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. 1987 May;31(5):686–690. doi: 10.1128/aac.31.5.686

Chloramphenicol accumulation by Haemophilus influenzae.

J L Burns, A L Smith
PMCID: PMC174815  PMID: 3496847

Abstract

The mechanism of chloramphenicol transport into susceptible strains of Haemophilus influenzae cells has not been reported previously. We examined apparent uptake of chloramphenicol by bacterial cells by using high-pressure liquid chromatography to quantitate drug disappearance from liquid media. Cell-associated chloramphenicol concentration is 1,000-fold greater than the extracellular drug concentration. Under incubation conditions associated with chloramphenicol disappearance from media, cellular protein synthesis was inhibited; however, if accumulation was inhibited, protein synthesis occurred in the presence of the drug. Chloramphenicol uptake appeared saturable (Km = 0.96 mM, Vmax = 0.9 mumol/min per mg of protein) and energy dependent: disappearance from media was markedly decreased by 2,4-dinitrophenol and carbonyl cyanide m-chlorophenylhydrazone, compounds which disrupt the proton motive force. Uptake occurred only in median which can support growth and was dependent upon temperature and pH. Drug accumulation was minimally affected by inhibitors of electron transport or by gentamicin and puromycin, both inhibitors of protein synthesis. The rate of disappearance was inhibited by SCH24893, a fluorinated chloramphenicol analog which also inhibits protein synthesis. We conclude that chloramphenicol accumulation by H. influenzae occurs by energy-dependent transport.

