Multicenter Validation of the
            <i>cppB</i>
            Gene as a PCR Target for Detection of
            <i>Neisseria gonorrhoeae</i>

Bruisten, Sylvia M.; Noordhoek, G. T.; Brule, Adriaan J. C. van den; Duim, Birgitta; Boel, C. H. E.; El-Faouzi, K.; Maine, Robin du; Mulder, S.J.; Luijt, D.; Schirm, J

doi:10.1128/jcm.42.9.4332-4334.2004

Cited by 40 publications

(36 citation statements)

References 15 publications

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“…However, subsequent melting curve analysis clearly distinguished NgNS from N. gonorrhoeae, with the N. gonorrhoeae T m always being above 64°C and the NgNS T m always being below 57°C. These results were confirmed in a study in which 450 well-characterized N. gonorrhoeae strains interspersed with negative controls and 20 NgNS strains were tested (2).…”

Section: Resultssupporting

confidence: 67%

Evaluation of Conventional and Real-Time PCR Assays Using Two Targets for Confirmation of Results of the COBAS AMPLICOR Chlamydia trachomatis / Neisseria gonorrhoeae Test for Detection of Neisseria gonorrhoeae in Clinical Samples

et al. 2005

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Two conventional PCR-enzyme immunoassays (PCR-EIAs) and two real-time PCR assays (LightCycler system; Roche Diagnostics) were evaluated as confirmation assays with cppB and 16S rRNA genes as targets. Of 765 male and female genitourinary and nasopharyngeal specimens positive for Neisseria gonorrhoeae in the COBAS AMPLICOR Chlamydia trachomatis/Neisseria gonorrhoeae PCR test (Roche Diagnostics), 229 (30%) were confirmed positive; 13 of these (5.7%) were lacking the cppB gene. Of the 534 samples (70%) that could not be confirmed, 81 (15%) showed a positive crossing point. However, melting curve analysis revealed an aberrant melting temperature in the LightCycler 16S rRNA assay; therefore, these samples were considered non-N. gonorrhoeae Neisseria species. Both of the 16S rRNA assays performed well, with positive predictive values of 99.1% and 100% for the PCR-EIAs and the real-time assays, respectively, and a negative predictive value of 99.8% for both. The cppB assays were compromised by the absence of the cppB gene in 5.7% of the N. gonorrhoeae-positive samples, resulting in negative predictive values of 96.8% and 97.6% for the PCR-EIAs and the real-time assays, respectively. Therefore, the 16S rRNA gene is preferable to the cppB gene as a target for confirmation assays. The melting curve analysis of the real-time assays provides useful additional information.Chlamydia trachomatis and Neisseria gonorrhoeae are the two most common causes of sexually transmitted infections (STI). Infection by these microorganisms can lead to clinical syndromes ranging from mild urethritis to pelvic inflammatory disease. However, both organisms can also cause asymptomatic infections in women and men; these infections can proceed in women to an ascending infection of the fallopian tubes (3,10,14). The outcome of the ascending infection can be reduced fertility, infertility, or even extrauterine pregnancies. Therefore, early diagnosis is necessary to prevent these complications and control the spread of infection.Several studies have shown that nucleic acid amplification tests are superior to conventional tests for the detection of C. trachomatis and N. gonorrhoeae (4,5,8). Because these organisms are regularly found together, the COBAS AMPLICOR Chlamydia trachomatis/Neisseria gonorrhoeae (CA CT/NG) multiplex PCR test (Roche Diagnostics, Mannheim, Germany) provides a useful platform for STI evaluation. An internal control greatly reduces false-negative results. The CA CT component has proven to be a useful diagnostic tool for the detection of C. trachomatis in cervical scraping and urine specimens (13,22,23). The CA NG component, however, lacks specificity, as some non-N. gonorrhoeae Neisseria species (NgNS) (8, 16) and even lactobacilli (23) have been reported to give falsepositive results. Therefore, confirmation by a second, more specific PCR assay is required, especially in low-prevalence communities (8,15,22). This second PCR assay should be convenient and fast to ensure that the delay introduced by required confirmation is kept ...

