories using commercial or in house NAATs that do not detect the nvCT are encouraged to carefully monitor their C. trachomatis incidence, participate in appropriate external quality assurance and controls schemes, and consider altering their testing system. The reliable detection of low amounts of the wildtype C. trachomatis strain in other samples of the ring trial set indicates a good diagnostic performance of all applied commercial NAATs while also detecting the nvCT strain.

High-throughput multistrain polymerase chain reaction quantification of Chlamydia trachomatis from clinical and preclinical urogenital specimens

Chlamydia trachomatis (CT) is the most prevalent sexually transmitted bacterial pathogen worldwide and causes severe reproductive tract infections. Currently, nucleic acid amplification tests (NAATs) are the gold standard for clinical diagnosis, but most NAATs are labor intensive and limited to specific CT serovars. We developed and validated a quantitative polymerase chain reaction (qPCR) assay that reproducibly detected CT serovars D, E, F, Ia, and Chlamydia muridarum over a linear range of 2 log 10 to 10 log 10 genomes with low coefficients of variation from both experimental and human urine samples. CT DNA loads from human vaginal, endocervical, and male urethral swabs correlated well with the BD ProbeTec ET assay (Becton Dickinson Diagnostic Systems, Franklin Lakes, NJ) run in parallel. In a preclinical microbicide evaluation, C. muridarum DNA loads in mouse swabs and tissues correlated well with an immunofluorescence assay. The optimized qPCR system provided enhanced sensitivity and facilitated the quantitative evaluation of clinical and experimental preclinical samples for anti-CT therapeutic and microbicide evaluation.

Nucleic Acid Amplification Tests for Diagnosis of Neisseria gonorrhoeae Oropharyngeal Infections

The optimal methods for the diagnosis of pharyngeal Neisseria gonorrhoeae infection are uncertain. The objective of this study was to define the performance of culture and nucleic acid amplification tests (NAATs) for the diagnosis of pharyngeal N. gonorrhoeae. In this cross-sectional study, males and females >15 years old who acknowledged performing fellatio or cunnilingus (in the previous 2 months) were recruited from three clinics (two human immunodeficiency virus clinics and one sexually transmitted diseases clinic) located in Birmingham, AL. The test performance of culture for N. gonorrhoeae, the Gen-Probe Aptima Combo 2 transcription-mediated amplification assay (TMA), the BD ProbeTec ET amplified DNA strand displacement assay (SDA), and the Roche Cobas Amplicor PCR was defined by using a rotating "gold standard" of any positive results by two or three of the three tests that excluded the test being evaluated. A total of 961 evaluable test sets were collected. On the basis of a rotating gold standard of positive results by two of three comparator tests, the sensitivity and the specificity were as follows: culture for N. gonorrhoeae, 50.0% and 99.4%, respectively; PCR, 80.3% and 73.0%, respectively; TMA, 83.6% and 98.6%, respectively; and SDA, 93.2% and 96.3%, respectively. On the basis of a rotating gold standard of positive results by three of three comparator tests, the sensitivity and specificity were as follows: culture for N. gonorrhoeae, 65.4% and 99.0%, respectively; PCR, 91.9% and 71.8%, respectively; TMA, 100% and 96.2%, respectively; and SDA, 97.1% and 94.2%, respectively. In conclusion, currently available NAATs are more sensitive than culture for the detection of pharyngeal gonorrhea in at-risk patients. PCR is substantially less specific than culture, TMA, or SDA and should not be used for the detection of pharyngeal gonorrhea.

Detection of Neisseria gonorrhoeae and Chlamydia trachomatis in pharyngeal and rectal specimens using the BD Probetec ET system, the Gen-Probe Aptima Combo 2 assay and culture

Objectives: This study compared the sensitivity and specificity of culture and two nucleic acid amplification tests (NAATs): the BD Probetec ET system (PT) and the Aptima Combo 2 (AC2) in...

Evaluation of the new COBAS TaqMan CT test v2.0 and impact on the proportion of new variant Chlamydia trachomatis by the introduction of diagnostics detecting new variant C trachomatis in Örebro county, Sweden

Background: The new variant of Chlamydia trachomatis (nvCT), discovered in Sweden in 2006, contains a 377-bp cryptic plasmid deletion, which includes the targets for the COBAS Amplicor/TaqMan C trachomatis/Neisseria gonorrhoea...

