Skip to main content
PMC home page
Antimicrobial Agents and Chemotherapy logoLink to Antimicrobial Agents and Chemotherapy
. 1996 May;40(5):1157–1163. doi: 10.1128/aac.40.5.1157

Alterations in the DNA topoisomerase IV grlA gene responsible for quinolone resistance in Staphylococcus aureus.

J Yamagishi 1, T Kojima 1, Y Oyamada 1, K Fujimoto 1, H Hattori 1, S Nakamura 1, M Inoue 1
PMCID: PMC163283  PMID: 8723458

Abstract

A 4.2-kb DNA fragment conferring quinolone resistance was cloned from a quinolone-resistant clinical isolate of Staphylococcus aureus and was shown to possess a part of the grlB gene and a mutated grlA gene. S-80-->F and E-84-->K mutations in the grlA gene product were responsible for the quinolone resistance. The mutated grlA genes responsible for quinolone resistance were dominant over the wild-type allele, irrespective of gene dosage in a transformation experiment with the grlA gene alone. However, dominance by mutated grlA genes depended on gene dosage when bacteria were transformed with the grlA and grlB genes in combination. Quinolone-resistant gyrA mutants were easily isolated from a strain, S. aureus RN4220, carrying a plasmid with the mutated grlA gene, though this was not the case for other S. aureus strains lacking the plasmid. The elimination of this plasmid from such quinolone-resistant gyrA mutants resulted in marked increases in quinolone susceptibility. These results suggest that both DNA gyrase and DNA topoisomerase IV may be targets of quinolones and that the quinolone susceptibility of organisms may be determined by which of these enzymes is most quinolone sensitive.

