The Poplar-Poplar Rust Interaction: Insights from Genomics and Transcriptomics

Poplars are extensively cultivated worldwide, and their susceptibility to the leaf rust fungus Melampsora larici-populina leads to considerable damages in plantations. Despite a good knowledge of the poplar rust life cycle, and particularly the epidemics on poplar, the perennial status of the plant host and the obligate biotrophic lifestyle of the rust fungus are bottlenecks for molecular investigations. Following the completion of both M. larici-populina and Populus trichocarpa genome sequences, gene families involved in poplar resistance or in rust fungus virulence were investigated, allowing the identification of key genetic determinants likely controlling the outcome of the interaction. Specific expansions of resistance and defense-related genes in poplar indicate probable innovations in perennial species in relation with host-pathogen interactions. The genome of M. Larici-populina contains a strikingly high number of genes encoding small secreted proteins (SSPs) representing hundreds of candidate effectors. Transcriptome analyses of interacting partners in compatible and incompatible interactions revealed conserved set of genes involved in poplar defense reactions as well as timely regulated expression of SSP transcripts during host tissues colonisation. Ongoing functional studies of selected candidate effectors will be achieved mainly on the basis of recombinant protein purification and subsequent characterisation

The Concepts of Heritage and Heritage Resource Applied to Karsts: Protecting the Choranche Caves (Vercors, France)

In 2005, French Ministry of Ecology started procedure to inscribe 18 Caves of Chorance into the World Heritage list of UNESCO. The application has to answer to three objectives: the scientific interest, definition of the territory, and to propose the management model. For the first all the heritage sources has to be identified, such as flowstone formations, karst water objects and historical curiosities. This are very important and sensitive questions specially because they have been not answered before the procedure of the inscription started off

Microbiota and Host Nutrition across Plant and Animal Kingdoms

Plants and animals each have evolved specialized organs dedicated to nutrient acquisition, and these harbor specific bacterial communities that extend the host's metabolic repertoire. Similar forces driving microbial community establishment in the gut and plant roots include diet/soil-type, host genotype, and immune system as well as microbe-microbe interactions. Here we show that there is no overlap of abundant bacterial taxa between the microbiotas of the mammalian gut and plant roots, whereas taxa overlap does exist between fish gut and plant root communities. A comparison of root and gut microbiota composition in multiple host species belonging to the same evolutionary lineage reveals host phylogenetic signals in both eukaryotic kingdoms. The reasons underlying striking differences in microbiota composition in independently evolved, yet functionally related, organs in plants and animals remain unclear but might include differences in start inoculum and niche-specific factors such as oxygen levels, temperature, pH, and organic carbon availability

Sequential Delivery of Host-Induced Virulence Effectors by Appressoria and Intracellular Hyphae of the Phytopathogen Colletotrichum higginsianum

Phytopathogens secrete effector proteins to manipulate their hosts for effective colonization. Hemibiotrophic fungi must maintain host viability during initial biotrophic growth and elicit host death for subsequent necrotrophic growth. To identify effectors mediating these opposing processes, we deeply sequenced the transcriptome of Colletotrichum higginsianum infecting Arabidopsis. Most effector genes are host-induced and expressed in consecutive waves associated with pathogenic transitions, indicating distinct effector suites are deployed at each stage. Using fluorescent protein tagging and transmission electron microscopy-immunogold labelling, we found effectors localised to stage-specific compartments at the host-pathogen interface. In particular, we show effectors are focally secreted from appressorial penetration pores before host invasion, revealing new levels of functional complexity for this fungal organ. Furthermore, we demonstrate that antagonistic effectors either induce or suppress plant cell death. Based on these results we conclude that hemibiotrophy in Colletotrichum is orchestrated through the coordinated expression of antagonistic effectors supporting either cell viability or cell death

Contribution à l'étude des déterminants génétiques impliqués dans le processus infectieux de Melampsora larici-populina, l'agent de la rouille foliaire du peuplier

