1887

Abstract

Spontaneous multidrug-resistant (Mdr) mutants of strain ECL8 arose at a frequency of 2-2 ° 10 and showed increased resistance to a range of unrelated antibiotics, including chloramphenicol, tetracycline, nalidixic acid, ampicillin, norfloxacin, trimethoprim and puromycin. A chromosomal fragment from one such mutant was cloned, and found to confer an Mdr phenotype on K12 cells that was essentially identical to that of the mutant. Almost complete loss of the OmpF porin in the transformant, and of the corresponding porin in the mutant, was observed. The presence of the Mdr mutation in or the cloned antibiotic ) locus in also resulted in active efflux of tetracycline, and increased active efflux of chloramphenicol. After transformation of a plasmid into expression of chloramphenicol resistance occurred later than expression of resistance to tetracycline, puromycin, trimethoprim and nalidixic acid. The gene was localized and sequenced. It encodes a putative positive transcriptional activator that is weakly related to the MarA and SoxS proteins. A gene was also found to be present in an fragment that has previously been shown to confer an Mdr phenotype, and it appears that , rather than the gene identified in that study, is responsible for multidrug resistance. The gene from the wild-type was identical to that of the mutant strain and also conferred an Mdr phenotype on indicating that the mutation responsible for Mdr in had not been cloned.

Loading

Article metrics loading...

/content/journal/micro/10.1099/13500872-141-8-1909
1995-08-01
2024-03-29
Loading full text...

Full text loading...

/deliver/fulltext/micro/141/8/mic-141-8-1909.html?itemId=/content/journal/micro/10.1099/13500872-141-8-1909&mimeType=html&fmt=ahah

