Caution Required When Using Supplementary Nucleic Acid Amplification Tests for Gonorrhea

Research results highlighted the need for continued supplemental NAATs for oropharyngeal samples being tested for Nisseria gonorrhoeae.

Supplementary nucleic acid amplification tests (NAATs) for Neisseria gonorrhoeae should be chosen with caution to avoid discrepancies with the screening NAAT, according to a study recently published in the European Journal of Clinical Microbiology & Infectious Diseases.

There are an estimated 78 million cases of gonorrhea globally each year. Management of gonorrhea is dependent on accurate diagnosis and the subsequent treatment with effective antimicrobials. While microscopy and culture were historically used for the microbiological diagnosis of N gonorrhoeae, these techniques have been replaced with more sensitive NAATs over the last 2 decades.

Unfortunately, early N gonorrhoeae-NAATs have demonstrated specificity problems, mostly associated with oropharyngeal swab testing. Most commercial N gonorrhoeae-NAATs are approved for urogenital sample testing but do not have specific claims for extragenital sites, including oropharyngeal and anorectal swabs; however, NAATs are still widely used and recommended for these samples due to their superior sensitivity, especially in high-risk patients.

Australian guidelines recommend using a supplemental confirmatory test to help facilitate testing of extragenital sites to circumvent these specificity problems; therefore, a positive sample test via screening N gonorrhoeae-NAAT is confirmed by a second NAAT. However, there have been increasing observations and concerns from local sexual health physicians over increased discrepancies between the screening and supplementary test results. In the study lab, increased discrepancies were noticed between Roche cobas 4800 CT/NG (c4800) and the in-house supplementary N gonorrhoeae-PCR (NG-duplex) when compared with Abbott RealTime CT/NG (m2000) for oropharyngeal samples. Researchers set out to investigate these discrepancies.

In total, 3 banks of N gonorrhoeae-positive samples were used to test the confirmatory rates of the NAATs used in the study lab: Bank 1 (n=344), Bank 2 (n=344), and Bank 3 (n=400). Samples in Bank 1 were screened using m2000, while samples in Banks 2 and 3 were screened using c4800. Bank 2 samples were further tested using m2000, and some samples were selectively tested using Cepheid Xpert CT/NG. Bank 3 samples were further tested using cobas CT/NG (cobas 6800 system).

Study results emphasized the need for careful selection of N gonorrhoeae supplemental nucleic acid testing methods for oropharyngeal samples. When compared with c4800, the confirmatory rates for the m2000 were significantly higher especially for oropharyngeal samples (P <.0001). The NG-duplex also failed to confirm some true-positive N gonorrhoeae samples. When an expanded gold standard was applied, confirmatory rates for both m2000 and c4800 exceeded 90% for all anatomical sites except for the confirmatory rate for c4800 with oropharyngeal samples, which was 78%. The discrepancies observed in this study were due to a combination of false-positive results for oropharyngeal samples from c4800 testing and NG-duplex-related sensitivity issues.

Study limitations include the use of cobas PCR media, which are not an approved collection media for m2000 or Xpert, resulting in potential for false-negative test results, and the decision to not conduct all tests at the same time. Although samples should be stable for 12 months, the investigators noted that some retrospective testing took place 3 to 6 months after the initial sample collection.

“[O]ur experiences here further highlight the need for maintaining a solid rapport between the laboratory and the clinicians receiving the results, particularly so as to minimize any potential negative impact that changes in testing practice may have upon patient management,” the researchers concluded.

Disclosure: Several study authors declared affiliations with the pharmaceutical industry. Please see the original reference for a full list of authors’ disclosures.


Pryce TM, Hiew VJ, Haygarth EJ, Whiley DM. Second- and third-generation commercial Neisseria gonorrhoeae screening assays and the ongoing issues of false-positive results and confirmatory testing [published online August 7, 2020]. Eur J Clin Microbiol Infect Dis. doi: 10.1007/s10096-020-04004-5