Korean J Clin Microbiol.  2009 Mar;12(1):24-29. 10.5145/KJCM.2009.12.1.24.

Use of Boronic Acid Disks for the Detection of Extended-spectrum beta-lactamase and AmpC beta-lactamase in Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca and Proteus mirabilis

Affiliations
  • 1Department of Laboratory Medicine, Yonsei University Wonju College of Medicine, Wonju, Korea. mizpark66@empal.com
  • 2Department of Infectious Diseases, Yonsei University Wonju College of Medicine, Wonju, Korea.

Abstract

BACKGROUND
Accurate detection of organisms producing extended-spectrum beta-lactamase (ESBL) and AmpC beta-lactamase is very important for treatment of patients. However, unlike the ESBL confirmatory test, there are no guidelines for detection of organisms producing AmpC beta-lactamase. We evaluated a detection method using boronic acid (BA) for ESBL and AmpC beta-lactamase.
METHODS
Clinical isolates of Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca, and Proteus mirabilis showing intermediate resistance or resistance to cefoxitin (FOX) or positive for ESBL were tested. A > or =5 mm increase in zone diameter of ceftazidime/clavulanic acid/BA (CAZ/CA/BA) and/or cefotaxime/clavulanic acid/BA (CTX/CA/BA) versus CAZ/BA and/or CTX /BA was considered positive for ESBL. Likewise, a > or =5 mm increase in zone diameter of FOX/BA and/or cefotetan/BA (CTT/BA) versus FOX and/or CTT alone was considered positive for AmpC beta-lactamase.
RESULTS
Among 622 clinical isolates, ESBL positive rates by the CLSI ESBL confirmatory test or by the BA method were 18.1% or 18.4% for E. coli, 38.3% or 40.4% for K. pneumoniae, 8.7% or 8.7% for K. oxytoca, and 14.8% or 14.8% for P. mirabilis, respectively. AmpC beta-lactamase positive rates using the BA method were 3.7% for E. coli, 33.3% for K. pneumoniae, 0% for K. oxytoca, and 7.4% for P. mirabilis. The detection rates of coproducing ESBL and AmpC beta-lactamase were 2.4% in E. coli 27.1% in K. pneumoniae, and 3.7% in P. mirabilis.
CONCLUSION
The ESBL confirmatory method using BA was found to enhance the detection of ESBLs, even when potentially masked by AmpC beta-lactamase.

Keyword

Extended-spectrum beta-lactamase; Plasmid- mediated AmpC beta-lactamase; Boronic acid; Coproducers

MeSH Terms

Bacterial Proteins
beta-Lactamases
Boron
Cefoxitin
Escherichia
Escherichia coli
Humans
Klebsiella
Klebsiella oxytoca
Klebsiella pneumoniae
Masks
Mirabilis
Penicillinase
Pneumonia
Proteus
Proteus mirabilis
Bacterial Proteins
Boron
Cefoxitin
Penicillinase
beta-Lactamases

Figure

  • Fig. 1. Representative results using the Clinical and Laboratory Standards Institute extended-spectrum β-lactamase (ESBL) confirmatory test and AmpC disk test without and with boronic acid (two disks positioned in right side on media). Abbreviations: FOX, cefoxitin; CAZ, ceftazidime; CV, clavulanic acid; CTX, cefotaxime; CTX, cefotaxime. Left media shows AmpC positive, center media shows ESBL positive, and right media shows that isolate is negative in CLSI method and positive in boronic acid disk.


