Skip to main content
PMC home page
Antimicrobial Agents and Chemotherapy logoLink to Antimicrobial Agents and Chemotherapy
. 1997 Oct;41(10):2188–2195. doi: 10.1128/aac.41.10.2188

OXA-18, a class D clavulanic acid-inhibited extended-spectrum beta-lactamase from Pseudomonas aeruginosa.

L N Philippon 1, T Naas 1, A T Bouthors 1, V Barakett 1, P Nordmann 1
PMCID: PMC164091  PMID: 9333046

Abstract

Clinical isolate Pseudomonas aeruginosa Mus showed resistance both to extended-spectrum cephalosporins and to aztreonam. We detected a typical double-disk synergy image when ceftazidime or aztreonam was placed next to a clavulanic acid disk on an agar plate. This resistance phenotype suggested the presence of an extended-spectrum beta-lactamase. Isoelectric focusing revealed that this strain produced three beta-lactamases, of pI 5.5, 7.4, and 8.2. A 2.6-kb Sau3A fragment encoding the extended-spectrum beta-lactamase of pI 5.5 was cloned from P. aeruginosa Mus genomic DNA. This enzyme, named OXA-18, had a relative molecular mass of 30.6 kDa. OXA-18 has a broad substrate profile, hydrolyzing amoxicillin, ticarcillin, cephalothin, ceftazidime, cefotaxime, and aztreonam, but not imipenem or cephamycins. Its activity was totally inhibited by clavulanic acid at 2 microg/ml. Hydrolysis constants of OXA-18 (Vmax, Km) confirmed the MIC results. Cloxacillin and oxacillin hydrolysis was noticeable with the partially purified OXA-18. The blaOXA-18 gene encodes a 275-amino-acid protein which has weak identity with all class D beta-lactamases except OXA-9 and OXA-12 (45 and 42% amino acid identity, respectively). OXA-18 is likely to be chromosomally encoded since no plasmid was found in the strain and because attempts to transfer the resistance marker failed. OXA-18 is peculiar since it is a class D beta-lactamase which confers high resistance to extended-spectrum cephalosporins and seems to have unique hydrolytic properties among non-class A enzymes.

Full Text

The Full Text of this article is available as a PDF (295.9 KB).

