The Q fever bacterium Coxiella burnetii replicates inside host cells within a large Coxiella-containing vacuole (CCV) whose biogenesis relies on the Dot/Icm-dependent secretion of bacterial effectors. Several membrane trafficking pathways contribute membranes, proteins, and lipids for CCV biogenesis. These include the endocytic and autophagy pathways, which are characterized by phosphatidylinositol 3-phosphate [PI(3)P]-positive membranes. Here we show that the C. burnetii secreted effector Coxiella vacuolar protein B (CvpB) binds PI(3)P and phosphatidylserine (PS) on CCVs and early endosomal compartments and perturbs the activity of the phosphatidylinositol 5-kinase PIKfyve to manipulate PI(3)P metabolism. CvpB association to early endosome triggers vacuolation and clustering, leading to the channeling of large PI(3)P-positive membranes to CCVs for vacuole expansion. At CCVs, CvpB binding to early endosome-and autophagy-derived PI(3)P and the concomitant inhibition of PIKfyve favor the association of the autophagosomal machinery to CCVs for optimal homotypic fusion of the Coxiella-containing compartments. The importance of manipulating PI(3)P metabolism is highlighted by mutations in cvpB resulting in a multivacuolar phenotype, rescuable by gene complementation, indicative of a defect in CCV biogenesis. Using the insect model Galleria mellonella, we demonstrate the in vivo relevance of defective CCV biogenesis by highlighting an attenuated virulence phenotype associated with cvpB mutations.Coxiella burnetii | host-pathogen interactions | phosphoinositides

New insights into Legionella pneumophila biofilm regulation by c-di-GMP signaling.(2016) Biofouling, Open Archive TOULOUSE Archive Ouverte (OATAO) OATAO is an open access repository that collects the work of some Toulouse researchers and makes it freely available over the web where possible. This is an author's version published in : http://oatao.univ-toulouse.fr/19753Official URL : https://doi.org/10. 1080/08927014.2016.1212988 Any correspondence concerning this service should be sent to the repository administrator : tech-oatao@listes-diff.inp-toulouse.fr The waterborne pathogen Legionella pneumophila grows as a bio lm, freely or inside amoebae. Cyclic-di-GMP (c-di-GMP), a bacterial second messenger frequently implicated in bio lm 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 bio lm regulation, the consequences on bio lm 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 di erent aspects of bio lm formation, while two proteins with dual activity (DGC/PDE) inhibit bio lm growth. Surprisingly, only two mutants exhibited a change in global c-di-GMP concentration. This study highlights that speci c c-di-GMP pathways control L. pneumophila bio lm 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 bio lm in response to nitric oxide, a signal for bio lm dispersion in many other species. New insights into Legionella pneumophila bio lm regulation by c-di-GMP signaling