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Antimicrob Agents Chemother. In vivo emergence of colistin resistance in Klebsiella pneumoniae producing KPC-type carbapenemase mediated by insertional inactivation of the PhoQ/PhoP mgrB regulator
[Source: Antimicrobial Agents and Chemotherapy, full page: (LINK). Abstract, edited.]
In vivo emergence of colistin resistance in Klebsiella pneumoniae producing KPC-type carbapenemase mediated by insertional inactivation of the PhoQ/PhoP mgrB regulator
Antonio Cannatelli 1, Marco Maria D?Andrea 1, Tommaso Giani 1, Vincenzo Di Pilato 1, Fabio Arena 1, Simone Ambretti 2, Paolo Gaibani 3 and Gian Maria Rossolini 1,4,5*
Author Affiliations: <SUP>1</SUP> Department of Medical Biotechnologies, University of Siena, Siena, Italy <SUP>2</SUP> Bacteriology Laboratory <SUP>3</SUP> Regional Laboratory for Microbiological Emergencies, Unit of Clinical Microbiology, St. Orsola University Hospital, Bologna, Italy <SUP>4</SUP> Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy <SUP>5</SUP> Clinical Microbiology and Virology Unit, Florence Careggi University Hospital, Florence, Italy
ABSTRACT
Colistin is one of the few agents which retains activity against extensively drug resistant strains of Klebsiella pneumoniae producing KPC-type carbapenemases (KPC-KP). However, colistin resistance is increasingly reported among KPC-KP. Comparative genomic analysis of a pair of sequential KPC-KP isolates from the same patient, including a colistin-susceptible (KKBO-1) and a colistin-resistant (KKBO-4) isolate selected after colistin exposure, revealed that insertional inactivation of the mgrB gene, encoding a negative regulator of the PhoQ/PhoP signaling system, is a genetic mechanism for acquired colistin-resistance. The role of mgrB inactivation in acquired colistin resistance was confirmed by complementation experiments with wild-type mgrB, which restored colistin susceptibility in KKBO-4, and by construction of an mgrB deletion mutant from KKBO-1, which exhibited a colistin-resistant phenotype. Insertional mgrB inactivation was also detected in 60% of colistin-resistant mutants selected from KKBO-1 in vitro, following plating on colistin-containing medium, confirming the role (although not unique) of this mechanism in the emergence of acquired colistin resistance. In colistin-resistant mutants carrying insertional inactivation or deletion of the mgrB gene, upregulated transciption of phoP, phoQ and pmrK (which is part of the pmrHFIJKLM operon) was detected. These findings confirmed the MgrB regulatory role in K. pneumoniae, and were in agreement with the known association between upregulation of the PhoQ/PhoP system and activation of the pmrHFIJKLM operon which eventually leads to resistance to polymyxins by modification of the lipopolysaccharide target.
FOOTNOTES
*Corresponding author. Mailing address: Department of Experimental and Clinical Medicine, University of Florence, 50139 - Florence, Italy. Phone: +39-055-7949239; fax: +39-055-7949289; E-mail: gianni.rossolini@gmail.com, gianmaria.rossolini@unisi.it
Copyright ? 2013, American Society for Microbiology. All Rights Reserved.
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In vivo emergence of colistin resistance in Klebsiella pneumoniae producing KPC-type carbapenemase mediated by insertional inactivation of the PhoQ/PhoP mgrB regulator
Antonio Cannatelli 1, Marco Maria D?Andrea 1, Tommaso Giani 1, Vincenzo Di Pilato 1, Fabio Arena 1, Simone Ambretti 2, Paolo Gaibani 3 and Gian Maria Rossolini 1,4,5*
Author Affiliations: <SUP>1</SUP> Department of Medical Biotechnologies, University of Siena, Siena, Italy <SUP>2</SUP> Bacteriology Laboratory <SUP>3</SUP> Regional Laboratory for Microbiological Emergencies, Unit of Clinical Microbiology, St. Orsola University Hospital, Bologna, Italy <SUP>4</SUP> Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy <SUP>5</SUP> Clinical Microbiology and Virology Unit, Florence Careggi University Hospital, Florence, Italy
ABSTRACT
Colistin is one of the few agents which retains activity against extensively drug resistant strains of Klebsiella pneumoniae producing KPC-type carbapenemases (KPC-KP). However, colistin resistance is increasingly reported among KPC-KP. Comparative genomic analysis of a pair of sequential KPC-KP isolates from the same patient, including a colistin-susceptible (KKBO-1) and a colistin-resistant (KKBO-4) isolate selected after colistin exposure, revealed that insertional inactivation of the mgrB gene, encoding a negative regulator of the PhoQ/PhoP signaling system, is a genetic mechanism for acquired colistin-resistance. The role of mgrB inactivation in acquired colistin resistance was confirmed by complementation experiments with wild-type mgrB, which restored colistin susceptibility in KKBO-4, and by construction of an mgrB deletion mutant from KKBO-1, which exhibited a colistin-resistant phenotype. Insertional mgrB inactivation was also detected in 60% of colistin-resistant mutants selected from KKBO-1 in vitro, following plating on colistin-containing medium, confirming the role (although not unique) of this mechanism in the emergence of acquired colistin resistance. In colistin-resistant mutants carrying insertional inactivation or deletion of the mgrB gene, upregulated transciption of phoP, phoQ and pmrK (which is part of the pmrHFIJKLM operon) was detected. These findings confirmed the MgrB regulatory role in K. pneumoniae, and were in agreement with the known association between upregulation of the PhoQ/PhoP system and activation of the pmrHFIJKLM operon which eventually leads to resistance to polymyxins by modification of the lipopolysaccharide target.
FOOTNOTES
*Corresponding author. Mailing address: Department of Experimental and Clinical Medicine, University of Florence, 50139 - Florence, Italy. Phone: +39-055-7949239; fax: +39-055-7949289; E-mail: gianni.rossolini@gmail.com, gianmaria.rossolini@unisi.it
Copyright ? 2013, American Society for Microbiology. All Rights Reserved.
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