In Vivo Emergence of Colistin Resistance in Klebsiella pneumoniae Producing KPC-Type Carbapenemases Mediated by Insertional Inactivation of the PhoQ/PhoP mgrB Regulator

Abstract

e Colistin is one of the few agents that retain activity against extensively drug-resistant strains of Klebsiella pneumoniae producing KPC-type carbapenemases (KPC-KP). However, resistance to colistin 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 isolate (KKBO-1) and a colistin-resistant isolate (KKBO-4) 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 transcription 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.

Tracking Nosocomial Klebsiella pneumoniae Infections and Outbreaks by Whole-Genome Analysis: Small-Scale Italian Scenario within a Single Hospital

“…Genomic analysis aimed at detecting the determinants for this resistance trait found that the 10 resistant isolates demonstrated insertion of an IS5-like transposon in the mgrB, a gene that regulates a pathway of lipopolysaccharide biosynthesis (43). Insertion of an IS5-like sequence in the mgrB gene has indeed been proposed as a determinant of colistin resistance (32). Our results appear to support this causative link, considering that none of the six colistin-sensitive strains presented the aforementioned insertion.…”

“…All colistin-resistant strains harbored a variant of the mgrB gene interrupted by an IS5-like transposon ( Table 3). Insertion of a transposon in this gene has been reported as a determinant of colistin resistance (32).…”

Colistin Resistance in Carbapenem-Resistant Klebsiella Pneumoniae Strains

“…However, excessive use of colistin has led to resistance to this drug [13]. The resistance to colistin seen in K. pneumoniae strains is reported to be due to reduced affinity of colistin to lipopolysaccharide target [5]. In this study, colistin resistance in CRKP strains has been identified and evaluated.…”

“…As a result of increased use of carbapenems, carbapenemases are widespread in Enterobacteriaceae family and particularly in Klebsiella pneumoniae; so effective treatment options are decreasing [2,4]. Carbapenemase-producing K. pneumoniae emerged in the late 1990s and has become a serious health problem worldwide [5].…”

Large Nosocomial Outbreak of Colistin-Resistant, Carbapenemase-Producing Klebsiella pneumoniae Traced to Clonal Expansion of an mgrB Deletion Mutant

“…Inactivation of the mgrB gene, which encodes a negative feedback regulator of the PhoQ/PhoP signal transduction system, was found to be one of the most common mutational mechanisms responsible for polymyxin resistance among clinical isolates of CRKP (12)(13)(14). Alterations of the PmrA/PmrB and other twocomponent signal transduction systems have also been identified as causes of polymyxin resistance in K. pneumoniae (15)(16)(17)(18)(19).…”