Evaluation of the New BD Max GC Real-Time PCR Assay, Analytically and Clinically as a Supplementary Test for the BD ProbeTec GC Qx Amplified DNA Assay, for Molecular Detection of Neisseria gonorrhoeae

Abstract

The new BD Max GC real-time PCR assay showed high clinical and analytical sensitivity and specificity. It can be an effective and accurate supplementary test for the BD ProbeTec GC Qx amplified DNA assay, which had suboptimal specificity, and might also be used for initial detection of Neisseria gonorrhoeae. N eisseria gonorrhoeae is estimated to cause 106 million gonorrhea cases every year, resulting in substantial morbidity and economic costs globally (1). The need for highly sensitive and specific laboratory diagnostics and subsequent effective therapy is crucial. Nucleic acid amplification tests (NAATs) have increasingly replaced culture diagnostics internationally, due to their higher sensitivity, speed, automation, and use of noninvasive specimens (2-7). However, due to suboptimal specificity, especially of several early generation gonococcal NAATs, in several regions (e.g., Europe and Australia), it is recommended to verify positive samples with another NAAT targeting a different genetic sequence, particularly in low-prevalence populations and in pharyngeal infections (6,(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18). The psychological, social, and legal consequences of false-positive gonococcal test results can be substantial.The BD Viper System with XTR technology (BD Diagnostics, Sparks, MD) is a third-generation platform that when operating in "extraction mode" provides automated DNA extraction using ferric oxide and strand displacement amplification (19). The BD ProbeTec GC Qx amplified DNA assay, targeting a pilin-inverting gene, is used on the BD Viper system to detect N. gonorrhoeae. However, suboptimal (particularly analytical) specificity and cross-reaction with commensal Neisseria species have been described for this assay as well as most other gonococcal NAATs (6,8,13,20). Recently, the BD Max GC real-time PCR assay, targeting the gonococcal opcA gene, was developed to be run on the BD Max system, which provides automated DNA extraction and realtime PCR.We evaluated the performance of the new BD Max GC realtime PCR assay by examining clinical specimens positive in the BD ProbeTec GC Qx amplified DNA assay and samples spiked with isolates of gonococci, nongonococcal Neisseria species, and other closely related bacteria.During July to October 2014, 23,815 individuals (14,846 females and 8,969 males representing asymptomatic individuals presenting for screening and symptomatic patients) were tested with the BD ProbeTec GC Qx amplified DNA assay in a single replicate according to routine diagnostic protocol. All positive clinical specimens were subsequently stored in the primary tube (including BD transportation medium) prior to analysis (DNA extraction and real-time PCR) with the BD Max GC real-time PCR assay, which was performed within 1 to 12 h. Specimens negative in the BD Max GC real-time PCR assay were further tested with the Aptima Combo 2 assay (Hologic, Bedford, MA) and a gonococcal dual-target real-time PCR targeting the porA pseudogene and opa genes (21).To challenge the analytical sensitiv...

Background review for the ‘2020 European guideline for the diagnosis and treatment of gonorrhoea in adults’

“…1,3,36–40,54–57 Accordingly, vulvo-vaginal swabs (health care worker- or self-collected) are recommended in women due to their superior sensitivity and ease of sampling [1A]. 1,3,36–41,47,54–57 The performance characteristics of different commercially available or in-house gonococcal NAATs can differ substantially in sensitivity and particularly specificity, 1,3,30,58–60 and appropriate validation and quality assurance of all diagnostic NAATs are crucial.NAATs are significantly more sensitive than culture 1,3,30,35,38–41,49,52,53,61–67 for detection of rectal and oropharyngeal gonorrhoea due to frequently lower gonococcal load compared to urogenital infection. 61,62 Optimal sampling technique 68 and appropriate, validated and quality assured NAATs are recommended for testing and/or screening for infections at these sites.…”

“…61,62 Optimal sampling technique 68 and appropriate, validated and quality assured NAATs are recommended for testing and/or screening for infections at these sites. 1,3,38–41,69,70 However, most commercially available gonococcal NAATs are not licensed for testing oropharyngeal and rectal specimens, and differ substantially in their specificity, 1,3,30,38,40,58–60 particularly when examining oropharyngeal specimens due to the frequent presence of non-gonococcal Neisseria species. At present, only two commercially available gonococcal NAATs are approved by the US Food and Drug Administration (FDA) for testing rectal and oropharyngeal specimens.…”

“…1,3,30,58,60 If the diagnostic NAAT used does not display a PPV exceeding 90%, positive specimens should be subjected to confirmatory testing, i.e. repeated testing with a NAAT targeting another genetic sequence, particularly if oropharyngeal specimens are tested [1C]; 1,3,30,38,39,59,60,69–73 Point-of-care tests (POCTs): rapid, validated and quality-assured POCTs for diagnosis of gonorrhoea with sufficient sensitivity compared to NAATs are still lacking; however, several NAAT-based POCTs with high sensitivity and specificity are in late development (https://www.who.int/reproductivehealth/topics/rtis/Diagnostic-Landscape-for-STIs-2019.pdf). 1,74–76 Ideally, such tests should combine detection of N. gonorrhoeae and antimicrobial resistance prediction.…”

“…1,3,36–40,54–57 Accordingly, vulvo-vaginal swabs (health care worker- or self-collected) are recommended in women due to their superior sensitivity and ease of sampling [1A]. 1,3,36–41,47,54–57 The performance characteristics of different commercially available or in-house gonococcal NAATs can differ substantially in sensitivity and particularly specificity, 1,3,30,58–60 and appropriate validation and quality assurance of all diagnostic NAATs are crucial.…”

Comparison between DiaPlexQ™ STI6 and GeneFinder™ STD I/STD II multiplex Real‐time PCR Kits in the detection of six sexually transmitted disease pathogens

“…Recent advances in molecular testing for the detection of STD pathogens have led to changes in the diagnosis of STIs. NAATs have increasingly replaced conventional diagnostics due to their superior sensitivity and speed . Variable molecular technologies for the detection of STD pathogens from clinical specimens have been introduced including strand displacement amplification, transcription‐mediated amplification, and real‐time PCR .…”