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Selected References

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

  1. Bryan L. E., Kwan S. Roles of ribosomal binding, membrane potential, and electron transport in bacterial uptake of streptomycin and gentamicin. Antimicrob Agents Chemother. 1983 Jun;23(6):835–845. doi: 10.1128/aac.23.6.835. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bryan L. E., Van den Elzen H. M. Streptomycin accumulation in susceptible and resistant strains of Escherichia coli and Pseudomonas aeruginosa. Antimicrob Agents Chemother. 1976 Jun;9(6):928–938. doi: 10.1128/aac.9.6.928. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Burns J. L., Mendelman P. M., Levy J., Stull T. L., Smith A. L. A permeability barrier as a mechanism of chloramphenicol resistance in Haemophilus influenzae. Antimicrob Agents Chemother. 1985 Jan;27(1):46–54. doi: 10.1128/aac.27.1.46. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Catlin B. W., Bendler J. W., 3rd, Goodgal S. H. The type b capsulation locus of Haemophilus influenzae: map location and size. J Gen Microbiol. 1972 May;70(3):411–422. doi: 10.1099/00221287-70-3-411. [DOI] [PubMed] [Google Scholar]
  5. Catlin B. W. Nutritional profiles of Neisseria gonorrhoeae, Neisseria meningitidis, and Neisseria lactamica in chemically defined media and the use of growth requirements for gonococcal typing. J Infect Dis. 1973 Aug;128(2):178–194. doi: 10.1093/infdis/128.2.178. [DOI] [PubMed] [Google Scholar]
  6. Chopra I., Eccles S. J. Diffusion of tetracycline across the outer membrane of Escherichia coli K-12: involvement of protein Ia. Biochem Biophys Res Commun. 1978 Jul 28;83(2):550–557. doi: 10.1016/0006-291x(78)91025-2. [DOI] [PubMed] [Google Scholar]
  7. Eisenberg E. S., Mandel L. J., Kaback H. R., Miller M. H. Quantitative association between electrical potential across the cytoplasmic membrane and early gentamicin uptake and killing in Staphylococcus aureus. J Bacteriol. 1984 Mar;157(3):863–867. doi: 10.1128/jb.157.3.863-867.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. George A. M., Levy S. B. Amplifiable resistance to tetracycline, chloramphenicol, and other antibiotics in Escherichia coli: involvement of a non-plasmid-determined efflux of tetracycline. J Bacteriol. 1983 Aug;155(2):531–540. doi: 10.1128/jb.155.2.531-540.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hedstrom R. C., Crider B. P., Eagon R. G. Comparison of kinetics of active tetracycline uptake and active tetracycline efflux in sensitive and plasmid RP4-containing Pseudomonas putida. J Bacteriol. 1982 Oct;152(1):255–259. doi: 10.1128/jb.152.1.255-259.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hurwitz C., Braun C. B. Temperature-sensitivity of the weak bonds by which chloramphenicol is held in intact cells. Biochim Biophys Acta. 1968 Apr 22;157(2):392–403. doi: 10.1016/0005-2787(68)90093-2. [DOI] [PubMed] [Google Scholar]
  11. Kobayashi Y., Takahashi I., Nakae T. Diffusion of beta-lactam antibiotics through liposome membranes containing purified porins. Antimicrob Agents Chemother. 1982 Nov;22(5):775–780. doi: 10.1128/aac.22.5.775. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  13. Lindley E. V., Munske G. R., Magnuson J. A. Kinetic analysis of tetracycline accumulation by Streptococcus faecalis. J Bacteriol. 1984 Apr;158(1):334–336. doi: 10.1128/jb.158.1.334-336.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lovering A. M., White L. O., Reeves D. S. The assay of chloramphenicol acetyltransferase activity by high performance liquid chromatography. J Antimicrob Chemother. 1986 Jun;17(6):821–825. doi: 10.1093/jac/17.6.821. [DOI] [PubMed] [Google Scholar]
  15. McMurry L. M., Cullinane J. C., Petrucci R. E., Jr, Levy S. B. Active uptake of tetracycline by membrane vesicles from susceptible Escherichia coli. Antimicrob Agents Chemother. 1981 Sep;20(3):307–313. doi: 10.1128/aac.20.3.307. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Mendelman P. M., Chaffin D. O., Stull T. L., Rubens C. E., Mack K. D., Smith A. L. Characterization of non-beta-lactamase-mediated ampicillin resistance in Haemophilus influenzae. Antimicrob Agents Chemother. 1984 Aug;26(2):235–244. doi: 10.1128/aac.26.2.235. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Munske G. R., Lindley E. V., Magnuson J. A. Streptococcus faecalis proton gradients and tetracycline transport. J Bacteriol. 1984 Apr;158(1):49–54. doi: 10.1128/jb.158.1.49-54.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Nakae R., Nakae T. Diffusion of aminoglycoside antibiotics across the outer membrane of Escherichia coli. Antimicrob Agents Chemother. 1982 Oct;22(4):554–559. doi: 10.1128/aac.22.4.554. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Nierhaus D., Nierhaus K. H. Identification of the chloramphenicol-binding protein in Escherichia coli ribosomes by partial reconstitution. Proc Natl Acad Sci U S A. 1973 Aug;70(8):2224–2228. doi: 10.1073/pnas.70.8.2224. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Reeve E. C. Characteristics of some single-step mutants to chloramphenicol resistance in Escherichia coli K12 and their interactions with R-factor genes. Genet Res. 1966 Apr;7(2):281–286. doi: 10.1017/s0016672300009708. [DOI] [PubMed] [Google Scholar]
  21. Roberts M. C., Swenson C. D., Owens L. M., Smith A. L. Characterization of chloramphenicol-resistant Haemophilus influenzae. Antimicrob Agents Chemother. 1980 Oct;18(4):610–615. doi: 10.1128/aac.18.4.610. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Smith A. L., Smith D. H. Improved enzymatic assay of chloramphenicol. Clin Chem. 1978 Sep;24(9):1452–1457. [PubMed] [Google Scholar]
  23. Thomson T. B., Crider B. P., Eagon R. G. The kinetics of dihydrostreptomycin uptake in Pseudomonas putida membrane vesicles: absence of inhibition by cations. J Antimicrob Chemother. 1985 Aug;16(2):157–163. doi: 10.1093/jac/16.2.157. [DOI] [PubMed] [Google Scholar]
  24. VAZQUEZ D. ANTIBIOTICS WHICH AFFECT PROTEIN SYNTHESIS: THE UPTAKE OF 14C-CHLORAMPHENICOL BY BACTERIA. Biochem Biophys Res Commun. 1963 Aug 14;12:409–413. doi: 10.1016/0006-291x(63)90115-3. [DOI] [PubMed] [Google Scholar]
  25. Vachon V., Lyew D. J., Coulton J. W. Transmembrane permeability channels across the outer membrane of Haemophilus influenzae type b. J Bacteriol. 1985 Jun;162(3):918–924. doi: 10.1128/jb.162.3.918-924.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Walsh C. T., Kaback H. R. Membrane transport as a potential target for antibiotic action. Ann N Y Acad Sci. 1974 May 10;235(0):519–541. doi: 10.1111/j.1749-6632.1974.tb43288.x. [DOI] [PubMed] [Google Scholar]

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