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Journal of Bacteriology, May 2008, p. 3203-3212, Vol. 190, No. 9
0021-9193/08/$08.00+0     doi:10.1128/JB.01911-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Sustained Axenic Metabolic Activity by the Obligate Intracellular Bacterium Coxiella burnetii{triangledown} ,{dagger}

Anders Omsland,1 Diane C. Cockrell,1 Elizabeth R. Fischer,2 and Robert A. Heinzen1*

Coxiella Pathogenesis Section, Laboratory of Intracellular Parasites,1 Microscopy Unit, Research Technology Section, Research Technology Branch, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana 598402

Received 6 December 2007/ Accepted 14 February 2008

Growth of Coxiella burnetii, the agent of Q fever, is strictly limited to colonization of a viable eukaryotic host cell. Following infection, the pathogen replicates exclusively in an acidified (pH 4.5 to 5) phagolysosome-like parasitophorous vacuole. Axenic (host cell free) buffers have been described that activate C. burnetii metabolism in vitro, but metabolism is short-lived, with bacterial protein synthesis halting after a few hours. Here, we describe a complex axenic medium that supports sustained (>24 h) C. burnetii metabolic activity. As an initial step in medium development, several biological buffers (pH 4.5) were screened for C. burnetii metabolic permissiveness. Based on [35S]Cys-Met incorporation, C. burnetii displayed optimal metabolic activity in citrate buffer. To compensate for C. burnetii auxotrophies and other potential metabolic deficiencies, we developed a citrate buffer-based medium termed complex Coxiella medium (CCM) that contains a mixture of three complex nutrient sources (neopeptone, fetal bovine serum, and RPMI cell culture medium). Optimal C. burnetii metabolism occurred in CCM with a high chloride concentration (140 mM) while the concentrations of sodium and potassium had little effect on metabolism. CCM supported prolonged de novo protein and ATP synthesis by C. burnetii (>24 h). Moreover, C. burnetii morphological differentiation was induced in CCM as determined by the transition from small-cell variant to large-cell variant. The sustained in vitro metabolic activity of C. burnetii in CCM provides an important tool to investigate the physiology of this organism including developmental transitions and responses to antimicrobial factors associated with the host cell.

* Corresponding author. Mailing address: Rocky Mountain Laboratories, 903 South 4th Street, Hamilton, MT 59840. Phone: (406) 375-9695. Fax: (406) 363-9380. E-mail: rheinzen{at}niaid.nih.gov

{triangledown} Published ahead of print on 29 February 2008.

{dagger} Supplemental material for this article may be found at http://jb.asm.org/.

Journal of Bacteriology, May 2008, p. 3203-3212, Vol. 190, No. 9
0021-9193/08/$08.00+0     doi:10.1128/JB.01911-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Omsland, A., Cockrell, D. C., Howe, D., Fischer, E. R., Virtaneva, K., Sturdevant, D. E., Porcella, S. F., Heinzen, R. A. (2009). Host cell-free growth of the Q fever bacterium Coxiella burnetii. Proc. Natl. Acad. Sci. USA 106: 4430-4434 [Abstract] [Full Text]  
  • Beare, P. A., Howe, D., Cockrell, D. C., Omsland, A., Hansen, B., Heinzen, R. A. (2009). Characterization of a Coxiella burnetii ftsZ Mutant Generated by Himar1 Transposon Mutagenesis. J. Bacteriol. 191: 1369-1381 [Abstract] [Full Text]  


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