title: Unstable tandem gene amplification generates heteroresistance (variation in resistance within a population) to colistin in Salmonella enterica.
abstract: Heteroresistance, a phenomenon where subpopulations of a bacterial isolate exhibit different susceptibilities to an antibiotic, is a growing clinical problem where the underlying genetic mechanisms in most cases remain unknown. We isolated colistin resistant mutants in Escherichia coli and Salmonella enterica serovar Typhimurium at different concentrations of colistin. Genetic analysis showed that genetically stable pmrAB point mutations were responsible for colistin resistance during selection at high drug concentrations for both species and at low concentrations for E. coli. In contrast, for S. Typhimurium mutants selected at low colistin concentrations, amplification of different large chromosomal regions conferred a heteroresistant phenotype. All amplifications included the pmrD gene, which encodes a positive regulator that up-regulates proteins that modify lipid A, and as a result increase colistin resistance. Inactivation and over-expression of the pmrD gene prevented and conferred resistance, respectively, demonstrating that the PmrD protein is required and sufficient to confer resistance. The heteroresistance phenotype is explained by the variable gene dosage of pmrD in a population, where sub-populations with different copy number of the pmrD gene show different levels of colistin resistance. We propose that variability in gene copy number of resistance genes can explain the heteroresistance observed in clinically isolated pathogenic bacteria.
authors.affiliation: Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden.
authors.forename: Karin
authors.initials: K
authors.lastname: Hjort
authors.affiliation: Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden.
authors.forename: Hervé
authors.initials: H
authors.lastname: Nicoloff
authors.affiliation: Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden. dan.andersson@imbim.uu.se.
authors.forename: Dan I
authors.initials: DI
authors.lastname: Andersson
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics; D003091:Colistin / Q000494:pharmacology*; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005784:Gene Amplification; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D008826:Microbial Sensitivity Tests; D009154:Mutation; D012486:Salmonella typhimurium / Q000187:drug effects* / Q000235:genetics* / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; C409039:PmrB protein, bacteria; D014157:Transcription Factors; C081902:pmrA protein, Bacteria; D003091:Colistin