1887

Abstract

The ability of to persist in its human host despite extensive chemotherapy is thought to be based on subpopulations of non-replicating phenotypically drug-resistant bacilli. To study the non-growing pathogen, culture models that generate quiescent organisms by either oxygen depletion in nutrient-rich medium (Wayne model) or nutrient deprivation in oxygen-rich medium (Loebel model) have been developed. In contrast to the energy metabolism of Wayne bacilli, little is known about Loebel bacilli. Here we analysed under nutrient-starvation conditions. Upon shifting to the non-replicating state the pathogen maintained a fivefold reduced but constant intracellular ATP level. Chemical probing of the FF ATP synthase demonstrated the importance of this enzyme for ATP homeostasis and viability of the nutrient-starved organism. Surprisingly, the specific ATP synthase inhibitor TMC207 did not affect viability and only moderately reduced the intracellular ATP level of nutrient-starved organisms. Depletion of oxygen killed Loebel bacilli, whereas death was prevented by nitrate, suggesting that respiration and an exogenous electron acceptor are required for maintaining viability. Nutrient-starved bacilli lacking the glyoxylate shunt enzyme isocitrate lyase failed to reduce their intracellular ATP level and died, thus establishing a link between ATP control and intermediary metabolism. We conclude that reduction of the ATP level might be an important step in the adaptation of to non-growing survival.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.033084-0
2010-01-01
2024-03-29
Loading full text...

Full text loading...

/deliver/fulltext/micro/156/1/81.html?itemId=/content/journal/micro/10.1099/mic.0.033084-0&mimeType=html&fmt=ahah

