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. 1996 Sep;40(9):1988–1994. doi: 10.1128/aac.40.9.1988

Molecular aspects of high-level resistance to sulbactam-cefoperazone in Klebsiella oxytoca clinical isolates.

K Kimura 1, Y Arakawa 1, S Ohsuka 1, H Ito 1, K Suzuki 1, H Kurokawa 1, N Kato 1, M Ohta 1
PMCID: PMC163460  PMID: 8878568

Abstract

Nine Klebsiella oxytoca strains which demonstrated resistance to the combination of sulbactam and cefoperazone were isolated from geographically separate hospitals in Japan in 1995. Among them, K. oxytoca SB23 showed high-level resistance to sulbactam-cefoperazone (MIC > 128 micrograms/ml) and aztreonam (MIC, 128 micrograms/ml). The sulbactam-cefoperazone resistance was not transferred from strain SB23 to Escherichia coli CSH2 by conjugation, beta-Lactamase RbiA, produced by strain SB23, was purified, and the molecular mass was estimated to be 29 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Kinetic parameters for RbiA revealed that cefoperazone and aztreonam were hydrolyzed efficiently by this enzyme. Moreover, ceftazidime and imipenem were also hydrolyzed weakly by RbiA, although strain SB23 did not show any resistance to these agents. Clavulanate, sulbactam, and tazobactam failed to block the hydrolysis of cefoperazone by RbiA. The structural gene of RbiA (blaRBI) was cloned and sequenced, and the deduced amino acid sequence of RbiA demonstrated high-level similarities to those of the beta-lactamases found in K. oxytoca D488, E23004, and plasmid-mediated MEN-1, which have been classified into Bush functional group 2be. Although RbiA demonstrates high-level molecular similarity to the enzymes in group 2be, from an enzymological point of view, this enzyme might be differentiated from the enzymes in that group. Hybridization analysis revealed that beta-lactamase genes highly similar to blaRBI were generally encoded on the chromosome of the sulbactam-cefoperazone-resistant clinical isolates of K. oxytoca tested in the study, despite their different derivations. This observation suggests that sulbactam-cefoperazone-resistant A. oxytoca strains which produce RbiA-type beta-lactamases have been proliferating in many hospitals in Japan.

