RamA upregulates multidrug resistance efflux pumps AcrAB and OqxAB in Klebsiella pneumoniae

Abstract

Overexpression of the acrAB genes regulated by RamA and overexpression of oqxAB regulated by RarA have been reported to mediate multidrug resistance in Gram-negative bacilli. In this study, regulation of acrAB and oqxAB simultaneously by the global regulator RamA was investigated in a multidrug-resistant Klebsiella pneumoniae clinical isolate (KP22) resistant to tigecycline and other antimicrobials. KP22 overexpressed ramA due to a ramR mutation, along with an unexpected overexpression of oqxB. Deletion of ramA led to a 16-fold decrease in the tigecycline minimum inhibitory concentration (MIC) with decreased expression of acrB (4.3-fold) and oqxB (7.1-fold) compared with KP22. Transcomplementation of KP22ΔramA with the wild-type ramA gene restored the tigecycline MIC and upregulation of the acrB (3.9-fold) and oqxB (4.0-fold) genes compared with KP22. When oqxB was knocked out, MICs of ciprofloxacin, olaquindox and nitrofurantoin were considerably decreased, while deletion of acrB led to MIC decreases for cefepime, piperacillin/tazobactam and tigecycline in addition to the above three antimicrobials. The results of electrophoretic mobility shift assay showed that RamA could bind the promoter regions of both the acrAB and oqxAB operons. This study demonstrates for the first time that RamA can directly regulate multidrug resistance efflux pumps AcrAB and OqxAB in K. pneumoniae.