TNPHYTO

Plant pathogenic bacteria are a main threat for food production worldwide and no efficient means of control exist. Crop protection methods require knowledge of the genetic and phenotypic diversity of plant pathogens, of the extent of their host range and of habitats in which reservoirs of inoculum are found. Studies on plant pathogenic bacteria have often been conducted on a limited number of model laboratory strains and model plants. While some bacteria are found only in infected plants, others can be isolated in various environments such as soil, water or healthy plants. These strains could represent a potential source of disease outbreak and nothing is known on how environmental reservoirs weight on their survival, multiplication and pathogeny. From the perspective of developing more comprehensive approaches to plant disease studies, including environment, we propose to investigate genetic determinants of bacterial lifestyle both in planta and in fresh water, a major vector of pathogen dissemination through the river and irrigation networks. Availability of large collections of environmental strains and new high throughput sequencing techniques now allow to identify simultaneously, in a large panel of strains, genes required for growth in a given condition, like in planta, in water or in soil. In this project, we will use Tn-Seq, an innovative mutagenesis and high throughput sequencing technique to find such genetic determinants, in three wide spectrum pathogenic bacteria present in France: Pseudomonas syringae, Pectobacterium spp and Dickeya spp. Strains of these bacteria will be selected in our collections to represent the largest phylogenetic diversity and origin. The Tn-seq analysis will be performed on several plants and in river water. To analyse this data, we will develop a dedicated open-source bioinformatics pipeline, that will address the challenge of multiple conditions and multiple strains comparison. Validation of the results obtained will necessitate construction of a large number of mutants. In addition to the techniques commonly used to make mutants, we will use CrispRi gene silencing method in the three species. All these approaches will allow the identification of new genetic determinants involved in virulence on plants and those required for fitness in water, a major environmental habitats. Comparison of the two habitats will identify genetic determinants in common and lead to hypothesis on how environmental reservoirs could drive growth, survival and pathogeny of pathogens. Studying various pathogenic bacteria will allow to identify the virulence genetic determinants conserved at the genus, species or strain level. The results obtained in this work could help predict strain virulence, develop antibacterial strategies.