New insights into Legionella pneumophila biofilm regulation by c-di-GMP signaling

Abstract

The waterborne pathogen Legionella pneumophila grows as a biofilm, freely or inside amoebae. Cyclic-di-GMP (c-di-GMP), a bacterial second messenger frequently implicated in biofilm formation, is synthesized and degraded by diguanylate cyclases (DGCs) and phosphodiesterases (PDEs), respectively. To characterize the c-di-GMP-metabolizing enzymes involved in L. pneumophila biofilm regulation, the consequences on biofilm formation and the c-di-GMP concentration of each corresponding gene inactivation were assessed in the Lens strain. The results showed that one DGC and two PDEs enhance different aspects of biofilm formation, while two proteins with dual activity (DGC/PDE) inhibit biofilm growth. Surprisingly, only two mutants exhibited a change in global c-di-GMP concentration. This study highlights that specific c-di-GMP pathways control L. pneumophila biofilm formation, most likely via temporary and/or local modulation of c-di-GMP concentration. Furthermore, Lpl1054 DGC is required to enable the formation a dense biofilm in response to nitric oxide, a signal for biofilm dispersion in many other species.

Keywords:

• Biofilm
• Legionella
• c-di-GMP signaling
• second messenger
• NO response

Acknowledgements

The authors are grateful to Claire Andréa and Angeline Rizzo for their technical assistance. c-di-GMP quantifications by UPLC-MS were performed at the Service Commun d’HPLC (Institut de Chimie de Toulouse). Confocal observations were carried out at FRAIB (TRI-GENOTOUL) in Castanet-Tolosan, France. The authors are also grateful to Dr C. Zedde and Dr A. Le Ru for their technical knowledge of UPLC-MS analysis and confocal microscopy respectively.