Search

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.

ABSTRACTThe opportunistic pathogen Acinetobacter baumanniiis able to persist in the environment and is often multidrug resistant (MDR), causing difficulties in the treatment of infections. Here, we show that the two-component system AdeRS, which regulates the production of the AdeABC multidrug resistance efflux pump, is required for the formation of a protective biofilm in an ex vivoporcine mucosal model, which mimics a natural infection of the human epithelium. Interestingly, deletion of adeBimpacted only on the ability of strain AYE to form a biofilm on plastic and only on the virulence of strain Singapore 1 for Galleria mellonella. RNA-Seq revealed that loss of AdeRS or AdeB significantly altered the transcriptional landscape, resulting in the changed expression of many genes, notably those associated with antimicrobial resistance and virulence interactions. For example, A. baumanniilacking AdeRS displayed decreased expression of adeABC, pilgenes, comgenes, and a pgaC-like gene, whereas loss of AdeB resulted in increased expression of piland comgenes and decreased expression of ferric acinetobactin transport system genes. These data define the scope of AdeRS-mediated regulation, show that changes in the production of AdeABC mediate important phenotypes controlled by AdeRS, and suggest that AdeABC is a viable target for antimicrobial drug and antibiofilm discovery.IMPORTANCEAcinetobacter baumanniiis a nosocomial pathogen and is an increasing problem in hospitals worldwide. This organism is often multidrug resistant, can persist in the environment, and forms a biofilm on environmental surfaces and wounds. Overproduction of efflux pumps can allow specific toxic compounds to be pumped out of the cell and can lead to multidrug resistance. This study demonstrates the role of the A. baumanniiefflux pump AdeB, and its regulator AdeRS, in multidrug resistance, epithelial cell killing, and biofilm formation. Deletion of the genes encoding these systems led to increased susceptibility to antibiotics, decreased biofilm formation on biotic and abiotic surfaces, and decreased virulence. Our data suggest that inhibition of AdeB could prevent biofilm formation or colonization in patients by A. baumanniiand provides a good target for drug discovery.