In the study, the ciprofloxacin resistance rate was 100%. High-level ciprofloxacin resistance rate was 63.55%. Sixteen different mutation patterns involved in the formation of ciprofloxacin resistance were identified. The most prevalent were patterns P7 (25.2%), P8 (15.0%), P9 (11.2%), P1 (10.3%), and P5 (10.3%). All of the 107 NG isolates analyzed for mutations in the study have demonstrated a change of Ser-91 → Phe in the gyrA gene, and all except one have demonstrated a change in position 95 of the amino acid sequence. All of the 68 high-level QRNG isolates had double mutations in gyrA gene combined with a single or two mutations in parC gene. It is most important that a new mutation site of Ile-97 → Met in gyrA and a new mutation of Leu-106 → Ile in parC were found in the study, both leading to high-level ciprofloxacin resistance (MIC values, 8 µg/mL, 32 µg/mL, respectively). Therefore, we confim that gyrA mutations are necessary for the fluoroquinolone resistance phenotype and parC mutations are correlated intimately with high-level fluoroquinolone resistance. In China fluoroquinolone resistance in Neisseria gonorrhoeae strains is very serious and the new mutation sites in the fluoroquinolone resistance-determining regions emerge more and more quickly. Hence, in China fluoroquinolones, which are used to treat gonorrhoea presently, should be substituted by a new antibiotics.