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Background Fluoroquinolone agents, such as enrofloxacin and marbofloxacin, are commonly used in pseudomonal infection in veterinary medicine. However, the resistance rate to fluoroquinolone is rapidly increasing according to multiple studies in various countries. The point mutation in quinolone resistance determining region (QRDR) is closely related to increased fluoroquinolone resistance of Pseudomonas aeruginosa . The aim of this study was to investigate current antimicrobial susceptibility and fluoroquinolone resistance in Pseudomonas aeruginosa strains isolated from dogs. The presence of the point mutations in the QRDR was confirmed by gyrA and parC polymerase chain reaction and nucleotide sequencing analysis. Results A total of 84 nonduplicated P. aeruginosa isolates were obtained from 228 healthy dogs (healthy group) and 260 dogs with clinical signs (infected group). From these, 38 isolates from the healthy group were detected in several samples, whereas 46 isolates from the infected group were mostly obtained from dogs’ ears with otitis externa (41/260, 15.8%). All isolates were resistant to nalidixic acid, while some were also resistant to enrofloxacin (23/84, 27.4%), marbofloxacin (17/84, 20.2%), levofloxacin (12/84, 14.3%), or ciprofloxacin (11/84, 13.1%). Enrofloxacin resistance was significantly higher in strains from the infected group than that of the healthy group ( p |