Selected References

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

  1. Ambler R. P. The structure of beta-lactamases. Philos Trans R Soc Lond B Biol Sci. 1980 May 16;289(1036):321–331. doi: 10.1098/rstb.1980.0049. [DOI] [PubMed] [Google Scholar]
  2. Bush K., Jacoby G. A., Medeiros A. A. A functional classification scheme for beta-lactamases and its correlation with molecular structure. Antimicrob Agents Chemother. 1995 Jun;39(6):1211–1233. doi: 10.1128/aac.39.6.1211. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Collis C. M., Hall R. M. Site-specific deletion and rearrangement of integron insert genes catalyzed by the integron DNA integrase. J Bacteriol. 1992 Mar;174(5):1574–1585. doi: 10.1128/jb.174.5.1574-1585.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Couture F., Lachapelle J., Levesque R. C. Phylogeny of LCR-1 and OXA-5 with class A and class D beta-lactamases. Mol Microbiol. 1992 Jun;6(12):1693–1705. doi: 10.1111/j.1365-2958.1992.tb00894.x. [DOI] [PubMed] [Google Scholar]
  5. Dale J. W., Godwin D., Mossakowska D., Stephenson P., Wall S. Sequence of the OXA2 beta-lactamase: comparison with other penicillin-reactive enzymes. FEBS Lett. 1985 Oct 21;191(1):39–44. doi: 10.1016/0014-5793(85)80989-3. [DOI] [PubMed] [Google Scholar]
  6. Danel F., Hall L. M., Gur D., Akalin H. E., Livermore D. M. Transferable production of PER-1 beta-lactamase in Pseudomonas aeruginosa. J Antimicrob Chemother. 1995 Feb;35(2):281–294. doi: 10.1093/jac/35.2.281. [DOI] [PubMed] [Google Scholar]
  7. Danel F., Hall L. M., Gur D., Livermore D. M. OXA-14, another extended-spectrum variant of OXA-10 (PSE-2) beta-lactamase from Pseudomonas aeruginosa. Antimicrob Agents Chemother. 1995 Aug;39(8):1881–1884. doi: 10.1128/aac.39.8.1881. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Danel F., Hall L. M., Gur D., Livermore D. M. OXA-15, an extended-spectrum variant of OXA-2 beta-lactamase, isolated from a Pseudomonas aeruginosa strain. Antimicrob Agents Chemother. 1997 Apr;41(4):785–790. doi: 10.1128/aac.41.4.785. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Feng D. F., Doolittle R. F. Progressive alignment and phylogenetic tree construction of protein sequences. Methods Enzymol. 1990;183:375–387. doi: 10.1016/0076-6879(90)83025-5. [DOI] [PubMed] [Google Scholar]
  10. Hall L. M., Livermore D. M., Gur D., Akova M., Akalin H. E. OXA-11, an extended-spectrum variant of OXA-10 (PSE-2) beta-lactamase from Pseudomonas aeruginosa. Antimicrob Agents Chemother. 1993 Aug;37(8):1637–1644. doi: 10.1128/aac.37.8.1637. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hall R. M., Vockler C. The region of the IncN plasmid R46 coding for resistance to beta-lactam antibiotics, streptomycin/spectinomycin and sulphonamides is closely related to antibiotic resistance segments found in IncW plasmids and in Tn21-like transposons. Nucleic Acids Res. 1987 Sep 25;15(18):7491–7501. doi: 10.1093/nar/15.18.7491. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hansen J. B., Olsen R. H. Isolation of large bacterial plasmids and characterization of the P2 incompatibility group plasmids pMG1 and pMG5. J Bacteriol. 1978 Jul;135(1):227–238. doi: 10.1128/jb.135.1.227-238.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Huovinen P., Huovinen S., Jacoby G. A. Sequence of PSE-2 beta-lactamase. Antimicrob Agents Chemother. 1988 Jan;32(1):134–136. doi: 10.1128/aac.32.1.134. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Jacoby G. A., Carreras I. Activities of beta-lactam antibiotics against Escherichia coli strains producing extended-spectrum beta-lactamases. Antimicrob Agents Chemother. 1990 May;34(5):858–862. doi: 10.1128/aac.34.5.858. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Joris B., Ledent P., Dideberg O., Fonzé E., Lamotte-Brasseur J., Kelly J. A., Ghuysen J. M., Frère J. M. Comparison of the sequences of class A beta-lactamases and of the secondary structure elements of penicillin-recognizing proteins. Antimicrob Agents Chemother. 1991 Nov;35(11):2294–2301. doi: 10.1128/aac.35.11.2294. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kado C. I., Liu S. T. Rapid procedure for detection and isolation of large and small plasmids. J Bacteriol. 1981 Mar;145(3):1365–1373. doi: 10.1128/jb.145.3.1365-1373.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Levesque R. C., Jacoby G. A. Molecular structure and interrelationships of multiresistance beta-lactamase transposons. Plasmid. 1988 Jan;19(1):21–29. doi: 10.1016/0147-619x(88)90059-5. [DOI] [PubMed] [Google Scholar]
