Analysis of antimicrobial susceptibility testing methods and variables and in vitro activity of gepotidacin against urogenital Neisseria gonorrhoeae in men

•New antibiotics are needed to treat antibiotic-resistant N. gonorrhoeae infections.•Gepotidacin is a novel triazaacenaphthylene antibiotic.•Gepotidacin demonstrated activity against N. gonorrhoeae isolates in vitro.•Media type, inoculum, CO2, and pH can impact MIC testing with gepotidacin. Gepotidacin is a triazaacenaphthylene antibiotic with activity against Neisseria gonorrhoeae including strains resistant to current agents. We tested 145 N. gonorrhoeae isolates by agar dilution according to Gonococcal Isolate Surveillance Program and Clinical and Laboratory Standards Institute methodologies. Gepotidacin demonstrated a minimum inhibitory concentration (MIC)50 of 0.25 μg/mL and a MIC90 of 0.5 μg/mL (highest gepotidacin MIC was 1 μg/mL) against the 145 N. gonorrhoeae isolates tested. We also assessed the impact of test variables on antimicrobial susceptibility test results for gepotidacin, ciprofloxacin, and ceftriaxone against 10 N. gonorrhoeae isolates. Media type had the biggest effect but wasn't specific to gepotidacin. Gepotidacin MICs were also affected by inoculum, pH, and 10% CO2. These in vitro data indicate that further study of gepotidacin is warranted for potential use in treating gonorrhea infections and highlight the importance of controlling for media type, inoculum, CO2, and pH when performing MIC testing with gepotidacin.