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. 1991 Sep;173(18):5854–5860. doi: 10.1128/jb.173.18.5854-5860.1991

A novel locus conferring fluoroquinolone resistance in Staphylococcus aureus.

M Trucksis 1, J S Wolfson 1, D C Hooper 1
PMCID: PMC208319  PMID: 1653224

Abstract

Fluoroquinolones such as ciprofloxacin and ofloxacin are potent antimicrobial agents that antagonize the A subunit of DNA gyrase. We selected and mapped a novel fluoroquinolone resistance gene on the Staphylococcus aureus chromosome. Resistant mutants were selected with ciprofloxacin or ofloxacin and were uniformly localized to the A fragment of chromosomal DNA digested with SmaI and arrayed by pulsed-field gel electrophoresis. Several mutants (cfxB, ofxC) were genetically mapped between the thr and trp loci in the A fragment. A majority of A fragment fluoroquinolone resistance mutations were associated with reduced susceptibility to novobiocin, an antagonist of the B subunit of DNA gyrase. Two genes previously associated with fluoroquinolone resistance, the gyrA gene of DNA gyrase and the norA gene (associated with decreased drug accumulation), were localized to the G and D fragments, respectively. Thus, the fluoroquinolone resistance mutations in the A fragment are distinct from previously identified fluoroquinolone resistance mutations in gyrA and norA. Whether mutations in the A fragment after a second topoisomerase or another gene controlling supercoiling or affect drug permeation is unknown.

