The first wide-scale drug repurposing screen using the Prestwick Chemical Library (1200 bioactive molecules) against Neisseria gonorrhoeae identifies high in vitro activity of auranofin and many additional drugs.

Treatment options for gonorrhoea are scarce. Drug repurposing of bioactive molecules approved for other conditions might therefore be of value. We developed a method for wide-scale, systematic drug repurposing screen to identify molecules with activity against Neisseria gonorrhoeae and screened the Prestwick Chemical Library (1200 FDA-approved drugs). As a proof-of-concept, we further examined one promising and interesting screening hit (auranofin; antirheumatic agent). Three WHO gonococcal reference strains (WHO F, O, P) were used for the Library screening. The strains were grown in presence of a fixed concentration of the library drugs in 384-well plates for 12 h, and the remaining bacterial respiration, to reflect growth, was then quantitatively measured using optical density (OD) 450 nm and a resazurin assay. The activity of auranofin was further examined using in vitro susceptibility testing (minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC)) against genetically diverse antimicrobial-resistant N. gonorrhoeae strains and time-kill assays. Sixty-eight molecules significantly inhibited bacterial growth of WHO F, O and P. Auranofin showed potent in vitro bactericidal activity (in MIC-, MBC- and time-kill assays) against four WHO reference strains. No cross-resistance between auranofin and any antimicrobial currently or previously used for gonorrhoea treatment was found when examining 51 selected antimicrobial-resistant gonococcal strains. In conclusion, this is the first wide-scale systematic screening effort for repurposing drugs for future treatment of gonorrhoea. Additional studies examining mechanism(s) of action, resistance development, in vivo anti-gonococcal activity and pharmacokinetics/pharmacodynamics for gonococcal infections of auranofin and several other significant screening hits would be valuable.

A new rapid resazurin-based microdilution assay for antimicrobial susceptibility testing of Neisseria gonorrhoeae.

Objectives: Rapid, cost-effective and objective methods for antimicrobial susceptibility testing of Neisseria gonorrhoeae would greatly enhance surveillance of antimicrobial resistance. Etest, disc diffusion and agar dilution methods are subjective, mostly laborious for large-scale testing and take ∼24 h. We aimed to develop a rapid broth microdilution assay using resazurin (blue), which is converted into resorufin (pink fluorescence) in the presence of viable bacteria. Methods: The resazurin-based broth microdilution assay was established using 132 N. gonorrhoeae strains and the antimicrobials ceftriaxone, cefixime, azithromycin, spectinomycin, ciprofloxacin, tetracycline and penicillin. A regression model was used to estimate the MICs. Assay results were obtained in ∼7.5 h. Results: The EC 50 of the dose-response curves correlated well with Etest MIC values (Pearson's r = 0.93). Minor errors resulting from misclassifications of intermediate strains were found for 9% of the samples. Major errors (susceptible strains misclassified as resistant) occurred for ceftriaxone (4.6%), cefixime (3.3%), azithromycin (0.6%) and tetracycline (0.2%). Only one very major error was found (a ceftriaxone-resistant strain misclassified as susceptible). Overall the sensitivity of the assay was 97.1% (95% CI 95.2-98.4) and the specificity 78.5% (95% CI 74.5-82.9). Conclusions: A rapid, objective, high-throughput, quantitative and cost-effective broth microdilution assay was established for gonococci. For use in routine diagnostics without confirmatory testing, the specificity might remain suboptimal for ceftriaxone and cefixime. However, the assay is an effective low-cost method to evaluate novel antimicrobials and for high-throughput screening, and expands the currently available methodologies for surveillance of antimicrobial resistance in gonococci.