A novel commensal Neisseria species harboring the gonococcal diagnostic marker DR-9 causes false-positive Roche cobas NAAT results

Abstract

Neisseria gonorrhoeae (Ng), the etiologic agent of gonorrhea, is the second most reported bacterial sexually transmitted infection globally. In the USA, nucleic acid amplification tests (NAATs) are considered the gold standard for diagnosis. Although NAATs are sensitive and offer high throughput, cross-reactivity with commensal Neisseria can compromise specificity. Here, we report the isolation and characterization of a novel commensal Neisseria sp. from the oropharynx of a patient with suspected gonococcal treatment failure. An initial diagnosis was made using the Roche cobas CT/NG test on the cobas 8800 system, which repeatedly yielded positive results post-treatment. A confirmatory Aptima Combo 2 (AC2, Hologic) test was negative, and multiple culture attempts failed to isolate Ng. Instead, a commensal Neisseria strain was recovered. Whole-genome sequencing revealed this isolate shared partial genomic identity with several Neisseria spp., including Ng, and carried a distinct region encoding the gonococcal diagnostic marker DR-9, the target of cobas assays. Cross-reactivity was confirmed with the cobas 4800 and 6800 platforms, but not with the AC2 assay targeting 16S rRNA. We discuss the possible origins of the DR-9 marker on this strain as well as examine its antimicrobial susceptibility profile and genomic resistance markers. These findings highlight the potential for misdiagnosis and unnecessary treatment when commensal Neisseria species harbor Ng diagnostic targets. Our study underscores the need for genomic surveillance of Ng and commensal Neisseria, not only to monitor diagnostic performance but also to track commensals that may serve as reservoirs of antimicrobial resistance determinants and contribute to the spread of resistance.IMPORTANCEAccurate diagnosis of gonorrhea is critical for effective treatment and antimicrobial stewardship. Nucleic acid amplification tests, the mainstay of gonococcal diagnostic testing, can yield false-positive results due to genetic overlap between Neisseria gonorrhoeae and commensal Neisseria species, especially from extragenital sites like the oropharynx. Prior studies, such as Hopkins et al. (2023), have recognized this limitation and proposed supplemental tests to improve specificity for oropharyngeal specimens. Here, we describe a novel commensal Neisseria strain isolated from a patient with suspected treatment failure that harbors the gonococcal diagnostic marker DR-9. This case highlights the need for confirmatory testing using an alternate gene target in cases where repeated positive tests are obtained with extragenital specimens and demonstrates the need for improved test specificity, particularly for anatomical sites such as the pharynx, which has high commensal diversity. Enhanced molecular surveillance of commensal Neisseria populations will be vital for understanding and minimizing diagnostic cross-reactivity.