title: Multidrug resistance in Klebsiella pneumoniae: a novel gene, ramA, confers a multidrug resistance phenotype in Escherichia coli.
abstract: Spontaneous multidrug-resistant (Mdr) mutants of Klebsiella pneumoniae strain ECL8 arose at a frequency of 2.2 x 10(-8) and showed increased resistance to a range of unrelated antibiotics, including chloramphenicol, tetracycline, nalidixic acid, ampicillin, norfloxacin, trimethoprim and puromycin. A chromosomal fragment from one such mutant was cloned, and found to confer an Mdr phenotype on Escherichia coli K12 cells that was essentially identical to that of the K. pneumoniae mutant. Almost complete loss of the OmpF porin in the E. coli transformant, and of the corresponding porin in the K. pneumoniae mutant, was observed. The presence of the Mdr mutation in K. pneumoniae or the cloned K. pneumoniae ramA (resistance antibiotic multiple) locus in E. coli also resulted in active efflux of tetracycline, and increased active efflux of chloramphenicol. After transformation of a ramA plasmid into E. coli, expression of chloramphenicol resistance occurred later than expression of resistance to tetracycline, puromycin, trimethoprim and nalidixic acid. The ramA gene was localized and sequenced. It encodes a putative positive transcriptional activator that is weakly related to the E. coli MarA and SoxS proteins. A ramA gene was also found to be present in an Enterobacter cloacae fragment that has previously been shown to confer an Mdr phenotype, and it appears that ramA, rather than the romA gene identified in that study, is responsible for multidrug resistance. The ramA gene from the wild-type K. pneumoniae was identical to that of the mutant strain and also conferred an Mdr phenotype on E. coli, indicating that the mutation responsible for Mdr in K. pneumoniae had not been cloned.
authors.affiliation: Department of Biochemistry and Physiology, University of Technology Sydney, PO Box 123, Broadway NSW 2007, Australia.
authors.forename: Anthony M
authors.initials: AM
authors.lastname: George
authors.affiliation: CSIRO Division of Biomolecular Engineering, Laboratory for Molecular Biology, PO Box 184, North Ryde NSW 2113, Australia.
authors.forename: Ruth M
authors.initials: RM
authors.lastname: Hall
authors.affiliation: School of Biological Sciences, Macquarie University, Sydney NSW 2109, Australia.
authors.forename: H W
authors.initials: HW
authors.lastname: Stokes
mesh_terms: D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D001693:Biological Transport, Active; D002701:Chloramphenicol / Q000378:metabolism / Q000494:pharmacology; D003001:Cloning, Molecular; D018432:Drug Resistance, Multiple / Q000235:genetics*; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D005798:Genes, Bacterial; D007711:Klebsiella pneumoniae / Q000235:genetics* / Q000378:metabolism; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D015183:Restriction Mapping; D016415:Sequence Alignment; D013752:Tetracycline / Q000378:metabolism; D013997:Time Factors; D014157:Transcription Factors / Q000235:genetics
chemical_list: D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D008565:Membrane Proteins; C095947:RamA protein, Klebsiella; D014157:Transcription Factors; C064777:romA protein, Enterobacter cloacae; D002701:Chloramphenicol; D013752:Tetracycline