References

  1. Amaral L., Kristiansen J. E., Abebe L. S., Millett W. 1996; Inhibition of the respiration of multi-drug resistant clinical isolates of Mycobacterium tuberculosis by thioridazine: potential use for initial therapy of freshly diagnosed tuberculosis. J Antimicrob Chemother 38:1049–1053
    [Google Scholar]
  2. Andries K., Verhasselt P., Guillemont J., Göhlmann H. W., Neefs J. M., Winkler H., Van Gestel J., Timmerman P., Zhu M. other authors 2005; A diarylquinoline drug active on the ATP synthase of Mycobacterium tuberculosis. Science 307:223–227
    [Google Scholar]
  3. Bardarov S., Bardarov S. Jr, Pavelka M. S. Jr, Sambandamurthy V., Larsen M., Tufariello J., Chan J., Hatfull G., Jacobs W. R. Jr 2002; Specialized transduction: an efficient method for generating marked and unmarked targeted gene disruptions in Mycobacterium tuberculosis, M.bovis BCG and M. smegmatis. Microbiology 148:3007–3017
    [Google Scholar]
  4. Betts J. C., Lukey P. T., Robb L. C., McAdam R. A., Duncan K. 2002; Evaluation of a nutrient starvation model of Mycobacterium tuberculosis persistence by gene and protein expression profiling. Mol Microbiol 43:717–731
    [Google Scholar]
  5. Boshoff H. I., Barry C. E. III 2005; Tuberculosis – metabolism and respiration in the absence of growth. Nat Rev Microbiol 3:70–80
    [Google Scholar]
  6. Cattell K. J., Lindop C. R., Knight I. G., Beechey R. B. 1971; The identification of the site of action of N, N′-dicyclohexylcarbodi-imide as a proteolipid in mitochondrial membranes. Biochem J 125:169–177
    [Google Scholar]
  7. Dahl J. L., Kraus C. N., Boshoff H. I., Doan B., Foley K., Avarbock D., Kaplan G., Mizrahi V., Rubin H., Barry C. E. III: 2003; The role of RelMtb-mediated adaptation to stationary phase in long-term persistence of Mycobacterium tuberculosis in mice. Proc Natl Acad Sci U S A 100:10026–10031
    [Google Scholar]
  8. Dick T. 2001; Dormant tubercle bacilli: the key to more effective TB chemotherapy?. J Antimicrob Chemother 47:117–118
    [Google Scholar]
  9. Fenhalls G., Stevens L., Moses L., Bezuidenhout J., Betts J. C., Helden P. van., Lukey P. T., Duncan K. 2002; In situ detection of Mycobacterium tuberculosis transcripts in human lung granulomas reveals differential gene expression in necrotic lesions. Infect Immun 70:6330–6338
    [Google Scholar]
  10. Gordhan B. G., Smith D. A., Kana B. D., Bancroft G., Mizrahi V. 2006; The carbon starvation-inducible genes Rv2557 and Rv2558 of Mycobacterium tuberculosis are not required for long-term survival under carbon starvation and for virulence in SCID mice. Tuberculosis (Edinb 86:430–437
    [Google Scholar]
  11. Harries A. D., Dye C. 2006; Tuberculosis. Ann Trop Med Parasitol 100:415–431
    [Google Scholar]
  12. Hinds J., Mahenthiralingam E., Kempsell K. E., Duncan K., Stokes R. W., Parish T., Stoker N. G. 1999; Enhanced gene replacement in mycobacteria. Microbiology 145:519–527
    [Google Scholar]
  13. Honer Zu Bentrup K., Miczak A., Swenson D. L., Russell D. G. 1999; Characterization of activity and expression of isocitrate lyase in Mycobacterium avium and Mycobacterium tuberculosis. J Bacteriol 181:7161–7167
    [Google Scholar]
  14. Koul A., Dendouga N., Vergauwen K., Molenberghs B., Vranckx L., Willebrords R., Ristic Z., Lill H., Dorange I. other authors 2007; Diarylquinolines target subunit c of mycobacterial ATP synthase. Nat Chem Biol 3:323–324
    [Google Scholar]
  15. Koul A., Vranckx L., Dendouga N., Balemans W., Van den Wyngaert I., Vergauwen K., Göhlmann H. W., Willebrords R., Poncelet A. other authors 2008; Diarylquinolines are bactericidal for dormant mycobacteria as a result of disturbed ATP homeostasis. J Biol Chem 283:25273–25280
    [Google Scholar]
  16. Lee M. H., Pascopella L., Jacobs W. R. Jr, Hatfull G. F. 1991; Site-specific integration of mycobacteriophage L5: integration-proficient vectors for Mycobacterium smegmatis, Mycobacterium tuberculosis, and bacille Calmette-Guerin. Proc Natl Acad Sci U S A 88:3111–3115
    [Google Scholar]
  17. Lim A., Eleuterio M., Hutter B., Murugasu-Oei B., Dick T. 1999; Oxygen depletion-induced dormancy in Mycobacterium bovis BCG. J Bacteriol 181:2252–2256
    [Google Scholar]
  18. Loebel R. O., Shorr E., Richardson H. B. 1933a; The influence of adverse conditions upon the respiratory metabolism and growth of human tubercle bacilli. J Bacteriol 26:167–200
    [Google Scholar]
  19. Loebel R. O., Shorr E., Richardson H. B. 1933b; The influence of foodstuffs upon the respiratory metabolism and growth of human tubercle bacilli. J Bacteriol 26:139–166
    [Google Scholar]
  20. McKinney J. D., Honer zu Bentrup K., Munoz-Elias E. J., Miczak A., Chen B., Chan W. T., Swenson D., Sacchettini J. C., Jacobs W. R. Jr, Russell D. G. 2000; Persistence of Mycobacterium tuberculosis in macrophages and mice requires the glyoxylate shunt enzyme isocitrate lyase. Nature 406:735–738
    [Google Scholar]
  21. Nyka W. 1974; Studies on the effect of starvation on mycobacteria. Infect Immun 9:843–850
    [Google Scholar]
  22. Ovchinnikov Y. A., Abdulaev N. G., Modyanov N. N. 1982; Structural basis of proton-translocating protein function. Annu Rev Biophys Bioeng 11:445–463
    [Google Scholar]
  23. Parish T., Stoker N. G. 2000; Use of a flexible cassette method to generate a double unmarked Mycobacterium tuberculosis tlyA plcABC mutant by gene replacement. Microbiology 146:1969–1975
    [Google Scholar]
  24. Primm T. P., Andersen S. J., Mizrahi V., Avarbock D., Rubin H., Barry C. E. III 2000; The stringent response of Mycobacterium tuberculosis is required for long-term survival. J Bacteriol 182:4889–4898
    [Google Scholar]
  25. Rao S. P., Alonso S., Rand L., Dick T., Pethe K. 2008; The protonmotive force is required for maintaining ATP homeostasis and viability of hypoxic, nonreplicating Mycobacterium tuberculosis. Proc Natl Acad Sci U S A 105:11945–11950
    [Google Scholar]
  26. Stover C. K., de la Cruz V. F., Fuerst T. R., Burlein J. E., Benson L. A., Bennett L. T., Bansal G. P., Young J. F., Lee M. H. 1991; New use of BCG for recombinant vaccines. Nature 351:456–460
    [Google Scholar]
  27. Teh J. S., Yano T., Rubin H. 2007; Type II NADH : menaquinone oxidoreductase of Mycobacterium tuberculosis. Infect Disord Drug Targets 7:169–181
    [Google Scholar]
  28. Via L. E., Lin P. L., Ray S. M., Carrillo J., Allen S. S., Eum S. Y., Taylor K., Klein E., Manjunatha U. other authors 2008; Tuberculous granulomas are hypoxic in guinea pigs, rabbits, and nonhuman primates. Infect Immun 76:2333–2340
    [Google Scholar]
  29. Wayne L. G., Hayes L. G. 1996; An in vitro model for sequential study of shiftdown of Mycobacterium tuberculosis through two stages of nonreplicating persistence. Infect Immun 64:2062–2069
    [Google Scholar]
  30. Wayne L. G., Sohaskey C. D. 2001; Nonreplicating persistence of Mycobacterium tuberculosis. Annu Rev Microbiol 55:139–163
    [Google Scholar]
  31. Weinstein E. A., Yano T., Li L. S. other authors 2005; Inhibitors of type II NADH : menaquinone oxidoreductase represent a class of antitubercular drugs. Proc Natl Acad Sci U S A 102:4548–4553
    [Google Scholar]
  32. Williams K. J., Duncan K. 2007; Current strategies for identifying and validating targets for new treatment-shortening drugs for TB. Curr Mol Med 7:297–307
    [Google Scholar]
  33. Xie Z., Siddiqi N., Rubin E. J. 2005; Differential antibiotic susceptibilities of starved Mycobacterium tuberculosis isolates. Antimicrob Agents Chemother 49:4778–4780
    [Google Scholar]
  34. Yano T., Li L. S., Weinstein E., Teh J. S., Rubin H. 2006; Steady-state kinetics and inhibitory action of antitubercular phenothiazines on Mycobacterium tuberculosis type-II NADH-menaquinone oxidoreductase (NDH-2. J Biol Chem 281:11456–11463
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.033084-0
Loading
/content/journal/micro/10.1099/mic.0.033084-0
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF
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