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Loss-of-function mutations in the Dpp and Opp permeases render Erwinia amylovora resistant to kasugamycin and blasticidin S

Yixin Ge: University of Illinois

<div>Extensive use of antibiotic streptomycin to control fire blight disease of apples and pears, caused by <em>Erwinia amylovora</em>, leads to the development of streptomycin-resistant strains in the United States and elsewhere. Kasugamycin (Ksg) has been considered as an alternative to control this disease. In this study, we investigated the role of two major peptide ABC-transporter systems in <em>E. amylovora</em>, the dipeptide permease (Dpp) and oligopeptide permease (Opp), in conferring sensitivity to Ksg and blasticidin S (BcS). Minimum inhibitory concentration and spot dilution assays showed that the <em>dpp </em>and <em>opp </em>deletion mutants exhibited enhanced resistance to Ksg and BcS. Deletion of both <em>dpp</em> and <em>opp</em> conferred higher level of resistance to Ksg as compared to the single mutant. In addition, bioinformatic analysis combined with qRT-PCR showed that the Rcs system negatively regulates <em>opp</em> expression and the <em>rcsB </em>mutant was more sensitive to both Ksg and BcS as compared to the wild-type. Electrophoresis motility shift assay further confirmed the direct binding of the RcsA/RcsB proteins to the promoter region of the <em>opp </em>operon. However, neither the Dpp nor the Opp permeases contributed to disease progress on immature pears, hypersensitive response on tobacco leaves, motility, and amylovoran production. These results suggested that Ksg and BcS hijack the Dpp and Opp permeases to enter <em>E. amylovora</em> cells, and the Dpp and Opp permeases act synergistically for illicit transport of antibiotics.</div>