Reference

1. Uh Y, Kim HY, et al. Antimicrobial Agents and Antimicrobial Susceptibility Test. 1st ed.Paju: KIS;2007. p. 118–9.
2. Beesley T, Gascoyne N, Knott-Hunziker V, Petursson S, Waley SG, Jaurin B, et al. The inhibition of class C β-lactamases by boronic acids. Biochem J. 1983; 209:229–33.
Article
3. Brenwald NP, Jevons G, Andrews J, Ang L, Fraise P. Disc methods for detecting AmpC β-lactamase-producing clinical isolates of Escherichia coli and Klebsiella pneumoniae. J Antimicrob Chemother. 2005; 56:600–1.
4. Coudron PE, Moland ES, Thomson KS. Occurrence and detection of AmpC β-lactamases among Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis isolates at a veterans medical center. J Clin Microbiol. 2000; 38:1791–6.
5. Yagi T, Wachino J, Kurokawa H, Suzuki S, Yamane K, Doi Y, et al. Practical methods using boronic acid compounds for identification of class C β-lactamase-producing Klebsiella pneumoniae and Escherichia coli. J Clin Microbiol. 2005; 43:2551–8.
6. Song W, Bae IK, Lee YN, Lee CH, Lee SH, Jeong SH. Detection of extended-spectrum β-lactamases by using boronic acid as an AmpC β-lactamase inhibitor in clinical isolates of Klebsiella spp. and Escherichia coli. J Clin Microbiol. 2007; 45:1180–4.
7. Song W, Jeong SH, Kim JS, Kim HS, Shin DH, Roh KH, et al. Use of boronic acid disk methods to detect the combined expression of plasmid-mediated AmpC β-lactamases and extended-spectrum β-lactamases in clinical isolates of Klebsiella spp., Salmonella spp., and Proteus mirabilis. Diagn Microbiol Infect Dis. 2007; 57:315–8.
8. Uh Y, Son JS, Hwang GY, Jang IH, Yoon KJ, Seo DM. Microplate identification system of Enterobacteriaceae. Korean J Clin Microbiol. 1999; 2:135–43.
9. Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing: 16th informational supplement. Document M100-S16. Wayne, PA; CLSI,. 2006.
10. Bauernfeind A, Stemplinger I, Jungwirth R, Giamarellou H. Characterization of the plasmidic β-lactamase CMY-2, which is responsible for cephamycin resistance. Antimicrob Agents Chemother. 1996; 40:221–4.
11. Queenan AM, Jenkins S, Bush K. Cloning and biochemical characterization of FOX-5, an AmpC-type plasmid-encoded β-lactamase from a New York City Klebsiella pneumoniae clinical isolate. Antimicrob Agents Chemother. 2001; 45:3189–94.
12. Walther-Rasmussen J, H⊘iby N. Plasmid-borne AmpC β-lactamases. Can J Microbiol. 2002; 48:479–93.
Article
13. Bauernfeind A, Hohl P, Schneider I, Jungwirth R, Frei R. Escherichia coli producing a cephamycinase (CMY-2) from a patient from the Libyan-Tunisian border region. Clin Microbiol Infect. 1998; 4:168–70.
14. Coudron PE, Hanson ND, Climo MW. Occurrence of extended-spectrum and AmpC beta-lactamases in bloodstream isolates of Klebsiella pneumoniae: isolates harbor plasmid-mediated FOX-5 and ACT-1 AmpC beta-lactamases. J Clin Microbiol. 2003; 41:772–7.
15. Bradford PA, Urban C, Mariano N, Projan SJ, Rahal JJ, Bush K. Imipenem resistance in Klebsiella pneumoniae is associated with the combination of ACT-1, a plasmid-mediated AmpC β-lactamase, and the loss of an outer membrane protein. Antimicrob Agents Chemother. 1997; 41:563–9.
16. Nadjar D, Rouveau M, Verdet C, Donay L, Herrmann J, Lagrange PH, et al. Outbreak of Klebsiella pneumoniae producing transferable AmpC-type β-lactamase (ACC-1) originating from Hafnia alvei. FEMS Microbiol Lett. 2000; 187:35–40.
17. Coudron PE. Inhibitor-based methods for detection of plasmid-mediated AmpC β-lactamases in Klebsiella spp., Escherichia coli, and Proteus mirabilis. J Clin Microbiol. 2005; 43:4163–7.
18. Song W, Kim JS, Kim HS, Yong D, Jeong SH, Park MJ, et al. Increasing trend in the prevalence of plasmid-mediated AmpC β-lactamases in Enterobacteriaceae lacking chromosomal ampC gene at a Korean university hospital from 2002 to 2004. Diagn Microbiol Infect Dis. 2006; 55:219–24.
19. Lee K, Hong SG, Park YJ, Lee HS, Song W, Jeong J, et al. Evaluation of phenotypic screening methods for detecting plasmid-mediated AmpC β-lactamases-producing isolates of Escherichia coli and Klebsiella pneumoniae. Diagn Microbiol Infect Dis. 2005; 53:319–23.
20. Black JA, Moland ES, Thomson KS. AmpC disk test for detection of plasmid-mediated AmpC β-lactamases in Enterobacteriaceae lacking chromosomal AmpC β-lactamases. J Clin Microbiol. 2005; 43:3110–3.
21. Pérez-Pérez FJ, Hanson ND. Detection of plasmid-mediated AmpC β-lactamase genes in clinical isolates by using multiplex PCR. J Clin Microbiol. 2002; 40:2153–62.
Article
22. Smith Moland E, Hanson ND, Herrera VL, Black JA, Lockhart TJ, Hossain A, et al. Plasmid-mediated, carbapenem-hydrolysing β-lactamase, KPC-2, in Klebsiella pneumoniae isolates. J Antimicrob Chemother. 2003; 51:711–4.
23. Jeong SH, Song W, Park MJ, Kim JS, Kim HS, Bae IK, et al. Boronic acid disk tests for identification of extended-spectrum β-lactamase production in clinical isolates of Enterobacteriaceae producing chromosomal AmpC β-lactamases. Int J Antimicrob Agents. 2008; 31:467–71.
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