References

  1. Abdel-Sayed S. 1987; Transport of chloramphenicol into sensitive strains of Escherichia coli and Pseudomonas aeruginosa. J Antimicrob Chemother 19:7–20
    [Google Scholar]
  2. Amabile-Cuevas C.F., Demple B. 1991; Molecular characterization of the soxRS genes of Escherichia coli: two genes control a superoxide stress regulon. Nucleic Acids Res 19:4479–4484
    [Google Scholar]
  3. Ariza R.R., Cohen S.P., Bachhawat N., Levy S.B., Demple B. 1994; Repressor mutations in the marRAB operon that activate oxidative stress genes and multiple antibiotic resistance in Escherichia coli. J Bacteriol 176:143–148
    [Google Scholar]
  4. Berger E.A., Heppel L.A. 1974; Different mechanisms of energy coupling for the shock-sensitive and shock-resistant amino acid permeases of Escherichia coli. J Biol Chem 249:7747–7755
    [Google Scholar]
  5. Birnboim H.C., Doly J. 1979; A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res 7:1513–1523
    [Google Scholar]
  6. Cohen S.P., Hooper D.C., Wolfson J.S., Souza K. S., McMurry L.M., Levy S.B. 1988a; Endogenous active efflux of norfloxacin in susceptible Escherichia coli. Antimicrob Agents Chemother 32:1187–1191
    [Google Scholar]
  7. Cohen S.P., McMurry L.M., Levy S.B. 1988b; mar A locus causes decreased expression of OmpF porin in multiple-antibiotic-resistant (Mar) mutants of Escherichia coli. J Bacteriol 170:5416–5422
    [Google Scholar]
  8. Cohen S.P., McMurry L.M., Hooper D.C., Wolfson J.S., Levy S.B. 1989; Cross-resistance to fluoroquinolones in multiple- antibiotic-resistant (Mar) Escherichia coli selected by tetracycline or chloramphenicol: decreased drug accumulation associated with membrane changes in addition to OmpF reduction. Antimicrob Agents Chemother 33:1318–1325
    [Google Scholar]
  9. Cohen S.P., Hachler H., Levy S.B. 1993a; Genetic and functional analysis of the multiple antibiotic resistance (mar) locus in Escherichia coli. J Bacteriol 175:1484–1492
    [Google Scholar]
  10. Cohen S.P., Yan W., Levy S.B. 1993b; A multidrug resistance regulatory chromosomal locus is widespread among enteric bacteria. J Infect Dis 168:484–488
    [Google Scholar]
  11. Dang P., Gutmann L., Quentin C., Williamson R., Collatz E. 1988; Some properties of Serratia marcescens, Salmonella paratyphi A, and Enterobacter cloacae with non-enzyme-dependent multiple resistance to β-lactam antibiotics, aminoglycosides, and quinolones. Rev Infect Dis 10:899–904
    [Google Scholar]
  12. Dayhoff M.O., Schwartz R.M., Orcott B.L. 1978 In Atlas Of Protein Sequence and Structure 5 suppl. 3 pp. 345–352 Edited by Dayhoff M. O. Washington, DC: National Biomedical Research Foundation;
    [Google Scholar]
  13. Devereux J., Haeberli P., Smithies O. 1984; A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res 12:387–395
    [Google Scholar]
  14. Dodd I.B., Egan J.B. 1987; Systematic method for the detection of potential λ Cro-like DNA binding regions in proteins. J Mol Biol 194:557–564
    [Google Scholar]
  15. Forage R.G., Lin E.C.C. 1982; DHA system mediating aerobic and anaerobic dissimilation of glycerol in Klebsiella pneumoniae NCIB 418. J Bacteriol 151:591–599
    [Google Scholar]
  16. Gambino L, Miller P.F. 1993; Overexpression of the MarA positive regulator is sufficient to confer multiple antibiotic resistance in Escherichia coli. J Bacteriol 175:2888–2894
    [Google Scholar]
  17. George A.M., Levy S.B. 1983a; Ampliflable resistance to tetracycline, chloramphenicol and other antibiotics in Escherichia coli: involvement of a non-plasmid determined efflux of tetracycline. J Bacteriol 155:531–540
    [Google Scholar]
  18. George A.M., Levy S.B. 1983b; Gene in the major cotransduction gap of the Escherichia coli K-12 linkage map required for the expression of chromosomal resistance to tetracycline and other antibiotics. J Bacteriol 155:541–548
    [Google Scholar]
  19. Greenberg J.T., Chou J.H., Monach P.A., Demple B. 1991; Activation of oxidative stress genes by mutations at the soxQ/cfxB/marA locus of Escherichia coli. J Bacteriol 173:4433–4439
    [Google Scholar]
  20. Gutmann L, Williamson R., Moreau R., Kitzis M.-D., Collatz E., Acar J.F., Goldstein F.W. 1985; Cross-resistance to nalidixic acid, trimethoprim, and chloramphenicol associated with alterations in the outer membrane proteins of Klebsiella, Enterobacter, and Serratia. J Infect Dis 151:501–507
    [Google Scholar]
  21. Hachler H., Cohen S.P., Levy S.B. 1991; mar A, a regulated locus which controls expression of chromosomal multiple antibiotic resistance in Escherichia coli. J Bacteriol 173:5532–5538
    [Google Scholar]
  22. Hirai K., Aoyama H., Suzue S., Irikura T., Iyobe S., Mitsuhashi S. 1986; Isolation and characterization of norfloxacin-resistant mutants of Escherichia coli K-12. Antimicrob Agents Chemother 30:248–253
    [Google Scholar]
  23. Hooper D.C., Wolfson J.S., Souza K.S., Ng E.Y., McHugh G.L., Swartz M.N. 1989; Mechanisms of quinolone resistance in Escherichia coli: characterization of nfxB and cfxB, two mutant resistance loci decreasing norfloxacin accumulation. Antimicrob Agents Chemother 33:283–290
    [Google Scholar]
  24. Hooper D.C., Wolfson J.S., Bozza M.A., Ng E.Y. 1992; Genetics and regulation of outer membrane protein expression by quinolone resistance loci nfxB, nfxC, and cfxB. Antimicrob Agents Chemother 36:1151–1154
    [Google Scholar]
  25. Higgins D.G., Bleasby A.J., Fuchs R. 1992; ClustalV: improved software for multiple sequence alignment. Comput Appl Biosci 8:189–191
    [Google Scholar]
  26. Innis M.A., Gelfand D.H. 1990; Optimization of RCR. In PCR Protocols: a Guide to Methods and Applications Edited by Innis M. A., Gelfand D. H., Asninsky J. J., White T. J. San Diego, CA: Academic Press;
    [Google Scholar]
