AIM: To identify the location and the relationship, and to analyze the genetic background of 2 plasmid-mediated quinolone resistance genes, qnrB4 and qnrS1, carried by a clinical strain of Klebsiella pneumoniae (K pneumoniae). METHODS: The plasmids carrying qnrB4 or qnrS1 were identified by Southern blotting. A HindIII fragment containing qnrB4 or qnrS1 was cloned into plasmid puc18 and sequenced. RESULTS: qnrB4 and qnrS1 were located on 2 different plasmids, pHS7 and pHS8, and were 180 and 45 kb in size, respectively. A transconjugant carrying plasmid pHS7 bearing qnrB4 and another transconjugant carrying pHS9 bearing qnrB4 and qnrS1 were obtained by conjugation. Plasmid pHS8 bearing qnrS1 was also transferred to J53 by transformation. The ciprofloxacin minimal inhibitory concentrations (MIC) for J53 transconjugants or the transformant carrying qnrB4 only, qnrS1 only, and both qnrB4 and qnrS1 were 0.19, 0.25, and 0.25 mg/L, respectively, while the parent clinical strain of K pneumoniae had a MIC of 0.75 mg/L. qnrB4 was located in a sul1-type integron with blaDHA-1, ampR and psp genes in upstream and insertion sequence IS26, and sap genes in downstream of qnrB4. qnrS1 was not located in an integron, but IS26 was found both upstream and downstream, and IS2 was found directly upstream of qnrS1. CONCLUSION: qnrB and qnrS can be harbored simultaneously by a single clinical strain of K pneumoniae. These 2 genes are carried by 2 different plasmids and have different genetic environments in plasmid DNA structure.