Skip to main content
NCBI home page
Journal of Clinical Microbiology logoLink to Journal of Clinical Microbiology
. 1994 Aug;32(8):1866–1869. doi: 10.1128/jcm.32.8.1866-1869.1994

Development of enzyme-labeled oligonucleotide probe for detection of mecA gene in methicillin-resistant Staphylococcus aureus.

M Shimaoka 1, M Yoh 1, A Segawa 1, Y Takarada 1, K Yamamoto 1, T Honda 1
PMCID: PMC263893  PMID: 7989534

Abstract

A DNA hybridization method with an enzyme-labeled oligonucleotide probe (mecA-ELONP) was developed to detect the methicillin-resistant gene (mecA) in methicillin-resistant Staphylococcus aureus. For rapid identification, bacterial colonies were transferred from agar plates directly onto nylon membranes. Lysis of cells, denaturation of DNA, and hybridization were performed on the membranes. These procedures required only 3 h for completion. The results obtained by this test closely corresponded with those obtained by determining the MICs of oxacillin against S. aureus. The results of the mecA-ELONP also correlated well with those of a commercially available PCR test. Thus, mecA-ELONP proved to be a reliable and convenient method for the rapid identification of methicillin-resistant S. aureus, which could be useful in clinical microbiology laboratories.

