The ABC of biofilm drug tolerance: The MerR-like regulator BrlR is an activator of ABC transport systems, with PA1874-77 contributing to the tolerance by Pseudomonas aeruginosa biofilms to tobramycin [PublishAheadOfPrint]

A hallmark of biofilms is their tolerance to killing by antimicrobial agents. In P. aeruginosa, biofilm drug tolerance requires the c-di-GMP responsive, MerR transcriptional regulator BrlR. However, the mechanism by which BrlR mediates biofilm drug tolerance has not been elucidated. Here, we demonstrate that BrlR activates the expression of at least 7 ABC transport systems, including PA1874-77, with ChIP and DNA binding assays confirming BrlR binding to the promoter region of PA1874-77. Insertional inactivation of the 7 ABC transport systems rendered P. aeruginosa PAO1 biofilms susceptible to tobramycin or norfloxacin. Susceptibility was linked to drug accumulation, with BrlR contributing to norfloxacin accumulation in a manner dependent on multidrug efflux pumps and the ABC transport system PA1874-77. Inactivation of the respective ABC transport system furthermore eliminated the recalcitrance of biofilms to killing by tobramycin but not norfloxacin, indicating that drug accumulation is not linked to biofilm drug tolerance. Our findings indicate for the first time that BrlR, a MerR-type transcriptional activator, activates genes encoding several ABC transport systems, in addition to multiple multi-drug efflux pump genes. Moreover, our data confirm a BrlR target contributing to drug tolerance, likely countering the prevailing dogma that biofilm tolerance arises from a multiplicity of factors.

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