Evaluation of Self-Collected Vaginal Swab, First Void Urine, and Endocervical Swab Specimens for the Detection of Chlamydia Trachomatis and Neisseria Gonorrhoeae in Adolescent Females

Objective To assess the concordance of self-obtained vaginal swabs (SVS), first void urine samples (FVU) and provider-collected endocervical swabs (PES) for the detection of Chlamydia trachomatis (CT) and Neisseria gonorrhea (NG) in adolescents. Methods A total of 342 adolescent women and 1080 baseline and semi-annual visits were analyzed. FVU, SVS and PES were collected at each biannual visit. All specimens were tested by BDProbeTec ET ™ Amplified DNA Assay. Sensitivity, specificity, positive predictive value (PPV) negative predictive value (NPV) and kappa coefficient were calculated to evaluate the ability to identify possible infected cases using samples from three anatomic sites and the test agreement between any two of these three specimen types. Positive results from at least two of the three specimens collected from same subject at the same study visit was considered true positive. Results The positivity rates for CT and NG were 26.6 and 11.7 per 100 women respectively. The sensitivities of SVS, FVU and PES for detecting CT were 97.3%, 89.2% and 90.1% respectively. For the detection of NG, the sensitivities of the three sampling methods were 100%, 88.6% and 95.5% respectively. The specificities were between 94.7% and 99.7% for both CT and NG. Kappa coefficients of CT test results were 0.89, 0.88 and 0.83 for specimen pairs SVS vs PES, SVS vs FVU and PES vs FVU respectively. For the detection of NG, kappa coefficients were 0.91, 0.87 and 0.91 for these three pairs (all P < 0.0001). Kappa > 0.75 is considered excellent agreement between specimens. Conclusion There were strong agreements among SVS, PES and FVU specimens on the detection of CT and NG infections in adolescent females using nucleic acid amplification test. SVS represented as high as or more sensitive an approach for detecting both CT and NG compared to PES. Although FVU was the least sensitive sampling method, it is also the least invasive method. Thus SVS and FVU may provide a reliable alternative to endocervical specimens for CT and NG screening.

Impact of a Chemistry-Based DNA Extraction Method on Performance of a Commercial Amplification Assay for Detection of Mycobacterium tuberculosis Complex

Impact of a Chemistry-Based DNA Extraction Method on Performance of a Commercial Amplification Assay for Detection of <i>Mycobacterium tuberculosis</i> Complex

Vaginal swabs are appropriate specimens for the diagnosis of genital tract infection with Chlamydia trachomatis

Objectives. Self-administered vaginal swabs were used to determine the prevalence of Chlamydia trachomatis in a population under 25 years of age in order to evaluate if this is an acceptable and accurate method compared to the traditional cervical swabs. Methods. Patients were asked to perform a double self-administered vaginal swab prior to a traditional cervical swab. One self-collected vaginal swab was used with the AMPLIFIED CT DNA PROBING (GENPROBE bioMérieux) method that is based on a DNA probe against an amplified target for the identification of the Chlamydia trachomatis ribosomal RNA. The remaining two swabs were processed using BD PROBETEC ET (Becton Dickinson) for the determination of amplified DNA, using standard displacement amplification technology. Results. 8.2% of the 182 patients that were analysed between September 2005 and March 2006 tested positive. The two methods had an equivalent sensitivity and a specificity. It is important to concentrate prevention on young females who may have greater contacts with local specialists and could benefit from the self-administered vaginal swab technique.

Evaluation of the BD ProbeTec ET system for the direct detection of Mycobacterium tuberculosis from clinical samples

Early detection of disease caused by members of the Mycobacterium tuberculosis complex (MTBC) is essential in controlling transmission of tuberculosis. Clinical evaluation of the BD ProbeTec ET system for the direct detection of MTBC organisms in clinical specimens is carried out by comparing results obtained with conventional liquid culture and reference laboratory identification. In 186 specimens (159 respiratory, 27 non-respiratory), the BD ProbeTec ET assay detected eight out of 11 MTBC culture-positive specimens, with overall sensitivity, specificity, positive predictive value and negative predictive value of 85%, 100%, 100% and 99%, respectively. The data show that the BD ProbeTec ET assay is a highly sensitive and specific technique, and gives an accurate result from a clinical specimen within three to four hours.

