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. 1989 Apr;33(4):551–559. doi: 10.1128/aac.33.4.551

Development of a DNA probe from the deoxyribonucleotide sequence of a 3-N-aminoglycoside acetyltransferase [AAC(3)-I] resistance gene.

F C Tenover 1, K L Phillips 1, T Gilbert 1, P Lockhart 1, P J O'Hara 1, J J Plorde 1
PMCID: PMC172477  PMID: 2658795

Abstract

The aacC1 gene encoding the 3-N-aminoglycoside acetyltransferase [AAC(3)-I] was cloned from enteric plasmid pJR88, and its deoxyribonucleotide sequence was determined. Significant nucleotide homology was noted in the region extending from the proposed -35 sequences through the first 59 base pairs of the aacC1 gene open reading frame (ORF) and the upstream flanking regions and ORFs of several other antibiotic resistance genes. Sequences were noted to be homologous with the 6'-N-aminoglycoside acetyltransferase [AAC(6')-I], 2''-O-aminoglycoside adenylyltransferase [AAD(2'')], and 3''-O-aminoglycoside adenylyltransferase [AAD(3'')] resistance genes; the OXA-1, OXA-2, and PSE-2 beta-lactamase genes; and several dihydrofolate reductase genes. Small regions of homology were noted in the 3'-flanking regions of these resistance genes as well. A DNA probe for the aacC1 gene was selected from the nucleotide sequence information and was tested against a series of genetically and enzymatically defined strains. The probe, which proved specific for the aacC1 gene, was then tested against a series of 58 gentamicin-susceptible and 219 gentamicin-resistant gram-negative bacilli isolated from patients at the Seattle Veterans Administration Medical Center. Only six clinical isolates were noted to carry the aacC1 gene. Each was resistant to gentamicin but susceptible to kanamycin, tobramycin, and amikacin. The presence of homologous regions of DNA at both the 3' and 5' ends of the aacC1 gene reinforces the importance of choosing probes from within the ORFs of genes and of avoiding flanking sequences. When the homology with other sequences extends into the ORF, as it does with the aacC1 gene, development of a specific probe may require determination of the nucleotide sequence.

