To explore the prevalence of the plasmid-mediated quinolone resistance gene qnrA in Gramnegative bacteria and to investigate its molecular genetic background and resistance profile in isolates harboring this gene, a total of 629 nalidixic acid-resistant isolates of non-repetitive Gram-negative bacteria were collected from clinical specimens between April 2004 and April 2006 and these isolates were screened for qnrA gene by PCR using specific primers combined with DNA sequencing. The extended spectnan β-lactamase (ESBL) or AmpC-producing isolates were distinguished by the phenotypic confirmatory test combined with DNA sequencing, and the antibiotics susceptibility test for qnrA-positive isolates was carried out by Kirby-Bauer and E-test method. To detect the location of the qnrA gene, plasmid conjugation and Southe hybridization were performed and the integron structure containing the qnrA gene was cloned by PCR strategy and sequenced by primer walking. It was demonstrated that the incidence of the qnrA-positive strains in nalidixic acid-resistant bacteria was 1.9% (12/629), in which the detection rates for Klebiesiella pneumoniae. Enterobacter cloacae, Enterobacter aerogenes,Citrobacter freundii and Salmonella choeraesuis were 2.2% (3/138), 17. 1% (6/35), 9. 1%(1/11), 12.5% (1/8), and 14.3% (1/7), respectively. The qnrA gene was found to be embedded in the complex su/1-type integron located on plasmids with varied size (80-180 kb). Among them, 4qnrA-positive isolates carried integron In37 and 8 isolates carried a novel integron, temporarily designated as InX. All the qnrA-positive isolates were ESBL-producing and transferable for the multi-drug resistance. It is concluded that the plasmid-mediated drug-resistance mechanism exists in the quinolone resistant strains of isolates from hospitals in Guangdong area, but the incidence was rather low. Nevertheless, it is still possible that the horizontal transfer of the resistant qnrA gene might lead to the spreading of drug-resistance.