男女尿検体におけるStrand displacement amplification (SDA) 法を用いた&lt;I&gt;Chlamydia trachomatis&lt;/I&gt; および&lt;I&gt;Neisseria gonorrhoeae&lt;/I&gt;の検出

Urine samples collected from 422 males and 53 females visiting a clinic in Kawasaki City who were suspected to have sexually transmitted infection were tested for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae by BD ProbeTecET (SDA method). The detection rates of C. trachomatis by the SDA method and polymerase chain reaction (PCR) method (control) were as high as 98.1% for C. trachomatis, and as high as 99.4% for N. gonorrhoeae, and the concordance rate of detection of both bacterial species was high. The detection sensitivity and specificity of the SDA method were 90.6 and 99.3%, respectively for C. trachomatis and 98.7% and 100% for N. gonorrhoeae, when PCR was used as the standard method. There were no differences in these results between males and females. The number of patients showing a discrepancy of the results obtained between the SDA method and the PCR method was 9 for C. trachomatis and 1 for N. gonorrhoeae, but the results of redetermination by the SDA method tended to coincide with those of the PCR method. Urine samples tested by the SDA method were positive for N. gonorrhoeae even in patients in whom the culture of secretions from the male urethra was negative for N. gonorrhoeae. Based on these results, the BD ProbeTecET (SDA method) was confirmed to have the equivalent capability to the PCR method for the detection of C. trachomatis and N. gonorrhoeae in urine samples.

Performance assessment of the Capilia TB assay and the BD ProbeTec ET system for rapid culture confirmation of Mycobacterium tuberculosis

Because of the increasing numbers of nontuberculous mycobacterial isolates from clinical specimens, rapid and accurate methods for culture confirmation of Mycobacterium tuberculosis are urgently needed. The study evaluated the performance of the Capilia TB immunochromatographic assay (TAUNS, Numazu, Japan) for culture confirmation of M. tuberculosis using 242 culture-positive liquid media in 2 mycobacterial laboratories from November 2005 to February 2006. Among the 242 samples, 183 were also tested with the BD ProbeTec ET (CTB) assay (Becton Dickinson, Sparks, MD). The results of both assays were compared to the culture results and to each other. The overall sensitivity and specificity of the Capilia TB assay were 98.6% and 97.9%, respectively, and for the CTB assay were 97.3% and 97.1%, respectively. The positive and negative predictive values for the Capilia TB assay were 98.6% and 97.9%, respectively, and for the CTB assay were 98.2% and 95.8%, respectively. Among the 183 samples tested with both assays, 8 had discrepant results, including Capilia-TB-false-positive in 2, CTB-false-positive in another 2, CTB-false-negative in 2, Capilia TB-false-negative in 1, and both assays with false-negative results in the remaining one. This study demonstrated that the Capilia TB assay has a similar diagnostic value with the CTB assay. In addition, with the immunochromatographic method, it is less time-consuming and does not require other laboratory equipment.

Evaluation of the BD ProbeTec ET System for Direct Detection of Mycobacterium bovis in Veterinary Specimens

We describe the application of the BD ProbeTec ET direct tuberculosis system for the detection of Mycobacterium bovis in bovine and cervine lymph node tissues. Compared to traditional culture, the overall sensitivity, specificity, and positive and negative predictive values of the BD ProbeTec were 87, 100, 100, and 87%, respectively.