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Selected References

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  1. Allmansberger R., Bräu B., Piepersberg W. Genes for gentamicin-(3)-N-acetyl-transferases III and IV. II. Nucleotide sequences of three AAC(3)-III genes and evolutionary aspects. Mol Gen Genet. 1985;198(3):514–520. doi: 10.1007/BF00332949. [DOI] [PubMed] [Google Scholar]
  2. Arthur M., Andremont A., Courvalin P. Distribution of erythromycin esterase and rRNA methylase genes in members of the family Enterobacteriaceae highly resistant to erythromycin. Antimicrob Agents Chemother. 1987 Mar;31(3):404–409. doi: 10.1128/aac.31.3.404. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Balbás P., Soberón X., Merino E., Zurita M., Lomeli H., Valle F., Flores N., Bolivar F. Plasmid vector pBR322 and its special-purpose derivatives--a review. Gene. 1986;50(1-3):3–40. doi: 10.1016/0378-1119(86)90307-0. [DOI] [PubMed] [Google Scholar]
  4. Bauer A. W., Kirby W. M., Sherris J. C., Turck M. Antibiotic susceptibility testing by a standardized single disk method. Am J Clin Pathol. 1966 Apr;45(4):493–496. [PubMed] [Google Scholar]
  5. Biddlecome S., Haas M., Davies J., Miller G. H., Rane D. F., Daniels P. J. Enzymatic modification of aminoglycoside antibiotics: a new 3-N-acetylating enzyme from a Pseudomonas aeruginosa isolate. Antimicrob Agents Chemother. 1976 Jun;9(6):951–955. doi: 10.1128/aac.9.6.951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Biggin M. D., Gibson T. J., Hong G. F. Buffer gradient gels and 35S label as an aid to rapid DNA sequence determination. Proc Natl Acad Sci U S A. 1983 Jul;80(13):3963–3965. doi: 10.1073/pnas.80.13.3963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bilofsky H. S., Burks C., Fickett J. W., Goad W. B., Lewitter F. I., Rindone W. P., Swindell C. D., Tung C. S. The GenBank genetic sequence databank. Nucleic Acids Res. 1986 Jan 10;14(1):1–4. doi: 10.1093/nar/14.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Brzezinska M., Benveniste R., Davies J., Daniels P. J., Weinstein J. Gentamicin resistance in strains of Pseudomonas aeruginosa mediated by enzymatic N-acetylation of the deoxystreptamine moiety. Biochemistry. 1972 Feb 29;11(5):761–765. doi: 10.1021/bi00755a013. [DOI] [PubMed] [Google Scholar]
  9. Bräu B., Pilz U., Piepersberg W. Genes for gentamicin-(3)-N-acetyltransferases III and IV: I. Nucleotide sequence of the AAC(3)-IV gene and possible involvement of an IS140 element in its expression. Mol Gen Genet. 1984;193(1):179–187. doi: 10.1007/BF00327434. [DOI] [PubMed] [Google Scholar]
  10. Bullock D. W., Bidwell J. L., Reeves D. S., White L. O., Turner A., Speller D. C., Wilkinson P. J. Outbreaks of hospital infection in southwest England caused by gentamicin-resistant Serratia marcescens. J Hosp Infect. 1982 Sep;3(3):263–273. doi: 10.1016/0195-6701(82)90045-7. [DOI] [PubMed] [Google Scholar]
  11. Cameron F. H., Groot Obbink D. J., Ackerman V. P., Hall R. M. Nucleotide sequence of the AAD(2'') aminoglycoside adenylyltransferase determinant aadB. Evolutionary relationship of this region with those surrounding aadA in R538-1 and dhfrII in R388. Nucleic Acids Res. 1986 Nov 11;14(21):8625–8635. doi: 10.1093/nar/14.21.8625. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Chinault A. C., Blakesley V. A., Roessler E., Willis D. G., Smith C. A., Cook R. G., Fenwick R. G., Jr Characterization of transferable plasmids from Shigella flexneri 2a that confer resistance to trimethoprim, streptomycin, and sulfonamides. Plasmid. 1986 Mar;15(2):119–131. doi: 10.1016/0147-619x(86)90048-x. [DOI] [PubMed] [Google Scholar]
  13. Cohen S. N., Chang A. C., Hsu L. Nonchromosomal antibiotic resistance in bacteria: genetic transformation of Escherichia coli by R-factor DNA. Proc Natl Acad Sci U S A. 1972 Aug;69(8):2110–2114. doi: 10.1073/pnas.69.8.2110. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Davies J., Smith D. I. Plasmid-determined resistance to antimicrobial agents. Annu Rev Microbiol. 1978;32:469–518. doi: 10.1146/annurev.mi.32.100178.002345. [DOI] [PubMed] [Google Scholar]