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Selected References

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  1. Ambler R. P., Coulson A. F., Frère J. M., Ghuysen J. M., Joris B., Forsman M., Levesque R. C., Tiraby G., Waley S. G. A standard numbering scheme for the class A beta-lactamases. Biochem J. 1991 May 15;276(Pt 1):269–270. doi: 10.1042/bj2760269. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. 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]
  3. Arakawa Y., Ohta M., Kido N., Fujii Y., Komatsu T., Kato N. Close evolutionary relationship between the chromosomally encoded beta-lactamase gene of Klebsiella pneumoniae and the TEM beta-lactamase gene mediated by R plasmids. FEBS Lett. 1986 Oct 20;207(1):69–74. doi: 10.1016/0014-5793(86)80014-x. [DOI] [PubMed] [Google Scholar]
  4. Arakawa Y., Ohta M., Kido N., Mori M., Ito H., Komatsu T., Fujii Y., Kato N. Chromosomal beta-lactamase of Klebsiella oxytoca, a new class A enzyme that hydrolyzes broad-spectrum beta-lactam antibiotics. Antimicrob Agents Chemother. 1989 Jan;33(1):63–70. doi: 10.1128/aac.33.1.63. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Barthélémy M., Péduzzi J., Bernard H., Tancrède C., Labia R. Close amino acid sequence relationship between the new plasmid-mediated extended-spectrum beta-lactamase MEN-1 and chromosomally encoded enzymes of Klebsiella oxytoca. Biochim Biophys Acta. 1992 Jul 13;1122(1):15–22. doi: 10.1016/0167-4838(92)90121-s. [DOI] [PubMed] [Google Scholar]
  6. Belaaouaj A., Lapoumeroulie C., Caniça M. M., Vedel G., Névot P., Krishnamoorthy R., Paul G. Nucleotide sequences of the genes coding for the TEM-like beta-lactamases IRT-1 and IRT-2 (formerly called TRI-1 and TRI-2). FEMS Microbiol Lett. 1994 Jul 1;120(1-2):75–80. doi: 10.1111/j.1574-6968.1994.tb07010.x. [DOI] [PubMed] [Google Scholar]
  7. Blazquez J., Baquero M. R., Canton R., Alos I., Baquero F. Characterization of a new TEM-type beta-lactamase resistant to clavulanate, sulbactam, and tazobactam in a clinical isolate of Escherichia coli. Antimicrob Agents Chemother. 1993 Oct;37(10):2059–2063. doi: 10.1128/aac.37.10.2059. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Boras G. J., Au S., Roy K. L., von Tigerstrom R. G. beta-Lactamase of Lysobacter enzymogenes: cloning, characterization and expression of the gene and comparison of the enzyme to other lactamases. J Gen Microbiol. 1993 Jun;139(Pt 6):1245–1252. doi: 10.1099/00221287-139-6-1245. [DOI] [PubMed] [Google Scholar]
  9. Brun T., Péduzzi J., Caniça M. M., Paul G., Névot P., Barthélémy M., Labia R. Characterization and amino acid sequence of IRT-4, a novel TEM-type enzyme with a decreased susceptibility to beta-lactamase inhibitors. FEMS Microbiol Lett. 1994 Jul 1;120(1-2):111–117. doi: 10.1111/j.1574-6968.1994.tb07016.x. [DOI] [PubMed] [Google Scholar]
  10. 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]
  11. Bush K., Sykes R. B. Methodology for the study of beta-lactamases. Antimicrob Agents Chemother. 1986 Jul;30(1):6–10. doi: 10.1128/aac.30.1.6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fournier B., Lu C. Y., Lagrange P. H., Krishnamoorthy R., Philippon A. Point mutation in the pribnow box, the molecular basis of beta-lactamase overproduction in Klebsiella oxytoca. Antimicrob Agents Chemother. 1995 Jun;39(6):1365–1368. doi: 10.1128/aac.39.6.1365. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Henquell C., Chanal C., Sirot D., Labia R., Sirot J. Molecular characterization of nine different types of mutants among 107 inhibitor-resistant TEM beta-lactamases from clinical isolates of Escherichia coli. Antimicrob Agents Chemother. 1995 Feb;39(2):427–430. doi: 10.1128/aac.39.2.427. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Herzberg O., Moult J. Bacterial resistance to beta-lactam antibiotics: crystal structure of beta-lactamase from Staphylococcus aureus PC1 at 2.5 A resolution. Science. 1987 May 8;236(4802):694–701. doi: 10.1126/science.3107125. [DOI] [PubMed] [Google Scholar]
  15. Horii T., Arakawa Y., Ohta M., Ichiyama S., Wacharotayankun R., Kato N. Plasmid-mediated AmpC-type beta-lactamase isolated from Klebsiella pneumoniae confers resistance to broad-spectrum beta-lactams, including moxalactam. Antimicrob Agents Chemother. 1993 May;37(5):984–990. doi: 10.1128/aac.37.5.984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Horii T., Arakawa Y., Ohta M., Sugiyama T., Wacharotayankun R., Ito H., Kato N. Characterization of a plasmid-borne and constitutively expressed blaMOX-1 gene encoding AmpC-type beta-lactamase. Gene. 1994 Feb 11;139(1):93–98. doi: 10.1016/0378-1119(94)90529-0. [DOI] [PubMed] [Google Scholar]