Full Text

The Full Text of this article is available as a PDF (283.3 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Fasching C. E., Tenover F. C., Slama T. G., Fisher L. M., Sreedharan S., Oram M., Willard K., Sinn L. M., Gerding D. N., Peterson L. R. gyrA mutations in ciprofloxacin-resistant, methicillin-resistant Staphylococcus aureus from Indiana, Minnesota, and Tennessee. J Infect Dis. 1991 Nov;164(5):976–979. doi: 10.1093/infdis/164.5.976. [DOI] [PubMed] [Google Scholar]
  2. Ferrero L., Cameron B., Crouzet J. Analysis of gyrA and grlA mutations in stepwise-selected ciprofloxacin-resistant mutants of Staphylococcus aureus. Antimicrob Agents Chemother. 1995 Jul;39(7):1554–1558. doi: 10.1128/aac.39.7.1554. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Ferrero L., Cameron B., Manse B., Lagneaux D., Crouzet J., Famechon A., Blanche F. Cloning and primary structure of Staphylococcus aureus DNA topoisomerase IV: a primary target of fluoroquinolones. Mol Microbiol. 1994 Aug;13(4):641–653. doi: 10.1111/j.1365-2958.1994.tb00458.x. [DOI] [PubMed] [Google Scholar]
  4. Goswitz J. J., Willard K. E., Fasching C. E., Peterson L. R. Detection of gyrA gene mutations associated with ciprofloxacin resistance in methicillin-resistant Staphylococcus aureus: analysis by polymerase chain reaction and automated direct DNA sequencing. Antimicrob Agents Chemother. 1992 May;36(5):1166–1169. doi: 10.1128/aac.36.5.1166. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Holmes D. S., Quigley M. A rapid boiling method for the preparation of bacterial plasmids. Anal Biochem. 1981 Jun;114(1):193–197. doi: 10.1016/0003-2697(81)90473-5. [DOI] [PubMed] [Google Scholar]
  6. Hooper D. C. Quinolone mode of action. Drugs. 1995;49 (Suppl 2):10–15. doi: 10.2165/00003495-199500492-00004. [DOI] [PubMed] [Google Scholar]
  7. Hori S., Ohshita Y., Utsui Y., Hiramatsu K. Sequential acquisition of norfloxacin and ofloxacin resistance by methicillin-resistant and -susceptible Staphylococcus aureus. Antimicrob Agents Chemother. 1993 Nov;37(11):2278–2284. doi: 10.1128/aac.37.11.2278. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Hoshino K., Kitamura A., Morrissey I., Sato K., Kato J., Ikeda H. Comparison of inhibition of Escherichia coli topoisomerase IV by quinolones with DNA gyrase inhibition. Antimicrob Agents Chemother. 1994 Nov;38(11):2623–2627. doi: 10.1128/aac.38.11.2623. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hudson M. C., Curtiss R., 3rd Regulation of expression of Streptococcus mutans genes important to virulence. Infect Immun. 1990 Feb;58(2):464–470. doi: 10.1128/iai.58.2.464-470.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Israel D. I. A PCR-based method for high stringency screening of DNA libraries. Nucleic Acids Res. 1993 Jun 11;21(11):2627–2631. doi: 10.1093/nar/21.11.2627. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ito H., Yoshida H., Bogaki-Shonai M., Niga T., Hattori H., Nakamura S. Quinolone resistance mutations in the DNA gyrase gyrA and gyrB genes of Staphylococcus aureus. Antimicrob Agents Chemother. 1994 Sep;38(9):2014–2023. doi: 10.1128/aac.38.9.2014. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Ito W., Ishiguro H., Kurosawa Y. A general method for introducing a series of mutations into cloned DNA using the polymerase chain reaction. Gene. 1991 Jun 15;102(1):67–70. doi: 10.1016/0378-1119(91)90539-n. [DOI] [PubMed] [Google Scholar]
  13. Kaatz G. W., Seo S. M., Ruble C. A. Efflux-mediated fluoroquinolone resistance in Staphylococcus aureus. Antimicrob Agents Chemother. 1993 May;37(5):1086–1094. doi: 10.1128/aac.37.5.1086. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Koga H., Itoh A., Murayama S., Suzue S., Irikura T. Structure-activity relationships of antibacterial 6,7- and 7,8-disubstituted 1-alkyl-1,4-dihydro-4-oxoquinoline-3-carboxylic acids. J Med Chem. 1980 Dec;23(12):1358–1363. doi: 10.1021/jm00186a014. [DOI] [PubMed] [Google Scholar]
  15. Kojima T., Inoue M., Mitsuhashi S. In vitro activity of AT-4140 against quinolone- and methicillin-resistant Staphylococcus aureus. Antimicrob Agents Chemother. 1990 Jun;34(6):1123–1127. doi: 10.1128/aac.34.6.1123. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kojima T., Yamagishi J., Oyamada Y., Yoshida H., Hattori H., Inoue M., Nakamura S. Analysis of quinolone resistance genes in a clinical isolate of quinolone-resistant MRSA. Drugs. 1995;49 (Suppl 2):182–184. doi: 10.2165/00003495-199500492-00034. [DOI] [PubMed] [Google Scholar]
  17. Kreuzer K. N., Cozzarelli N. R. Escherichia coli mutants thermosensitive for deoxyribonucleic acid gyrase subunit A: effects on deoxyribonucleic acid replication, transcription, and bacteriophage growth. J Bacteriol. 1979 Nov;140(2):424–435. doi: 10.1128/jb.140.2.424-435.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Miyamoto T., Matsumoto J., Chiba K., Egawa H., Shibamori K., Minamida A., Nishimura Y., Okada H., Kataoka M., Fujita M. Synthesis and structure-activity relationships of 5-substituted 6,8-difluoroquinolones, including sparfloxacin, a new quinolone antibacterial agent with improved potency. J Med Chem. 1990 Jun;33(6):1645–1656. doi: 10.1021/jm00168a018. [DOI] [PubMed] [Google Scholar]
  19. Nakamura S., Nakamura M., Kojima T., Yoshida H. gyrA and gyrB mutations in quinolone-resistant strains of Escherichia coli. Antimicrob Agents Chemother. 1989 Feb;33(2):254–255. doi: 10.1128/aac.33.2.254. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Ng E. Y., Trucksis M., Hooper D. C. Quinolone resistance mediated by norA: physiologic characterization and relationship to flqB, a quinolone resistance locus on the Staphylococcus aureus chromosome. Antimicrob Agents Chemother. 1994 Jun;38(6):1345–1355. doi: 10.1128/aac.38.6.1345. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. O'sullivan D. J., Klaenhammer T. R. Rapid Mini-Prep Isolation of High-Quality Plasmid DNA from Lactococcus and Lactobacillus spp. Appl Environ Microbiol. 1993 Aug;59(8):2730–2733. doi: 10.1128/aem.59.8.2730-2733.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Schaefler S. Methicillin-resistant strains of Staphylococcus aureus resistant to quinolones. J Clin Microbiol. 1989 Feb;27(2):335–336. doi: 10.1128/jcm.27.2.335-336.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Shalit I., Berger S. A., Gorea A., Frimerman H. Widespread quinolone resistance among methicillin-resistant Staphylococcus aureus isolates in a general hospital. Antimicrob Agents Chemother. 1989 Apr;33(4):593–594. doi: 10.1128/aac.33.4.593. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. 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]
  25. Sreedharan S., Peterson L. R., Fisher L. M. Ciprofloxacin resistance in coagulase-positive and -negative staphylococci: role of mutations at serine 84 in the DNA gyrase A protein of Staphylococcus aureus and Staphylococcus epidermidis. Antimicrob Agents Chemother. 1991 Oct;35(10):2151–2154. doi: 10.1128/aac.35.10.2151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Trucksis M., Wolfson J. S., Hooper D. C. A novel locus conferring fluoroquinolone resistance in Staphylococcus aureus. J Bacteriol. 1991 Sep;173(18):5854–5860. doi: 10.1128/jb.173.18.5854-5860.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Wilkie N. M., Clements J. B., Boll W., Mantei N., Lonsdale D., Weissmann C. Hybrid plasmids containing an active thymidine kinase gene of Herpes simplex virus 1. Nucleic Acids Res. 1979 Oct 25;7(4):859–877. doi: 10.1093/nar/7.4.859. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Yoshida H., Bogaki M., Nakamura M., Nakamura S. Quinolone resistance-determining region in the DNA gyrase gyrA gene of Escherichia coli. Antimicrob Agents Chemother. 1990 Jun;34(6):1271–1272. doi: 10.1128/aac.34.6.1271. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Yoshida H., Bogaki M., Nakamura M., Yamanaka L. M., Nakamura S. Quinolone resistance-determining region in the DNA gyrase gyrB gene of Escherichia coli. Antimicrob Agents Chemother. 1991 Aug;35(8):1647–1650. doi: 10.1128/aac.35.8.1647. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. 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]

Articles from Antimicrobial Agents and Chemotherapy are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES