This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Google Scholar
Right arrow Articles by Delgado-Viscogliosi, P.
Right arrow Articles by Delattre, J.-M.
PubMed
Right arrow PubMed Citation
Right arrow Articles by Delgado-Viscogliosi, P.
Right arrow Articles by Delattre, J.-M.
Agricola
Right arrow Articles by Delgado-Viscogliosi, P.
Right arrow Articles by Delattre, J.-M.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, June 2009, p. 3502-3512, Vol. 75, No. 11
0099-2240/09/$08.00+0     doi:10.1128/AEM.02878-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Viability PCR, a Culture-Independent Method for Rapid and Selective Quantification of Viable Legionella pneumophila Cells in Environmental Water Samples{triangledown}

Pilar Delgado-Viscogliosi,* Lydie Solignac, and Jean-Marie Delattre

Département Eaux et Environnement, Institut Pasteur de Lille, F-59019 Lille Cedex, France

Received 18 December 2008/ Accepted 31 March 2009

PCR-based methods have been developed to rapidly screen for Legionella pneumophila in water as an alternative to time-consuming culture techniques. However, these methods fail to discriminate between live and dead bacteria. Here, we report a viability assay (viability PCR [v-PCR]) for L. pneumophila that combines ethidium monoazide bromide with quantitative real-time PCR (qPCR). The ability of v-PCR to differentiate viable from nonviable L. pneumophila cells was confirmed with permeabilizing agents, toluene, or isopropanol. v-PCR suppressed more than 99.9% of the L. pneumophila PCR signal in nonviable cultures and was able to discriminate viable cells in mixed samples. A wide range of physiological states, from culturable to dead cells, was observed with 64 domestic hot-water samples after simultaneous quantification of L. pneumophila cells by v-PCR, conventional qPCR, and culture methods. v-PCR counts were equal to or higher than those obtained by culture and lower than or equal to conventional qPCR counts. v-PCR was used to successfully monitor in vitro the disinfection efficacy of heating to 70°C and glutaraldehyde and chlorine curative treatments. The v-PCR method appears to be a promising and rapid technique for enumerating L. pneumophila bacteria in water and, in comparison with conventional qPCR techniques used to monitor Legionella, has the advantage of selectively amplifying only viable cells.

* Corresponding author. Mailing address: Département Eaux et Environnement, Institut Pasteur de Lille, F-59019 Lille Cedex, France. Phone: 33 3 20 87 71 20. Fax: 33 3 20 87 73 83. E-mail: pilar.viscogliosi{at}pasteur-lille.fr

{triangledown} Published ahead of print on 10 April 2009.

Applied and Environmental Microbiology, June 2009, p. 3502-3512, Vol. 75, No. 11
0099-2240/09/$08.00+0     doi:10.1128/AEM.02878-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»