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Selected References

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  1. Bauer A. W., Kirby W. M., Sherris J. C., Turck M. Antibiotic susceptibility testing by a standardized single disk method. Am J Clin Pathol. 1966 Apr;45(4):493–496. [PubMed] [Google Scholar]
  2. Feinberg A. P., Vogelstein B. "A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity". Addendum. Anal Biochem. 1984 Feb;137(1):266–267. doi: 10.1016/0003-2697(84)90381-6. [DOI] [PubMed] [Google Scholar]
  3. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  4. Gellert M. DNA topoisomerases. Annu Rev Biochem. 1981;50:879–910. doi: 10.1146/annurev.bi.50.070181.004311. [DOI] [PubMed] [Google Scholar]
  5. Gellert M., Mizuuchi K., O'Dea M. H., Itoh T., Tomizawa J. I. Nalidixic acid resistance: a second genetic character involved in DNA gyrase activity. Proc Natl Acad Sci U S A. 1977 Nov;74(11):4772–4776. doi: 10.1073/pnas.74.11.4772. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Gellert M., Mizuuchi K., O'Dea M. H., Nash H. A. DNA gyrase: an enzyme that introduces superhelical turns into DNA. Proc Natl Acad Sci U S A. 1976 Nov;73(11):3872–3876. doi: 10.1073/pnas.73.11.3872. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Higgins C. F., Dorman C. J., Stirling D. A., Waddell L., Booth I. R., May G., Bremer E. A physiological role for DNA supercoiling in the osmotic regulation of gene expression in S. typhimurium and E. coli. Cell. 1988 Feb 26;52(4):569–584. doi: 10.1016/0092-8674(88)90470-9. [DOI] [PubMed] [Google Scholar]
  8. Hirai K., Aoyama H., Suzue S., Irikura T., Iyobe S., Mitsuhashi S. Isolation and characterization of norfloxacin-resistant mutants of Escherichia coli K-12. Antimicrob Agents Chemother. 1986 Aug;30(2):248–253. doi: 10.1128/aac.30.2.248. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hooper D. C., Wolfson J. S., Souza K. S., Tung C., McHugh G. L., Swartz M. N. Genetic and biochemical characterization of norfloxacin resistance in Escherichia coli. Antimicrob Agents Chemother. 1986 Apr;29(4):639–644. doi: 10.1128/aac.29.4.639. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hopewell R., Oram M., Briesewitz R., Fisher L. M. DNA cloning and organization of the Staphylococcus aureus gyrA and gyrB genes: close homology among gyrase proteins and implications for 4-quinolone action and resistance. J Bacteriol. 1990 Jun;172(6):3481–3484. doi: 10.1128/jb.172.6.3481-3484.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kato J., Nishimura Y., Imamura R., Niki H., Hiraga S., Suzuki H. New topoisomerase essential for chromosome segregation in E. coli. Cell. 1990 Oct 19;63(2):393–404. doi: 10.1016/0092-8674(90)90172-b. [DOI] [PubMed] [Google Scholar]
  12. Kuhl S. A., Pattee P. A., Baldwin J. N. Chromosomal map location of the methicillin resistance determinant in Staphylococcus aureus. J Bacteriol. 1978 Aug;135(2):460–465. doi: 10.1128/jb.135.2.460-465.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lampe M. F., Bott K. F. Cloning the gyrA gene of Bacillus subtilis. Nucleic Acids Res. 1984 Aug 10;12(15):6307–6323. doi: 10.1093/nar/12.15.6307. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lampe M. F., Bott K. F. Genetic and physical organization of the cloned gyrA and gyrB genes of Bacillus subtilis. J Bacteriol. 1985 Apr;162(1):78–84. doi: 10.1128/jb.162.1.78-84.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lindberg M., Sjöström J. E., Johansson T. Transformation of chromosomal and plasmid characters in Staphylococcus aureus. J Bacteriol. 1972 Feb;109(2):844–847. doi: 10.1128/jb.109.2.844-847.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Liu L. F., Wang J. C. Micrococcus luteus DNA gyrase: active components and a model for its supercoiling of DNA. Proc Natl Acad Sci U S A. 1978 May;75(5):2098–2102. doi: 10.1073/pnas.75.5.2098. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Luchansky J. B., Pattee P. A. Isolation of transposon Tn551 insertions near chromosomal markers of interest in Staphylococcus aureus. J Bacteriol. 1984 Sep;159(3):894–899. doi: 10.1128/jb.159.3.894-899.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Matsuhashi M., Song M. D., Ishino F., Wachi M., Doi M., Inoue M., Ubukata K., Yamashita N., Konno M. Molecular cloning of the gene of a penicillin-binding protein supposed to cause high resistance to beta-lactam antibiotics in Staphylococcus aureus. J Bacteriol. 1986 Sep;167(3):975–980. doi: 10.1128/jb.167.3.975-980.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Ohshita Y., Hiramatsu K., Yokota T. A point mutation in norA gene is responsible for quinolone resistance in Staphylococcus aureus. Biochem Biophys Res Commun. 1990 Nov 15;172(3):1028–1034. doi: 10.1016/0006-291x(90)91549-8. [DOI] [PubMed] [Google Scholar]
  20. Patel A. H., Foster T. J., Pattee P. A. Physical and genetic mapping of the protein A gene in the chromosome of Staphylococcus aureus 8325-4. J Gen Microbiol. 1989 Jul;135(7):1799–1807. doi: 10.1099/00221287-135-7-1799. [DOI] [PubMed] [Google Scholar]
  21. Pattee P. A. Distribution of Tn551 insertion sites responsible for auxotrophy on the Staphylococcus aureus chromosome. J Bacteriol. 1981 Jan;145(1):479–488. doi: 10.1128/jb.145.1.479-488.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Pattee P. A., Neveln D. S. Transformation analysis of three linkage groups in Staphylococcus aureus. J Bacteriol. 1975 Oct;124(1):201–211. doi: 10.1128/jb.124.1.201-211.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Raji A., Zabel D. J., Laufer C. S., Depew R. E. Genetic analysis of mutations that compensate for loss of Escherichia coli DNA topoisomerase I. J Bacteriol. 1985 Jun;162(3):1173–1179. doi: 10.1128/jb.162.3.1173-1179.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Rudd K. E., Menzel R. his operons of Escherichia coli and Salmonella typhimurium are regulated by DNA supercoiling. Proc Natl Acad Sci U S A. 1987 Jan;84(2):517–521. doi: 10.1073/pnas.84.2.517. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  26. Sreedharan S., Oram M., Jensen B., Peterson L. R., Fisher L. M. DNA gyrase gyrA mutations in ciprofloxacin-resistant strains of Staphylococcus aureus: close similarity with quinolone resistance mutations in Escherichia coli. J Bacteriol. 1990 Dec;172(12):7260–7262. doi: 10.1128/jb.172.12.7260-7262.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Stahl M. L., Pattee P. A. Confirmation of protoplast fusion-derived linkages in Staphylococcus aureus by transformation with protoplast DNA. J Bacteriol. 1983 Apr;154(1):406–412. doi: 10.1128/jb.154.1.406-412.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Sugino A., Bott K. F. Bacillus subtilis deoxyribonucleic acid gyrase. J Bacteriol. 1980 Mar;141(3):1331–1339. doi: 10.1128/jb.141.3.1331-1339.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sugino A., Peebles C. L., Kreuzer K. N., Cozzarelli N. R. Mechanism of action of nalidixic acid: purification of Escherichia coli nalA gene product and its relationship to DNA gyrase and a novel nicking-closing enzyme. Proc Natl Acad Sci U S A. 1977 Nov;74(11):4767–4771. doi: 10.1073/pnas.74.11.4767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Takahata M., Nishino T. DNA gyrase of Staphylococcus aureus and inhibitory effect of quinolones on its activity. Antimicrob Agents Chemother. 1988 Aug;32(8):1192–1195. doi: 10.1128/aac.32.8.1192. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Trucksis M., Depew R. E. Identification and localization of a gene that specifies production of Escherichia coli DNA topoisomerase I. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2164–2168. doi: 10.1073/pnas.78.4.2164. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Trucksis M., Hooper D. C., Wolfson J. S. Emerging resistance to fluoroquinolones in staphylococci: an alert. Ann Intern Med. 1991 Mar 1;114(5):424–426. doi: 10.7326/0003-4819-114-5-424. [DOI] [PubMed] [Google Scholar]
  33. Ubukata K., Itoh-Yamashita N., Konno M. Cloning and expression of the norA gene for fluoroquinolone resistance in Staphylococcus aureus. Antimicrob Agents Chemother. 1989 Sep;33(9):1535–1539. doi: 10.1128/aac.33.9.1535. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Yoshida H., Bogaki M., Nakamura S., Ubukata K., Konno M. Nucleotide sequence and characterization of the Staphylococcus aureus norA gene, which confers resistance to quinolones. J Bacteriol. 1990 Dec;172(12):6942–6949. doi: 10.1128/jb.172.12.6942-6949.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]

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