In Vitro Activity of Zoliflodacin Against Neisseria gonorrhoeae Isolates from Shanghai, China (2020–2023)

<b>Background/Objectives:</b> The escalating threat of drug-resistant <i>Neisseria gonorrhoeae</i> underscores the urgent need for novel therapeutic agents. Zoliflodacin, a first-in-class spiropyrimidinetrione antibiotic that targets bacterial DNA gyrase and topoisomerase IV, represents a promising candidate for gonorrhea treatment. <b>Methods:</b> From 2020 to 2023, a total of 876 urogenital <i>N. gonorrhoeae</i> isolates were collected from 35 hospitals across Shanghai, China. In vitro susceptibilities to zoliflodacin and six conventional antibiotics (penicillin, tetracycline, ciprofloxacin, azithromycin, ceftriaxone, and spectinomycin) were determined using the agar dilution method. Whole-genome sequencing was conducted to identify sequence types (STs) and amino-acid substitutions in GyrA, GyrB, ParC, ParE, and MtrR. <b>Results:</b> Zoliflodacin exhibited potent in vitro activity, with minimum inhibitory concentrations (MICs) ranging from ≤0.004 to 0.25 mg/L (MIC₅₀ = 0.06 mg/L; MIC₉₀ = 0.125 mg/L), all below the breakpoint (0.5 mg/L). Notably, zoliflodacin maintained high activity against isolates resistant to ceftriaxone, azithromycin, ciprofloxacin, penicillin, and tetracycline. Although all isolates were susceptible to zoliflodacin, elevated MIC values were observed in ST7363 and ST8123 compared with other clones. Genomic analysis identified no substitutions associated with increased zoliflodacin MICs, and most GyrB sequences, the key gene associated with zoliflodacin resistance, remained intact. <b>Conclusions:</b> These findings demonstrate that zoliflodacin possesses robust activity against circulating multidrug-resistant <i>N. gonorrhoeae</i> lineages in Shanghai and support its potential clinical use for the treatment of gonorrhea. Continued genomic and phenotypic surveillance is warranted to preserve the long-term efficacy of this novel agent.