MarA-mediated overexpression of the AcrAB efflux pump results in decreased susceptibility to tigecycline in Escherichia coli

Objectives: The purpose of this study was to characterize decreased susceptibility to tigecycline in clinical isolates of Escherichia coli obtained during Phase 3 clinical trials. Methods: Gene expression was analysed by transcriptional profile analysis and RT-PCR. Transposon mutagenesis with IS903 phi kan was used for selection of transposon mutants. Transposon insertions were mapped by DNA sequencing and PCR analyses. The MICs were determined by broth microdilution. Results: Both transcriptional profile analysis and Taqman RT-PCR demonstrated increased expression levels of MarA, a transcriptional activator, and AcrAB, an RND-type efflux pump, in the strains with elevated tigecycline MICs. Transposon mutagenesis generated nine mutants, the majority of which had either marA or acrB inactivated. Sequence analysis revealed a single nucleotide insertion in the open reading frame of the marR gene in less-susceptible strains of E. coli. Conclusions: This study suggested that a loss of MarR functionality due to a frameshift mutation resulted in constitutive overproduction of MarA and AcrAB and, consequently, in decreased susceptibility to tigecycline in clinical isolates of E. coli.