  17. Ishii Y., Ohno A., Taguchi H., Imajo S., Ishiguro M., Matsuzawa H. Cloning and sequence of the gene encoding a cefotaxime-hydrolyzing class A beta-lactamase isolated from Escherichia coli. Antimicrob Agents Chemother. 1995 Oct;39(10):2269–2275. doi: 10.1128/aac.39.10.2269. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Ito H., Arakawa Y., Ohsuka S., Wacharotayankun R., Kato N., Ohta M. Plasmid-mediated dissemination of the metallo-beta-lactamase gene blaIMP among clinically isolated strains of Serratia marcescens. Antimicrob Agents Chemother. 1995 Apr;39(4):824–829. doi: 10.1128/aac.39.4.824. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Joris B., Ghuysen J. M., Dive G., Renard A., Dideberg O., Charlier P., Frère J. M., Kelly J. A., Boyington J. C., Moews P. C. The active-site-serine penicillin-recognizing enzymes as members of the Streptomyces R61 DD-peptidase family. Biochem J. 1988 Mar 1;250(2):313–324. doi: 10.1042/bj2500313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Lemozy J., Sirot D., Chanal C., Huc C., Labia R., Dabernat H., Sirot J. First characterization of inhibitor-resistant TEM (IRT) beta-lactamases in Klebsiella pneumoniae strains. Antimicrob Agents Chemother. 1995 Nov;39(11):2580–2582. doi: 10.1128/aac.39.11.2580. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. 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]
  22. Ohsuka S., Arakawa Y., Horii T., Ito H., Ohta M. Effect of pH on activities of novel beta-lactamases and beta-lactamase inhibitors against these beta-lactamases. Antimicrob Agents Chemother. 1995 Aug;39(8):1856–1858. doi: 10.1128/aac.39.8.1856. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Osano E., Arakawa Y., Wacharotayankun R., Ohta M., Horii T., Ito H., Yoshimura F., Kato N. Molecular characterization of an enterobacterial metallo beta-lactamase found in a clinical isolate of Serratia marcescens that shows imipenem resistance. Antimicrob Agents Chemother. 1994 Jan;38(1):71–78. doi: 10.1128/aac.38.1.71. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Payne D. J., Cramp R., Winstanley D. J., Knowles D. J. Comparative activities of clavulanic acid, sulbactam, and tazobactam against clinically important beta-lactamases. Antimicrob Agents Chemother. 1994 Apr;38(4):767–772. doi: 10.1128/aac.38.4.767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Perilli M., Franceschini N., Segatore B., Amicosante G., Oratore A., Duez C., Joris B., Frère J. M. Cloning and nucleotide sequencing of the gene encoding the beta-lactamase from Citrobacter diversus. FEMS Microbiol Lett. 1991 Sep 15;67(1):79–84. doi: 10.1016/0378-1097(91)90448-j. [DOI] [PubMed] [Google Scholar]
  26. Péduzzi J., Reynaud A., Baron P., Barthélémy M., Labia R. Chromosomally encoded cephalosporin-hydrolyzing beta-lactamase of Proteus vulgaris RO104 belongs to Ambler's class A. Biochim Biophys Acta. 1994 Jul 20;1207(1):31–39. doi: 10.1016/0167-4838(94)90048-5. [DOI] [PubMed] [Google Scholar]
  27. Reynaud A., Péduzzi J., Barthélémy M., Labia R. Cefotaxime-hydrolysing activity of the beta-lactamase of Klebsiella oxytoca D488 could be related to a threonine residue at position 140. FEMS Microbiol Lett. 1991 Jun 15;65(2):185–192. doi: 10.1016/0378-1097(91)90301-p. [DOI] [PubMed] [Google Scholar]
  28. Rice L. B., Carias L. L., Etter L., Shlaes D. M. Resistance to cefoperazone-sulbactam in Klebsiella pneumoniae: evidence for enhanced resistance resulting from the coexistence of two different resistance mechanisms. Antimicrob Agents Chemother. 1993 May;37(5):1061–1064. doi: 10.1128/aac.37.5.1061. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Seoane A., García Lobo J. M. Nucleotide sequence of a new class A beta-lactamase gene from the chromosome of Yersinia enterocolitica: implications for the evolution of class A beta-lactamases. Mol Gen Genet. 1991 Aug;228(1-2):215–220. doi: 10.1007/BF00282468. [DOI] [PubMed] [Google Scholar]
  30. Stapleton P., Wu P. J., King A., Shannon K., French G., Phillips I. Incidence and mechanisms of resistance to the combination of amoxicillin and clavulanic acid in Escherichia coli. Antimicrob Agents Chemother. 1995 Nov;39(11):2478–2483. doi: 10.1128/aac.39.11.2478. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Thomson K. S., Weber D. A., Sanders C. C., Sanders W. E., Jr Beta-lactamase production in members of the family Enterobacteriaceae and resistance to beta-lactam-enzyme inhibitor combinations. Antimicrob Agents Chemother. 1990 Apr;34(4):622–627. doi: 10.1128/aac.34.4.622. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. 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]

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