  15. Dickie P., Bryan L. E., Pickard M. A. Effect of enzymatic adenylylation on dihydrostreptomycin accumulation in Escherichia coli carrying an R-factor: model explaining aminoglycoside resistance by inactivating mechanisms. Antimicrob Agents Chemother. 1978 Oct;14(4):569–580. doi: 10.1128/aac.14.4.569. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Flensburg J., Steen R. Nucleotide sequence analysis of the trimethoprim resistant dihydrofolate reductase encoded by R plasmid R751. Nucleic Acids Res. 1986 Jul 25;14(14):5933–5933. doi: 10.1093/nar/14.14.5933. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Fling M. E., Kopf J., Richards C. Nucleotide sequence of the transposon Tn7 gene encoding an aminoglycoside-modifying enzyme, 3"(9)-O-nucleotidyltransferase. Nucleic Acids Res. 1985 Oct 11;13(19):7095–7106. doi: 10.1093/nar/13.19.7095. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. George D. G., Barker W. C., Hunt L. T. The protein identification resource (PIR). Nucleic Acids Res. 1986 Jan 10;14(1):11–15. doi: 10.1093/nar/14.1.11. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Gootz T. D., Tenover F. C., Young S. A., Gordon K. P., Plorde J. J. Comparison of three DNA hybridization methods for detection of the aminoglycoside 2"-O-adenylyltransferase gene in clinical bacterial isolates. Antimicrob Agents Chemother. 1985 Jul;28(1):69–73. doi: 10.1128/aac.28.1.69. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Haas M. J., Dowding J. E. Aminoglycoside-modifying enzymes. Methods Enzymol. 1975;43:611–628. doi: 10.1016/0076-6879(75)43124-x. [DOI] [PubMed] [Google Scholar]
  21. 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]
  22. Hamm G. H., Cameron G. N. The EMBL data library. Nucleic Acids Res. 1986 Jan 10;14(1):5–9. doi: 10.1093/nar/14.1.5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hollingshead S., Vapnek D. Nucleotide sequence analysis of a gene encoding a streptomycin/spectinomycin adenylyltransferase. Plasmid. 1985 Jan;13(1):17–30. doi: 10.1016/0147-619x(85)90052-6. [DOI] [PubMed] [Google Scholar]
  24. Hsiang M. W., White T. J., Davies J. E. NH2-terminal sequence of the aminoglycoside acetyltransferase (3)-I mediated by plasmid RIP 135. FEBS Lett. 1978 Aug 1;92(1):97–99. doi: 10.1016/0014-5793(78)80730-3. [DOI] [PubMed] [Google Scholar]
  25. 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]
  26. Huovinen S., Huovinén P., Jacoby G. A. Detection of plasmid-mediated beta-lactamases with DNA probes. Antimicrob Agents Chemother. 1988 Feb;32(2):175–179. doi: 10.1128/aac.32.2.175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. John J. F., Jr, McNeill W. F. Characteristics of Serratia marcescens containing a plasmid coding for gentamicin resistance in nosocomial infections. J Infect Dis. 1981 Jun;143(6):810–817. doi: 10.1093/infdis/143.6.810. [DOI] [PubMed] [Google Scholar]
  28. Kagan S. A., Davies J. E. Enzymatic modification of aminocyclitol antibiotics: mutations affecting the expression of aminocyclitol acetyltransferase-3. Plasmid. 1980 May;3(3):312–318. doi: 10.1016/0147-619x(80)90043-8. [DOI] [PubMed] [Google Scholar]
  29. Kratz J., Schmidt F., Wiedemann B. Characterization of Tn2411 and Tn2410, two transposons derived from R-plasmid R1767 and related to Tn2603 and Tn21. J Bacteriol. 1983 Sep;155(3):1333–1342. doi: 10.1128/jb.155.3.1333-1342.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Lipman D. J., Pearson W. R. Rapid and sensitive protein similarity searches. Science. 1985 Mar 22;227(4693):1435–1441. doi: 10.1126/science.2983426. [DOI] [PubMed] [Google Scholar]
  31. Martin P., Trieu-Cuot P., Courvalin P. Nucleotide sequence of the tetM tetracycline resistance determinant of the streptococcal conjugative shuttle transposon Tn1545. Nucleic Acids Res. 1986 Sep 11;14(17):7047–7058. doi: 10.1093/nar/14.17.7047. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Maruyama T., Gojobori T., Aota S., Ikemura T. Codon usage tabulated from the GenBank genetic sequence data. Nucleic Acids Res. 1986;14 (Suppl):r151–r197. doi: 10.1093/nar/14.suppl.r151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  34. Mills D. R., Kramer F. R. Structure-independent nucleotide sequence analysis. Proc Natl Acad Sci U S A. 1979 May;76(5):2232–2235. doi: 10.1073/pnas.76.5.2232. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Novotný J., Auffray C. A program for prediction of protein secondary structure from nucleotide sequence data: application to histocompatibility antigens. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):243–255. doi: 10.1093/nar/12.1part1.243. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Rubens C. E., McNeill W. F., Farrar W. E., Jr Evolution of multiple-antibiotic-resistance plasmids mediated by transposable plasmid deoxyribonucleic acid sequences. J Bacteriol. 1979 Nov;140(2):713–719. doi: 10.1128/jb.140.2.713-719.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Sancar A., Hack A. M., Rupp W. D. Simple method for identification of plasmid-coded proteins. J Bacteriol. 1979 Jan;137(1):692–693. doi: 10.1128/jb.137.1.692-693.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Schmidt F. The role of insertions, deletions, and substitutions in the evolution of R6 related plasmids encoding aminoglycoside transferase ANT-(2"). Mol Gen Genet. 1984;194(1-2):248–259. doi: 10.1007/BF00383524. [DOI] [PubMed] [Google Scholar]