R2190 Comparison of real-time PCR and BDProbeTec ET system for rapid diagnosis of tuberculosis patients

R2190 Comparison of real-time PCR and BDProbeTec ET system for rapid diagnosis of tuberculosis patients

Characteristics of the m 2000 Automated Sample Preparation and Multiplex Real-Time PCR System for Detection of Chlamydia trachomatis and Neisseria gonorrhoeae

We evaluated a new real-time PCR-based prototype assay for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae developed by Abbott Molecular Inc. This assay is designed to be performed on an Abbott m2000 real-time instrument system, which consists of an m2000sp instrument for sample preparation and an m2000rt instrument for real-time PCR amplification and detection. The limit of detection of this prototype assay was determined to be 20 copies of target DNA for both C. trachomatis and N. gonorrhoeae, using serially diluted linearized plasmids. No cross-reactivity could be detected when 55 nongonococcal Neisseria isolates and 3 non-C. trachomatis Chlamydia isolates were tested at 1 million genome equivalents per reaction. Concordance with the Roche Amplicor, BDProbeTec ET, and Gen-Probe APTIMA Combo 2 tests was assessed using unlinked/deidentified surplus clinical specimens previously analyzed with these tests. For C. trachomatis, concordance for positive results ranged from 93.7% to 100%, while concordance for negative results ranged from 98.2% to 100%. For N. gonorrhoeae, concordance for positive and negative results ranged from 91.4% to 100% and 99.3% to 100%, respectively. A workflow analysis of the prototype assay was conducted to obtain information on throughput under laboratory conditions. At 48 samples/run, the time to first result for both C. trachomatis and N. gonorrhoeae was 4.5 h. A total of 135 patient specimens could be analyzed in 8.9 h, with 75 min of hands-on time. This study demonstrated the technical and clinical feasibility of the new Abbott real-time PCR C. trachomatis/N. gonorrhoeae assay.

A Fast Real-Time Polymerase Chain Reaction Method for Sensitive and Specific Detection of the Neisseria gonorrhoeae porA Pseudogene

Ever since the advent of molecular methods, the diagnostics of Neisseria gonorrhoeae has been troubled by false negative and false positive results compared with culture. Commensal Neisseria species and Neisseria meningitidis are closely related to N. gonorrhoeae and may cross-react when using molecular tests comprising too-low specificity. We have devised a real-time polymerase chain reaction (PCR), including an internal amplification control, that targets the N. gonorrhoeae porA pseudogene. DNA was automatically isolated on a BioRobot M48. Our subsequent PCR method amplified all of the different N. gonorrhoeae international reference strains (n = 34) and N. gonorrhoeae clinical isolates (n = 176) but not isolates of the 13 different nongonococcal Neisseria species (n = 68) that we tested. Furthermore, a panel of gram-negative bacterial (n = 18), gram-positive bacterial (n = 23), fungal (n = 1), and viral (n = 4) as well as human DNA did not amplify. The limit of detection was determined to be less than 7.5 genome equivalents/PCR reaction. In conclusion, the N. gonorrhoeae porA pseudogene real-time PCR developed in the present study is highly sensitive, specific, robust, rapid and reproducible, making it suitable for diagnosis of N. gonorrhoeae infection.

Detection of Chlamydia trachomatis by Nucleic Acid Amplification Testing: Our Evaluation Suggests that CDC-Recommended Approaches for Confirmatory Testing Are Ill-Advised

We evaluated three CDC-suggested approaches for confirming positive nucleic acid amplification tests (NAATs) for Chlamydia trachomatis: (i) repeat the original test on the original specimen, (ii) retest the original specimen with a different test, and (iii) perform a different test on a duplicate specimen. For approach 1, specimens (genital swabs or first-catch urine [FCU]) initially positive by the Abbott LCx Probe System Chlamydia trachomatis Assay (LCx; Abbott Laboratories), the APTIMA Combo 2 Assay (AC2; Gen-Probe Inc.), the Amplicor CT/NG Assay (PCR; Roche Diagnostics Corp.), or the BD ProbeTec ET System C. trachomatis amplified-DNA assay (SDA; Becton Dickinson Diagnostic Systems) were retested by the same NAAT. In several evaluations, multiple efforts were made to confirm the original positive result. For approach 2, specimens initially positive by SDA and the Hybrid Capture 2 CT-ID DNA Test (HC2; Digene Corp.) were retested by different NAATs (SDA, PCR, AC2, and the APTIMA assay for C. trachomatis [ACT]). For approach 3, duplicate male urethral or cervical swabs were tested by SDA or by both AC2 and ACT. FCU specimens were tested by all three tests. We found that 84 to 98% of SDA, LCx, PCR, and AC2 positive results were confirmed by a repeat test and that 89 to 99% of SDA and AC2 and 93% of HC2 positive results were confirmed by different NAATs, but that some NAATs cannot be used to confirm other NAATs. The use of repeat testing did not confirm 11% of C. trachomatis SDA positive results that could be confirmed by more extensive testing. Doing more testing confirms more positive results; >90% of all positive NAATs could be confirmed.