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Section: Resultssupporting

confidence: 67%

Evaluation of Conventional and Real-Time PCR Assays Using Two Targets for Confirmation of Results of the COBAS AMPLICOR Chlamydia trachomatis / Neisseria gonorrhoeae Test for Detection of Neisseria gonorrhoeae in Clinical Samples

et al. 2005

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“…Recently, two independent studies investigated the use of N. gonorrhoeae cppB PCR assays on populations in Australia and the Netherlands. 50, 51 Tabrizi et al 51 found no false-negative results and considered the cppB gene assay to be suitable for use in a population in Victoria, Australia, whereas Bruisten et al 50 found a high incidence (5.8%) of false-negative results and concluded that the cppB gene target was not suitable for use in the Dutch population. Interestingly, other Australian studies have revealed a high incidence of cppB-related false-negative PCR results in a patient population in the Northern Territory in Australia, due to the recent appearance and expansion of a non-PAU Ϫ gonococcal subtype.…”

Section: Sequence-related Challenges For Gonorrhea Naat Assaysmentioning

confidence: 99%

“…However, cppB gene-based NAAT assays are now no longer considered adequate for some patient populations because of the limitations outlined previously. 50 More recently, a PCR assay targeting the N. gonorrhoeae porA pseudogene showed considerable promise as a suitable supplementary test for the Amplicor assay. 91 So far, this assay has only been validated in an Australian population and will need further validation in other populations.…”

Section: Roche Cobas Amplicormentioning

confidence: 99%

“…Yet as previously discussed, the cppB gene has serious limitations as a NAAT target in many patient populations. 50 More recently, a PCR assay targeting the multicopy opa genes was described previously. 103 This assay was evaluated using 135 clinical samples and a panel of 173 microorganisms, including 73 nongonococcal Neisseria strains, and reported 100% sensitivity and specificity.…”

Section: In-house Assaysmentioning

confidence: 99%

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Nucleic Acid Amplification Testing for Neisseria gonorrhoeae

Whiley

Tapsall

Sloots

2006

The Journal of Molecular Diagnostics

178

163

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Nucleic acid amplification tests (NAATs) for the detection of Neisseria gonorrhoeae became available in the early 1990s. Although offering several advantages over traditional detection methods, N. gonorrhoeae NAATs do have some limitations. These include cost, risk of carryover contamination, inhibition, and inability to provide antibiotic resistance data. In addition, there are sequence-related limitations that are unique to N. gonorrhoeae NAATs. In particular, falsepositive results are a major consideration. These primarily stem from the frequent horizontal genetic exchange occurring within the Neisseria genus, leading to commensal Neisseria species acquiring N. gonorrhoeae genes. Furthermore, some N. gonorrhoeae subtypes may lack specific sequences targeted by a particular NAAT. Therefore, NAAT false-negative results because of sequence variation may occur in some gonococcal populations. Overall, the N. gonorrhoeae species continues to present a considerable challenge for molecular diagnostics. The need to evaluate N. gonorrhoeae NAATs before their use in any new patient population and to educate physicians on the limitations of these tests is emphasized in this review.

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“…Further, considerable recombination occurs within the DNA of the plasmid and some cppB gene sequences present in the chromosome were incomplete and in low copy numbers (6,7). It is thus not surprising that some decreased sensitivity with cppB gene-based nucleic acid amplification, arising from absent, altered, or low copy numbers of the chosen sequence, has now been acknowledged with different forms of this test (2,9).…”

mentioning

confidence: 99%

Cryptic-Plasmid-Free Gonococci May Contribute to Failure of cppB Gene-Based Assays To Confirm Results of BD ProbeTEC PCR for Identification of Neisseria gonorrhoeae

et al. 2005

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Multicenter Validation of the cppB Gene as a PCR Target for Detection of Neisseria gonorrhoeae

Abstract: The cppB gene is often used as a target for detection of Neisseria gonorrhoeae by PCR. Using a coded panel of 500 DNA samples, we determined that the cppB gene is missing in 5.8% of N. gonorrhoeae strains, and therefore we consider the cppB gene to be an unsuitable target

Cited by 40 publications

References 15 publications

Evaluation of Conventional and Real-Time PCR Assays Using Two Targets for Confirmation of Results of the COBAS AMPLICOR Chlamydia trachomatis / Neisseria gonorrhoeae Test for Detection of Neisseria gonorrhoeae in Clinical Samples

Evaluation of Conventional and Real-Time PCR Assays Using Two Targets for Confirmation of Results of the COBAS AMPLICOR Chlamydia trachomatis / Neisseria gonorrhoeae Test for Detection of Neisseria gonorrhoeae in Clinical Samples

Nucleic Acid Amplification Testing for Neisseria gonorrhoeae

Cryptic-Plasmid-Free Gonococci May Contribute to Failure of cppB Gene-Based Assays To Confirm Results of BD ProbeTEC PCR for Identification of Neisseria gonorrhoeae

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