Many diagnostic nucleic acid tests (NAATs) for Neisseria gonorrhoeae (NG), particularly earlier generation tests, have been beset with specificity problems associated with cross-reaction with commensal Neisseria. To address, supplementary testing (whereby samples testing positive in a screening NAAT are reflexively tested with a secondary NAAT) has been widely implemented. Many third-generation commercial NAATs are highly sensitive (<10 CFU/mL) and have progressed to include testing claims for oropharyngeal and anorectal swabs. Here we investigate the utility of the ResistancePlusGC (RP-GC) assay as a supplemental assay, which in addition to detecting NG, also predicts ciprofloxacin susceptibility via NG gyrA characterisation (1). We also describe a quantitative c6800 method to investigate discrepant screening and supplementary results.
We compared two nucleic acid extraction techniques for RP-GC: extracts from c4800 (c4800-RP-GC) and MagNA Pure 96 (MP96-RP-GC). NG-positive (n=300) and negative (n=150) samples from routine c4800 testing were retrospectively retested with c4800, c6800, c4800-RP-GC and MP-96-RP-GC. Selected samples were tested with Xpert CT/NG (Xpert) for discrepant analysis and compared to quantitative results. The gyrA status was also compared to ETEST ciprofloxacin susceptibility or non-susceptibility for recovered isolates (n=63).
Extragenital confirmatory rates were higher for MP96-RP-GC (131/140; 93.6%) compared to c4800-RP-GC (126/146; 86.3%), albeit not significantly (P=0.6677). Of 9 samples testing positive by c6800 and negative by MP-96-RP-GC, 7/9 (77.8%) were also negative by Xpert. These samples demonstrated NG loads ranging from 2 to 2500 CFU/mL yet were negative for RP-GC and Xpert (experimental sensitivity 100 CFU/mL and 10 CFU/mL, respectively).
By contrast, the number of samples returning a valid gyrA status were significantly (P=0.0003) higher for MP96-RP-GC (270/293; 92.2%) compared to c4800-RP-GC (245/298; 82.2%). The overall MP-96-RP-GC gyrA status correlated 98.4% (61/62) with the reported ciprofloxacin sensitive (35/36; 97.2%) or non-susceptible (26/26; 100%) phenotype.
Improved RP-GC confirmatory rates and reported gyrA status were observed using MP96 nucleic acids compared to c4800 extracts. We demonstrate the ongoing need for supplemental testing for oropharyngeal samples based on the quantitative discrepant analysis. We also highlight the importance of genotypic AMR detection, as a progressive step toward specific individualised treatment to reduce the growing state of AMR in gonococcus.