Performance Assessment of the DR. MTBC Screen Assay and the BD ProbeTec ET System for Direct Detection of Mycobacterium tuberculosis in Respiratory Specimens

The performance of the DR. MTBC PCR-based assay and the BD ProbeTec ET Mycobacterium tuberculosis Complex Direct Detection (DTB) assay for the direct detection of Mycobacterium tuberculosis was evaluated using 1,066 consecutive clinical respiratory samples collected from 494 patients who did not have old cases of pulmonary tuberculosis and were not receiving antituberculosis treatment at National Taiwan University Hospital from January to February 2005. The results of both assays were compared to the "gold standard" of combined culture results and clinical diagnosis. The overall sensitivity and specificity of the DR. MTBC Screen assay were 56.6% and 98.9%, respectively, and of the DTB assay were 63.2% and 98.4%, respectively. The positive and negative predictive values for the DR. MTBC Screen assay were 84.5% and 95.4%, respectively, and for the DTB assay were 81.7% and 96.0%, respectively. The DR. MTBC Screen assay produced 11 false-positive results for 11 patients, including three samples yielding non-M. tuberculosis mycobacteria (one each for M. abscessus, a mixture of M. abscessus and M. chelonae, and unidentified non-tuberculosis mycobacteria). The DTB assay produced 15 false-positive results for 13 patients, including five samples from four patients yielding non-tuberculosis mycobacteria (two for M. abscessus, one for a mixture of M. abscessus and M. chelonae, and two for unidentified non-tuberculosis mycobacteria). This study demonstrated that the DR. MTBC Screen assay has a similar diagnostic value but fewer false-positive results than the DTB assay for respiratory specimens.

子宮頸管および咽頭擦過検体, 尿検体に対するSDA法を原理とする新しい核酸増幅法を用いた&lt;I&gt;Chlamydia trachomatis&lt;/I&gt;および&lt;I&gt;Neisseria gonorrhoeae&lt;/I&gt;の検出

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.

Comparison of the COBAS AMPLICOR MTB and BDProbeTec ET Assays for Detection of Mycobacterium tuberculosis in Respiratory Specimens

The performances of the BDProbeTec ET (Becton Dickinson) and COBAS AMPLICOR MTB (Roche) were retrospectively evaluated for detecting Mycobacterium tuberculosis complex in various respiratory specimens. The BACTEC and MGIT liquid culture system (Becton Dickinson) was used as a reference method. A total of 824 respiratory specimens, comprised of sputa, bronchoalveolar lavage fluid, and bronchial and tracheal aspirates from 580 patients, were evaluated. Out of 824 clinical specimens, 109 specimens from 43 patients were culture positive for M. tuberculosis. Of these 109 specimens, 67 were smear positive, 85 were positive by the COBAS AMPLICOR MTB test, and 94 were positive by the BDProbeTec ET. Of the 715 culture-negative specimens, 17 were positive by the auramine staining, 11 were positive by the COBAS AMPLICOR MTB test, and 12 were positive by the BDProbeTec ET. After discrepancy analysis and review of the patients' clinical data, 130 specimens from 50 patients were considered "true-positive" specimens. This resulted in the following sensitivities: microscopy, 61.5%; COBAS AMPLICOR MTB test, 78.0%; and BDProbeTec ET, 86.2%. The specificities of each system, based on the clinical diagnosis, were 99.7% for microscopy, 99.9% for the COBAS AMPLICOR MTB test, and 99.9% for the BDProbeTec ET. The data presented represent a considerable number of specimens evaluated with a considerable number of culture- and auramine-positive and culture-positive and auramine-negative results and therefore give a realistic view of how the data should be interpreted in a daily routine situation. Specifically, the data with regard to the culture-positive and auramine-negative specimens are useful, because in a routine situation, auramine-negative specimens are sometimes accepted, on clinical indications, to be analyzed by an amplification method.