  27. Inokuchi K., Itoh M., Mizushima S. 1985; Domains involved in osmoregulation of the ompF gene in Escherichia coli. T Bacteriol 164:585–590
    [Google Scholar]
  28. Ishida H., Fuziwara H., Kaibori Y., Horiuchi T., Sato K., Osada Y. 1995; Cloning of the multidrug resistance gene pqrA from Proteus vulgaris. Antimicrob Agents Chemother 39:453–457
    [Google Scholar]
  29. Komatsu T., Ohta M., Kido N., Arakawa Y., Ito H., Mizuno T., Kato N. 1990; Molecular characterization of an Enterobacter cloacae gene (romA) which pleiotropically inhibits the expression of Escherichia coli outer membrane proteins. J Bacteriol 172:4082–4089
    [Google Scholar]
  30. Komatsu T., Ohta M., Kido N., Arakawa Y., Ito H., Kato N. 1991; Increased resistance to multiple drugs by introduction of the Enterobacter cloacae romA gene into OmpF porin-deficient mutants of Escherichia coli K-12. Antimicrob Agents Chemother 35:2155–2158
    [Google Scholar]
  31. Laemmli U.K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685
    [Google Scholar]
  32. Lambert B., LePecq J.-B. 1984; Effect of mutation, electric membrane potential, and metabolic inhibitors on the accessibility of nucleic acids to ethidium bromide in Escherichia coli cells. Biochemistry 23:166–176
    [Google Scholar]
  33. Lea D.E., Coulsen C.A. 1949; The distribution of the number of mutants in bacterial populations. J Genet 49:264–285
    [Google Scholar]
  34. Levy S.B. 1992; Active efflux mechanisms for antimicrobial resistance. Antimicrob Agents Chemother 36:695–703
    [Google Scholar]
  35. Maniatis T., Fritsch E.F., Sambrook J. 1982 Molecular Cloning: a Laboratory Manual Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  36. Markwell M.A.K., Haas S.M., Bieber L.L., Tolbert N.E. 1978; A modification of the Lowry procedure to simplify protein determination in membrane and lipoprotein samples. Anal Biochem 87:206–210
    [Google Scholar]
  37. McMurry L.M., Petrucci R.E. Jr Levy S. B. 1980; Active efflux of tetracycline encoded by four genetically different tetracycline resistance determinants in Escherichia coli. Proc Natl Acad Sci USA 77:3974–3977
    [Google Scholar]
  38. McMurry L.M., Aronson D.A., Levy S.B. 1983; Susceptible Escherichia coli cells can actively excrete tetracyclines. Antimicrob Agents Chemother 24:544–551
    [Google Scholar]
  39. McMurry L.M., Park B.H., Burdett V., Levy S.B. 1987; Energy-dependent efflux mediated by class L (TetL) tetracycline resistance determinant from Streptococci. Antimicrob Agents Chemother 31:1648–1650
    [Google Scholar]
  40. McMurry L.M., George A.M., Levy S.B. 1994; Active efflux of chloramphenicol in susceptible Escherichia coli strains and in multiple-antibiotic-resistant (Mar) mutants. Antimicrob Agents Chemother 38:542–546
    [Google Scholar]
  41. Miller P.F., Gambino L.F., Sulavik M.C., Gracheck S.J. 1994; Genetic relationship between soxRS and mar loci in promoting multiple antibiotic resistance in Escherichia coli. Antimicrob Agents Chemother 38:1773–1779
    [Google Scholar]
  42. Nikaido H. 1989; Outer membrane barrier as a mechanism of antimicrobial resistance. Antimicrob Agents Chemother 33:1831–1836
    [Google Scholar]
  43. Park B.H., Levy S.B. 1988; The cryptic tetracycline resistance determinant on Tn4400 mediates tetracycline degradation as well as tetracycline efflux. Antimicrob Agents Chemother 32:1797–1800
    [Google Scholar]
  44. Pfeiffer L. 1889; Ueber einen neuen Kapsel-Bacillus. Z Hyg 6:145–150
    [Google Scholar]
  45. Pugsley A.P., Schnaitman C.A. 1978; Identification of three genes controlling production of new outer membrane pore proteins in Escherichia coli K-12. J Bacteriol 135:1118–1129
    [Google Scholar]
  46. Ramos J.L., Rojo F., Zhou L., Timmis K.N. 1990; A family of positive regulators related to the Pseudomonas putida TOL plasmid XylS and the Escherichia coli AraC activators. Nucleic Acids Res 18:2149–2152
    [Google Scholar]
  47. Rosen B.P., Kashket E.R. 1978; Energetics of active transport. In Bacterial Transport pp. 559–620 Edited by Rosen B.P. New York: Marcel Dekker;
    [Google Scholar]
  48. Sanders C.C., Sanders W.E. Jr Goering R.V., Werner V. 1984; Selection of multiple antibiotic resistance by quinolones, β-lactams, and aminoglycosides with special reference to cross-resistance between unrelated drug classes. Antimicrob Agents Chemother 26:797–801
    [Google Scholar]
  49. Silhavy T.J., Berman M.L., Enquist L.W. 1984 Experiments with Gene Fusions Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  50. Szybalski W., Bryson V. 1952; Genetic studies on microbial cross resistance to toxic agents. I. Cross resistance of Escherichia coli to fifteen antibiotics. J Bacteriol 64:489–499
    [Google Scholar]
  51. Then R.L., Angehrn P. 1986; Multiply resistant mutants of Enterobacter cloacae selected by β-lactam antibiotics. Antimicrob Agents Chemother 30:684–688
    [Google Scholar]
  52. Traub W.H., Kleber I. 1977; Selected and spontaneous variants of Serratia marcescens with combined resistance against chloramphenicol, nalidixic acid, and trimethoprim. Chemotherapy 23:436–451
    [Google Scholar]
  53. Williams Smith H. 1976; Mutants of Klebsiella pneumoniae resistant to several antibiotics. Nature 259:307–308
    [Google Scholar]
  54. Wu J., Weiss B. 1991; Two divergently transcribed genes, roxR and soxS, control a superoxide response regulon of Escherichia coli. J Bacteriol 173:2864–2871
    [Google Scholar]
  55. Yanisch-Perron C., Vieira J., Messing J. 1985; Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mpl8 and pUC19 vectors. Gene 33:103–119
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/13500872-141-8-1909
Loading
/content/journal/micro/10.1099/13500872-141-8-1909
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error