ABSTRACT Since its discovery, qnrA has been found in most common Enterobacteriaceae . Ciprofloxacin MICs conferred by different qnrA -positive plasmids could range from 0.1 μg/ml to 2 μg/ml in Escherichia coli J53. The reasons for different ciprofloxacin MICs conferred by qnrA have not been fully clarified. Five hundred forty-one consecutive gram-negative clinical strains that were resistant or intermediate to ciprofloxacin and that were isolated in Shanghai in 2005 were screened for qnrA by PCR. For qnrA -positive isolates, the transferability of quinolone resistance was determined by conjugation and mutations within the quinolone resistance-determining region (QRDR) of gyrA and parC. aac ( 6 ′)- Ib-cr was detected and qnrA RNA expression was determined using real-time reverse transcription-PCR for transconjugants with different ciprofloxacin MICs. The qnrA gene was detected in 7 of the 541 clinical isolates. Quinolone resistance was transferred in four strains by conjugation. Mutations in the QRDR of gyrA and parC were detected in five qnrA -positive clinical strains with higher ciprofloxacin MICs. Of four qnrA -bearing plasmids in E. coli J53, pHS4 and pHS5 conferred ciprofloxacin MICs of 0.094 to 0.125 μg/ml; pHS3, which harbored the aac ( 6 ′)- Ib-cr gene as well, conferred a ciprofloxacin MIC of 0.25 μg/ml, and pHS6, which had both the aac ( 6 ′)- Ib-cr gene and a high expression level of qnrA , had a ciprofloxacin MIC of 1.0 μg/ml. The prevalence of qnrA appeared to be higher in Enterobacter cloacae than in other Enterobacteriaceae . The coexistence of qnrA and aac ( 6 ′)- Ib-cr in a single plasmid and increased qnrA expression can account for the different levels of ciprofloxacin resistance seen in transconjugants.