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TolC is a major outer membrane channel and it plays an important role in the excretion of a wide range of molecules, including antibiotics. A recent study has shown that Salmonella enterica serovar Typhimurium has nine functional drug efflux pumps; however, the TolC dependency of these efflux pumps remains to be studied in detail. The aim of this study was to investigate the TolC dependency of multidrug efflux pumps in this organism.All genes encoding the drug efflux systems were cloned into the pUC118 vector. Constructed plasmids were transformed into ΔacrB and ΔtolC mutants of S. enterica serovar Typhimurium ATCC 14028s, and then the drug susceptibilities of these transformants were determined.Plasmids carrying the acrAB, acrD, acrEF, mdsAB, mdtABC, emrAB or macAB genes did not confer resistance to the tolC mutant, whereas they conferred drug resistance to the acrB mutant. Only three plasmids carrying mdsABC, mdfA or mdtK conferred resistance to the tolC mutant.TolC is required for the function of seven drug efflux systems (AcrAB, AcrD, AcrEF, MdsAB, MdtABC, EmrAB and MacAB) in S. enterica serovar Typhimurium.