Full text

PDF
1866

Selected References

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

  1. Archer G. L., Pennell E. Detection of methicillin resistance in staphylococci by using a DNA probe. Antimicrob Agents Chemother. 1990 Sep;34(9):1720–1724. doi: 10.1128/aac.34.9.1720. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brumfitt W., Hamilton-Miller J. Methicillin-resistant Staphylococcus aureus. N Engl J Med. 1989 May 4;320(18):1188–1196. doi: 10.1056/NEJM198905043201806. [DOI] [PubMed] [Google Scholar]
  3. Chambers H. F., Archer G., Matsuhashi M. Low-level methicillin resistance in strains of Staphylococcus aureus. Antimicrob Agents Chemother. 1989 Apr;33(4):424–428. doi: 10.1128/aac.33.4.424. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Gerberding J. L., Miick C., Liu H. H., Chambers H. F. Comparison of conventional susceptibility tests with direct detection of penicillin-binding protein 2a in borderline oxacillin-resistant strains of Staphylococcus aureus. Antimicrob Agents Chemother. 1991 Dec;35(12):2574–2579. doi: 10.1128/aac.35.12.2574. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Hackbarth C. J., Chambers H. F. Methicillin-resistant staphylococci: detection methods and treatment of infections. Antimicrob Agents Chemother. 1989 Jul;33(7):995–999. doi: 10.1128/aac.33.7.995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hartman B. J., Tomasz A. Expression of methicillin resistance in heterogeneous strains of Staphylococcus aureus. Antimicrob Agents Chemother. 1986 Jan;29(1):85–92. doi: 10.1128/aac.29.1.85. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Jablonski E., Moomaw E. W., Tullis R. H., Ruth J. L. Preparation of oligodeoxynucleotide-alkaline phosphatase conjugates and their use as hybridization probes. Nucleic Acids Res. 1986 Aug 11;14(15):6115–6128. doi: 10.1093/nar/14.15.6115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Ligozzi M., Rossolini G. M., Tonin E. A., Fontana R. Nonradioactive DNA probe for detection of gene for methicillin resistance in Staphylococcus aureus. Antimicrob Agents Chemother. 1991 Mar;35(3):575–578. doi: 10.1128/aac.35.3.575. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Maidhof H., Reinicke B., Blümel P., Berger-Bächi B., Labischinski H. femA, which encodes a factor essential for expression of methicillin resistance, affects glycine content of peptidoglycan in methicillin-resistant and methicillin-susceptible Staphylococcus aureus strains. J Bacteriol. 1991 Jun;173(11):3507–3513. doi: 10.1128/jb.173.11.3507-3513.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Maple P. A., Hamilton-Miller J. M., Brumfitt W. World-wide antibiotic resistance in methicillin-resistant Staphylococcus aureus. Lancet. 1989 Mar 11;1(8637):537–540. doi: 10.1016/s0140-6736(89)90076-7. [DOI] [PubMed] [Google Scholar]
  11. McDougal L. K., Thornsberry C. The role of beta-lactamase in staphylococcal resistance to penicillinase-resistant penicillins and cephalosporins. J Clin Microbiol. 1986 May;23(5):832–839. doi: 10.1128/jcm.23.5.832-839.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Monzon-Moreno C., Aubert S., Morvan A., Solh N. E. Usefulness of three probes in typing isolates of methicillin-resistant Staphylococcus aureus (MRSA). J Med Microbiol. 1991 Aug;35(2):80–88. doi: 10.1099/00222615-35-2-80. [DOI] [PubMed] [Google Scholar]
  13. Murakami K., Minamide W., Wada K., Nakamura E., Teraoka H., Watanabe S. Identification of methicillin-resistant strains of staphylococci by polymerase chain reaction. J Clin Microbiol. 1991 Oct;29(10):2240–2244. doi: 10.1128/jcm.29.10.2240-2244.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Murakami K., Nomura K., Doi M., Yoshida T. Production of low-affinity penicillin-binding protein by low- and high-resistance groups of methicillin-resistant Staphylococcus aureus. Antimicrob Agents Chemother. 1987 Sep;31(9):1307–1311. doi: 10.1128/aac.31.9.1307. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Nishibuchi M., Arita M., Honda T., Miwatani T. Evaluation of a nonisotopically labeled oligonucleotide probe to detect the heat-stable enterotoxin gene of Escherichia coli by the DNA colony hybridization test. J Clin Microbiol. 1988 Apr;26(4):784–786. doi: 10.1128/jcm.26.4.784-786.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Rossi L., Tonin E., Cheng Y. R., Fontana R. Regulation of penicillin-binding protein activity: description of a methicillin-inducible penicillin-binding protein in Staphylococcus aureus. Antimicrob Agents Chemother. 1985 May;27(5):828–831. doi: 10.1128/aac.27.5.828. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Ryffel C., Tesch W., Birch-Machin I., Reynolds P. E., Barberis-Maino L., Kayser F. H., Berger-Bächi B. Sequence comparison of mecA genes isolated from methicillin-resistant Staphylococcus aureus and Staphylococcus epidermidis. Gene. 1990 Sep 28;94(1):137–138. doi: 10.1016/0378-1119(90)90481-6. [DOI] [PubMed] [Google Scholar]
  18. Song M. D., Wachi M., Doi M., Ishino F., Matsuhashi M. Evolution of an inducible penicillin-target protein in methicillin-resistant Staphylococcus aureus by gene fusion. FEBS Lett. 1987 Aug 31;221(1):167–171. doi: 10.1016/0014-5793(87)80373-3. [DOI] [PubMed] [Google Scholar]
  19. Struelens M. J., Bax R., Deplano A., Quint W. G., Van Belkum A. Concordant clonal delineation of methicillin-resistant Staphylococcus aureus by macrorestriction analysis and polymerase chain reaction genome fingerprinting. J Clin Microbiol. 1993 Aug;31(8):1964–1970. doi: 10.1128/jcm.31.8.1964-1970.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Tamatsukuri S., Yamamoto K., Shibata S., Leaño F., Honda T., Miwatani T. Detection of a heat-labile enterotoxin gene in enterotoxigenic Escherichia coli by densitometric evaluation using highly specific enzyme-linked oligonucleotide probes. Eur J Clin Microbiol Infect Dis. 1991 Dec;10(12):1048–1055. doi: 10.1007/BF01984927. [DOI] [PubMed] [Google Scholar]
  21. Tokue Y., Shoji S., Satoh K., Watanabe A., Motomiya M. Comparison of a polymerase chain reaction assay and a conventional microbiologic method for detection of methicillin-resistant Staphylococcus aureus. Antimicrob Agents Chemother. 1992 Jan;36(1):6–9. doi: 10.1128/aac.36.1.6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Ubukata K., Nakagami S., Nitta A., Yamane A., Kawakami S., Sugiura M., Konno M. Rapid detection of the mecA gene in methicillin-resistant staphylococci by enzymatic detection of polymerase chain reaction products. J Clin Microbiol. 1992 Jul;30(7):1728–1733. doi: 10.1128/jcm.30.7.1728-1733.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Ubukata K., Nonoguchi R., Matsuhashi M., Konno M. Expression and inducibility in Staphylococcus aureus of the mecA gene, which encodes a methicillin-resistant S. aureus-specific penicillin-binding protein. J Bacteriol. 1989 May;171(5):2882–2885. doi: 10.1128/jb.171.5.2882-2885.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Ubukata K., Yamashita N., Konno M. Occurrence of a beta-lactam-inducible penicillin-binding protein in methicillin-resistant staphylococci. Antimicrob Agents Chemother. 1985 May;27(5):851–857. doi: 10.1128/aac.27.5.851. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Yoh M., Miyagi K., Matsumoto Y., Hayashi K., Takarada Y., Yamamoto K., Honda T. Development of an enzyme-labeled oligonucleotide probe for the cholera toxin gene. J Clin Microbiol. 1993 May;31(5):1312–1314. doi: 10.1128/jcm.31.5.1312-1314.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. de Lencastre H., Sá Figueiredo A. M., Urban C., Rahal J., Tomasz A. Multiple mechanisms of methicillin resistance and improved methods for detection in clinical isolates of Staphylococcus aureus. Antimicrob Agents Chemother. 1991 Apr;35(4):632–639. doi: 10.1128/aac.35.4.632. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Clinical Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES