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Open Access Research article

Characterization and resuscitation of ‘non-culturable’ cells of Legionella pneumophila

Adrien Ducret12, Maïalène Chabalier1 and Sam Dukan1*

Author Affiliations

1 Aix Marseille Université, Laboratoire de Chimie Bactérienne (UMR7283), Institut de Microbiologie de la Méditerranée - CNRS, 31, Chemin Joseph Aiguier, 13402 Marseille, France

2 Present address: Department of Biology, Indiana University, 1001 East 3rd Street, Bloomington, IN 47405 USA

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BMC Microbiology 2014, 14:3  doi:10.1186/1471-2180-14-3

Published: 2 January 2014

Abstract

Background

Legionella pneumophila is a waterborne pathogen responsible for Legionnaires’ disease, an infection which can lead to potentially fatal pneumonia. After disinfection, L. pneumophila has been detected, like many other bacteria, in a “viable but non culturable” state (VBNC). The physiological significance of the VBNC state is unclear and controversial: it could be an adaptive response favoring long-term survival; or the consequence of cellular deterioration which, despite maintenance of certain features of viable cells, leads to death; or an injured state leading to an artificial loss of culturability during the plating procedure. VBNC cells have been found to be resuscitated by contact with amoebae.

Results

We used quantitative microscopic analysis, to investigate this “resuscitation” phenomenon in L. pneumophila in a model involving amending solid plating media with ROS scavengers (pyruvate or glutamate), and co-culture with amoebae. Our results suggest that the restoration observed in the presence of pyruvate and glutamate may be mostly due to the capacity of these molecules to help the injured cells to recover after a stress. We report evidence that this extracellular signal leads to a transition from a not-culturable form to a culturable form of L. pneumophila, providing a technique for recovering virulent and previously uncultivated forms of L. pneumophila.

Conclusion

These new media could be used to reduce the risk of underestimation of counts of virulent of L. pneumophila cells in environmental samples.