Background/aims: It has been shown that quinolone resistance arises due to mutations in the quinolone resistance-determining regions of the drug targets. This study aimed to optimise a multiplex PCR assay to track plasmid-mediated low-level quinolone resistance profiles.Subjects and methods: A multiplex PCR-based-method in which the primers were already established by our team. About 44 samples were collected from 44 patients who enrolled in this study by using surgical site infection (SSI).Results: By targeting the conserved domains of qnrA, qnrB, qnrS and the qnrVC gene families, the primer number was reduced significantly to only four pairs in one multiplex PCR. Using multiplex PCR, 3/44 SSI samples were found to be carrying fluoroquinolone-resistance genes (qnrA, qnrB, qnrS, qnrVC).Conclusion: A multiplex PCR for detecting pathogens as well as identifying quinolone resistance genes all in one reaction was successfully established.