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JAC Advance Access published online on May 9, 2005
Journal of Antimicrobial Chemotherapy, doi:10.1093/jac/dki141
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© The Author 2005. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oupjournals.org
Received January 5, 2005
Revised March 25, 2005
Accepted March 31, 2005

Original article

Antibiotic susceptibilities of Legionella pneumophila strain Paris in THP-1 cells as determined by real-time PCR assay

N. Roch 1 and M. Maurin 1*

1 CNRS UMR 5163, Université Joseph Fourier, Faculté de Médecine, Bâtiment Jean Roget, 38700 La Tronche, France

* To whom correspondence should be addressed.
M. Maurin, E-mail: mmaurin{at}chu-grenoble.fr


  Abstract

Objectives: Legionella species are facultative intracellular bacteria. Evaluation of the activity of antibiotics against intracellular L. pneumophila is more predictive of their in vivo efficacy than MICs as determined in axenic medium. However, current methodologies are based on cfu count determination, and are tedious because of the slow growth of Legionella spp. We investigated antibiotic susceptibilities of L. pneumophila strain Paris in THP-1-derived macrophages, using a real-time PCR assay for evaluation of bacterial growth.

Methods: Intracellular activities of seven antibiotic compounds against two human isolates of L. pneumophila strain Paris were determined in THP-1-derived macrophages in vitro. Bacterial growth was evaluated using either cfu methodology or a real-time PCR protocol targeting the mip gene.

Results: Bacterial titres as determined using real-time PCR were well correlated with cfu counts. Antibiotic susceptibilities for the two L. pneumophila isolates tested were comparable when using either of the two techniques. MICs were also similar to those previously reported for other L. pneumophila serogroup 1 strains. In particular, rifampicin and the fluoroquinolones were the most active compounds, both in extracellular medium and in THP-1 cells. Real-time PCR, however, was much less laborious than the traditional cfu method.

Conclusions: Real-time PCR is better adapted than cfu-based methods to evaluating the antibiotic susceptibilities of large series of Legionella strains to newer antibiotic compounds.

Keywords: antibacterials; intracellular; macrophages; quantitative PCR.
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