  40. Shimizu K., Kumada T., Hsieh W. C., Chung H. Y., Chong Y., Hare R. S., Miller G. H., Sabatelli F. J., Howard J. Comparison of aminoglycoside resistance patterns in Japan, Formosa, and Korea, Chile, and the United States. Antimicrob Agents Chemother. 1985 Aug;28(2):282–288. doi: 10.1128/aac.28.2.282. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Sivasubramanian N., Nayak D. P. Mutational analysis of the signal-anchor domain of influenza virus neuraminidase. Proc Natl Acad Sci U S A. 1987 Jan;84(1):1–5. doi: 10.1073/pnas.84.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Sundström L., Rådström P., Swedberg G., Sköld O. Site-specific recombination promotes linkage between trimethoprim- and sulfonamide resistance genes. Sequence characterization of dhfrV and sulI and a recombination active locus of Tn21. Mol Gen Genet. 1988 Aug;213(2-3):191–201. doi: 10.1007/BF00339581. [DOI] [PubMed] [Google Scholar]
  43. Swift G., McCarthy B. J., Heffron F. DNA sequence of a plasmid-encoded dihydrofolate reductase. Mol Gen Genet. 1981;181(4):441–447. doi: 10.1007/BF00428733. [DOI] [PubMed] [Google Scholar]
  44. Tabor S., Richardson C. C. DNA sequence analysis with a modified bacteriophage T7 DNA polymerase. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4767–4771. doi: 10.1073/pnas.84.14.4767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Taylor D. E. Plasmid-mediated tetracycline resistance in Campylobacter jejuni: expression in Escherichia coli and identification of homology with streptococcal class M determinant. J Bacteriol. 1986 Mar;165(3):1037–1039. doi: 10.1128/jb.165.3.1037-1039.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Tenover F. C. Diagnostic deoxyribonucleic acid probes for infectious diseases. Clin Microbiol Rev. 1988 Jan;1(1):82–101. doi: 10.1128/cmr.1.1.82. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Tenover F. C., Filpula D., Phillips K. L., Plorde J. J. Cloning and sequencing of a gene encoding an aminoglycoside 6'-N-acetyltransferase from an R factor of Citrobacter diversus. J Bacteriol. 1988 Jan;170(1):471–473. doi: 10.1128/jb.170.1.471-473.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Tenover F. C., Gootz T. D., Gordon K. P., Tompkins L. S., Young S. A., Plorde J. J. Development of a DNA probe for the structural gene of the 2"-O-adenyltransferase aminoglycoside-modifying enzyme. J Infect Dis. 1984 Nov;150(5):678–687. doi: 10.1093/infdis/150.5.678. [DOI] [PubMed] [Google Scholar]
  49. Tenover F. C. Studies of antimicrobial resistance genes using DNA probes. Antimicrob Agents Chemother. 1986 May;29(5):721–725. doi: 10.1128/aac.29.5.721. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Thomas F. E., Jackson R. T., Melly A., Alford R. H. Sequential hospitalwide outbreaks of resistant Serratia and Klebsiella infections. Arch Intern Med. 1977 May;137(5):581–584. [PubMed] [Google Scholar]
  51. Tran van Nhieu G., Collatz E. Primary structure of an aminoglycoside 6'-N-acetyltransferase AAC(6')-4, fused in vivo with the signal peptide of the Tn3-encoded beta-lactamase. J Bacteriol. 1987 Dec;169(12):5708–5714. doi: 10.1128/jb.169.12.5708-5714.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Wahl G. M., Stern M., Stark G. R. Efficient transfer of large DNA fragments from agarose gels to diazobenzyloxymethyl-paper and rapid hybridization by using dextran sulfate. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3683–3687. doi: 10.1073/pnas.76.8.3683. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Weinert T. A., Schaus N. A., Grindley N. D. Insertion sequence duplication in transpositional recombination. Science. 1983 Nov 18;222(4625):755–765. doi: 10.1126/science.6314502. [DOI] [PubMed] [Google Scholar]
  54. Witchitz J. L. Plasmid-mediated gentamicin resistance not associated with kanamycin resistance in Enterobacteriaceae. J Antibiot (Tokyo) 1972 Oct;25(10):622–624. doi: 10.7164/antibiotics.25.622. [DOI] [PubMed] [Google Scholar]
  55. Young S. A., Tenover F. C., Gootz T. D., Gordon K. P., Plorde J. J. Development of two DNA probes for differentiating the structural genes of subclasses I and II of the aminoglycoside-modifying enzyme 3'-aminoglycoside phosphotransferase. Antimicrob Agents Chemother. 1985 May;27(5):739–744. doi: 10.1128/aac.27.5.739. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Zolg J. W., Hänggi U. J. Characterization of a R plasmid-associated, trimethoprim-resistant dihydrofolate reductase and determination of the nucleotide sequence of the reductase gene. Nucleic Acids Res. 1981 Feb 11;9(3):697–710. doi: 10.1093/nar/9.3.697. [DOI] [PMC free article] [PubMed] [Google Scholar]

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