  18. Livermore D. M. beta-Lactamases in laboratory and clinical resistance. Clin Microbiol Rev. 1995 Oct;8(4):557–584. doi: 10.1128/cmr.8.4.557. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Massidda O., Rossolini G. M., Satta G. The Aeromonas hydrophila cphA gene: molecular heterogeneity among class B metallo-beta-lactamases. J Bacteriol. 1991 Aug;173(15):4611–4617. doi: 10.1128/jb.173.15.4611-4617.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Matagne A., Misselyn-Bauduin A. M., Joris B., Erpicum T., Granier B., Frère J. M. The diversity of the catalytic properties of class A beta-lactamases. Biochem J. 1990 Jan 1;265(1):131–146. doi: 10.1042/bj2650131. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Monks J., Waley S. G. Imipenem as substrate and inhibitor of beta-lactamases. Biochem J. 1988 Jul 15;253(2):323–328. doi: 10.1042/bj2530323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Mossakowska D., Ali N. A., Dale J. W. Oxacillin-hydrolysing beta-lactamases. A comparative analysis at nucleotide and amino acid sequence levels. Eur J Biochem. 1989 Mar 15;180(2):309–318. doi: 10.1111/j.1432-1033.1989.tb14649.x. [DOI] [PubMed] [Google Scholar]
  23. Mugnier P., Dubrous P., Casin I., Arlet G., Collatz E. A TEM-derived extended-spectrum beta-lactamase in Pseudomonas aeruginosa. Antimicrob Agents Chemother. 1996 Nov;40(11):2488–2493. doi: 10.1128/aac.40.11.2488. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Nicolas M. H., Jarlier V., Honore N., Philippon A., Cole S. T. Molecular characterization of the gene encoding SHV-3 beta-lactamase responsible for transferable cefotaxime resistance in clinical isolates of Klebsiella pneumoniae. Antimicrob Agents Chemother. 1989 Dec;33(12):2096–2100. doi: 10.1128/aac.33.12.2096. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Nordmann P., Naas T. Sequence analysis of PER-1 extended-spectrum beta-lactamase from Pseudomonas aeruginosa and comparison with class A beta-lactamases. Antimicrob Agents Chemother. 1994 Jan;38(1):104–114. doi: 10.1128/aac.38.1.104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Nordmann P., Ronco E., Naas T., Duport C., Michel-Briand Y., Labia R. Characterization of a novel extended-spectrum beta-lactamase from Pseudomonas aeruginosa. Antimicrob Agents Chemother. 1993 May;37(5):962–969. doi: 10.1128/aac.37.5.962. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Ouellette M., Bissonnette L., Roy P. H. Precise insertion of antibiotic resistance determinants into Tn21-like transposons: nucleotide sequence of the OXA-1 beta-lactamase gene. Proc Natl Acad Sci U S A. 1987 Nov;84(21):7378–7382. doi: 10.1073/pnas.84.21.7378. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Rasmussen B. A., Keeney D., Yang Y., Bush K. Cloning and expression of a cloxacillin-hydrolyzing enzyme and a cephalosporinase from Aeromonas sobria AER 14M in Escherichia coli: requirement for an E. coli chromosomal mutation for efficient expression of the class D enzyme. Antimicrob Agents Chemother. 1994 Sep;38(9):2078–2085. doi: 10.1128/aac.38.9.2078. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sanschagrin F., Couture F., Levesque R. C. Primary structure of OXA-3 and phylogeny of oxacillin-hydrolyzing class D beta-lactamases. Antimicrob Agents Chemother. 1995 Apr;39(4):887–893. doi: 10.1128/aac.39.4.887. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Scoulica E., Aransay A., Tselentis Y. Molecular characterization of the OXA-7 beta-lactamase gene. Antimicrob Agents Chemother. 1995 Jun;39(6):1379–1382. doi: 10.1128/aac.39.6.1379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Stokes H. W., Hall R. M. A novel family of potentially mobile DNA elements encoding site-specific gene-integration functions: integrons. Mol Microbiol. 1989 Dec;3(12):1669–1683. doi: 10.1111/j.1365-2958.1989.tb00153.x. [DOI] [PubMed] [Google Scholar]
  32. Sutcliffe J. G. Nucleotide sequence of the ampicillin resistance gene of Escherichia coli plasmid pBR322. Proc Natl Acad Sci U S A. 1978 Aug;75(8):3737–3741. doi: 10.1073/pnas.75.8.3737. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Takahashi S., Nagano Y. Rapid procedure for isolation of plasmid DNA and application to epidemiological analysis. J Clin Microbiol. 1984 Oct;20(4):608–613. doi: 10.1128/jcm.20.4.608-613.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Tolmasky M. E., Crosa J. H. Genetic organization of antibiotic resistance genes (aac(6')-Ib, aadA, and oxa9) in the multiresistance transposon Tn1331. Plasmid. 1993 Jan;29(1):31–40. doi: 10.1006/plas.1993.1004. [DOI] [PubMed] [Google Scholar]
  35. Tolmasky M. E. Sequencing and expression of aadA, bla, and tnpR from the multiresistance transposon Tn1331. Plasmid. 1990 Nov;24(3):218–226. doi: 10.1016/0147-619x(90)90005-w. [DOI] [PubMed] [Google Scholar]
  36. Vahaboglu H., Hall L. M., Mulazimoglu L., Dodanli S., Yildirim I., Livermore D. M. Resistance to extended-spectrum cephalosporins, caused by PER-1 beta-lactamase, in Salmonella typhimurium from Istanbul, Turkey. J Med Microbiol. 1995 Oct;43(4):294–299. doi: 10.1099/00222615-43-4-294. [DOI] [PubMed] [Google Scholar]
  37. West S. E., Iglewski B. H. Codon usage in Pseudomonas aeruginosa. Nucleic Acids Res. 1988 Oct 11;16(19):9323–9335. doi: 10.1093/nar/16.19.9323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]

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

RESOURCES