| Summary: | <p>Abstract</p> <p>Background</p> <p><it>Yersinia pestis </it>is the causative agent of plague and a potential agent of bioterrorism and biowarfare. The plague biothreat and the emergence of multidrug-resistant plague underscore the need to increase our understanding of the intrinsic potential of <it>Y. pestis </it>for developing antimicrobial resistance and to anticipate the mechanisms of resistance that may emerge in <it>Y. pestis</it>. Identification of <it>Y. pestis </it>genes that, when overexpressed, are capable of reducing antibiotic susceptibility is a useful strategy to expose genes that this pathogen may rely upon to evolve antibiotic resistance via a vertical modality. In this study, we explored the use of a multicopy suppressor, <it>Escherichia coli </it>host-based screening approach as a means to expose antibiotic resistance determinant candidates in <it>Y. pestis</it>.</p> <p>Results</p> <p>We constructed a multicopy plasmid-based, <it>Y. pestis </it>genome-wide expression library of nearly 16,000 clones in <it>E. coli </it>and screened the library for suppressors of the antimicrobial activity of ofloxacin, a fluoroquinolone antibiotic. The screen permitted the identification of a transcriptional regulator-encoding gene (<it>robA</it><sub>Yp</sub>) that increased the MIC<sub>99 </sub>of ofloxacin by 23-fold when overexpressed from a multicopy plasmid in <it>Y. pestis</it>. Additionally, we found that <it>robA</it><sub>Yp </sub>overexpression in <it>Y. pestis </it>conferred low-level resistance to many other antibiotics and increased organic solvent tolerance. Overexpression of <it>robA</it><sub>Yp </sub>also upregulated the expression of several efflux pumps in <it>Y. pestis</it>.</p> <p>Conclusion</p> <p>Our study provides proof of principle for the use of multicopy suppressor screening based on the tractable and easy-to-manipulate <it>E. coli </it>host as a means to identify antibiotic resistance determinant candidates of <it>Y. pestis</it>.</p>
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