The main objective of this study was to determine the prevalence of the Qnr determinants in clinical and environmental Aeromonas spp. A total of 52 Aeromonas sp. isolates (25 from natural waters, 27 from clinical samples in Valencia, Spain) were tested for quinolone resistance by the disk diffusion method. Among the isolates, only the clinical Aeromonas veronii A272 was resistant to both nalidixic acid and ciprofloxacin. Multiplex PCR screening for qnrA, qnrB, and qnrS genes showed A. veronii A272 carried qnrS2, which had 100% homology with qnrS2 previously reported in a wastewater treatment plant isolate from Germany and a non-Typhi Salmonella clinical isolate from the United States. The strain was identified as A. veronii by 16S rRNA gene sequencing. MICs for nalidixic acid, ciprofloxacin, and norfloxacin were 256, 8, and 12 mg/liter, respectively. Mutations in the quinolone resistance-determining regions of gyrA (Ser83→Ile) and parC (Ser80→Ile) were found. Conjugation experiments were negative, but transformation of rifampin-resistant E. coli J53 with the qnrS2-containing plasmid pA272 resulted in a 10- to more than 64-fold increase in quinolone MICs without increased resistance to other antibiotics. Genetic context analysis showed qnrS2 was inserted into an mpR gene. This is the first report of a qnrS-containing plasmid in a clinical Aeromonas isolate, highlighting the potential health risk of Qnr determinants spreading across different bacterial species.