Mechanisms of Increased Resistance to Chlorhexidine and Cross-Resistance to Colistin following Exposure of Klebsiella pneumoniaeClinical Isolates to Chlorhexidine

ABSTRACTKlebsiella pneumoniaeis an opportunistic pathogen that is often difficult to treat due to its multidrug resistance (MDR). We have previously shown that K. pneumoniaestrains are able to “adapt” (become more resistant) to the widely used bisbiguanide antiseptic chlorhexidine. Here, we investigated the mechanisms responsible for and the phenotypic consequences of chlorhexidine adaptation, with particular reference to antibiotic cross-resistance. In five of six strains, adaptation to chlorhexidine also led to resistance to the last-resort antibiotic colistin. Here, we show that chlorhexidine adaptation is associated with mutations in the two-component regulator phoPQand a putative Tet repressor gene (smvR) adjacent to the major facilitator superfamily (MFS) efflux pump gene, smvA. Upregulation of smvA(10- to 27-fold) was confirmed in smvRmutant strains, and this effect and the associated phenotype were suppressed when a wild-type copy of smvRwas introduced on plasmid pACYC. Upregulation of phoPQ(5- to 15-fold) and phoPQ-regulated genes, pmrD(6- to 19-fold) and pmrK(18- to 64-fold), was confirmed in phoPQmutant strains. In contrast, adaptation of K. pneumoniaeto colistin did not result in increased chlorhexidine resistance despite the presence of mutations in phoQand elevated phoPQ, pmrD, and pmrKtranscript levels. Insertion of a plasmid containing phoPQfrom chlorhexidine-adapted strains into wild-type K. pneumoniaeresulted in elevated expression levels of phoPQ, pmrD, and pmrKand increased resistance to colistin, but not chlorhexidine. The potential risk of colistin resistance emerging in K. pneumoniaeas a consequence of exposure to chlorhexidine has important clinical implications for infection prevention procedures.