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The Pseudomonas aeruginosa genome contains several different multidrug resistance (MDR) efflux pumps. Overproduction of these pumps reduces susceptibility to a variety of antibiotics. Some recently published works have analyzed the effect of the overproduction of MDR efflux pumps on bacterial virulence. Here we have studied the effect of overproduction of the efflux pumps MexAB-OprM, MexCD-OprJ, MexEF-OprN, and MexXY on type III secretion (T3S) in P. aeruginosa. The type III secretion system (T3SS) is used by P. aeruginosa to deliver toxins directly into the cytoplasm of the host cell. Our data indicate that overexpression of either MexCD-OprJ or MexEF-OprN is associated with the impairment of T3S in P. aeruginosa. No effect on overexpression of either MexAB-OprM or MexXY was detected. The observed defect in T3S was due to a lack of expression of genes belonging to the T3SS regulon …