The leaf rust disease caused by Melampsora larici-populina (Mlp) is the main disease affecting poplar plantations in Europe with severe economic losses. The recent sequencing of the genome of Mlp (strain 98AG31) opens new perspectives to identify key genes involved in the fungal infection process and particularly those encoding fungal effectors that could manipulate host cell structure and function to facilitate host colonization. Analysis of the rust transcriptome during time course infection of poplar leaves, based on NimbleGen oligoarrays and massive EST sequencing led to the identification of genes related to fungal germination, biotrophy and sporulation. A consistant induction of genes encoding small-secreted proteins (SSPs) was observed during the biotrophic growth at 96 hours post-inoculation (hpi) but also at 168 hpi in the palisade mesophyll using laser capture microdissection. Mlp Secretome analysis, based on annotation, evolution and expression of genes encoding SSPs helped in identifying candidate poplar rust effectors. Some, specifically expressed in planta or showing homologies with known rust effectors were localized around the haustorium. Interestingly, other candidate genes, belonging to multigenic families under diversifying selection are cystein-rich, specifically expressed in planta and harbour a translocation signal potentially involved in effector export inside host cell. This genome-wide analysis of putative fungal effectors will contribute to the general knowledge of rust biology and will help to set new approaches to prevent and control the diseaseLa maladie de la rouille foliaire du peuplier, causée par le basidiomycète Melampsora larici-populina (Mlp) cause des dégâts importants dans les peupleraies européennes. Le séquençage du génome de la souche 98AG31 de Mlp a ouvert de nouvelles perspectives pour l'identification de déterminants géniques impliqués dans le processus infectieux du champignon et notamment ceux codant des effecteurs fongiques capables de manipuler la structure et le fonctionnement de la cellule hôte pour assurer le succès de l'infection. L'analyse du transcriptome du champignon au cours des différentes phases du processus infectieux, basée sur l'utilisation de puces à oligonucléotides NimbleGen ou le séquençage massifs d'ESTs, a permis d?identifier des gènes marqueurs de la germination, de la phase de croissance biotrophe et de la sporulation du champignon. Nous avons notamment pu montrer l'induction importante de nombreux gènes codant des petites protéines sécrétées (SSPs) au cours de la phase biotrophe à 96 hpi heures post-inoculation (hpi) ainsi qu'au sein du parenchyme lacuneux à 168 hpi par microdissection à capture laser. L?analyse fine du sécrétome de Mlp, basée sur l'annotation, l'évolution et l'expression des gènes codant des SSPs a permis de mettre à jour des effecteurs candidats. Certains, spécifiquement exprimés in planta ou présentant des homologies de séquence avec des effecteurs de rouilles ont été localisés au niveau de l'haustorium. De manière intéressante, d'autre gènes candidats appartenant à des familles multigéniques sous pression de sélection positive, sont riches en cystéines, spécifiquement exprimés in planta et possèdent un motif de translocation potentiellement impliqué dans l'export de l'effecteur dans la cellule hôte. Ce travail d'analyse fine des effecteurs potentiels d'un agent de rouille à l'échelle génomique va contribuer à l'amélioration des connaissances sur la biologie de ces champignons biotrophes et contribuera à faciliter la recherche de nouvelles méthodes de lutte contre la maladi

Analysis of Melampsora larici-populina genetic determinants involved in the poplar leaf infection process. Genomic and transcriptomic approaches

La maladie de la rouille foliaire du peuplier, causée par le basidiomycète Melampsora larici-populina (Mlp) cause des dégâts importants dans les peupleraies européennes. Le séquençage du génome de la souche 98AG31 de Mlp a ouvert de nouvelles perspectives pour l'identification de déterminants géniques impliqués dans le processus infectieux du champignon et notamment ceux codant des effecteurs fongiques capables de manipuler la structure et le fonctionnement de la cellule hôte pour assurer le succès de l'infection. L'analyse du transcriptome du champignon au cours des différentes phases du processus infectieux, basée sur l'utilisation de puces à oligonucléotides NimbleGen ou le séquençage massifs d'ESTs, a permis d?identifier des gènes marqueurs de la germination, de la phase de croissance biotrophe et de la sporulation du champignon. Nous avons notamment pu montrer l'induction importante de nombreux gènes codant des petites protéines sécrétées (SSPs) au cours de la phase biotrophe à 96 hpi heures post-inoculation (hpi) ainsi qu'au sein du parenchyme lacuneux à 168 hpi par microdissection à capture laser. L?analyse fine du sécrétome de Mlp, basée sur l'annotation, l'évolution et l'expression des gènes codant des SSPs a permis de mettre à jour des effecteurs candidats. Certains, spécifiquement exprimés in planta ou présentant des homologies de séquence avec des effecteurs de rouilles ont été localisés au niveau de l'haustorium. De manière intéressante, d'autre gènes candidats appartenant à des familles multigéniques sous pression de sélection positive, sont riches en cystéines, spécifiquement exprimés in planta et possèdent un motif de translocation potentiellement impliqué dans l'export de l'effecteur dans la cellule hôte. Ce travail d'analyse fine des effecteurs potentiels d'un agent de rouille à l'échelle génomique va contribuer à l'amélioration des connaissances sur la biologie de ces champignons biotrophes et contribuera à faciliter la recherche de nouvelles méthodes de lutte contre la maladieThe leaf rust disease caused by Melampsora larici-populina (Mlp) is the main disease affecting poplar plantations in Europe with severe economic losses. The recent sequencing of the genome of Mlp (strain 98AG31) opens new perspectives to identify key genes involved in the fungal infection process and particularly those encoding fungal effectors that could manipulate host cell structure and function to facilitate host colonization. Analysis of the rust transcriptome during time course infection of poplar leaves, based on NimbleGen oligoarrays and massive EST sequencing led to the identification of genes related to fungal germination, biotrophy and sporulation. A consistant induction of genes encoding small-secreted proteins (SSPs) was observed during the biotrophic growth at 96 hours post-inoculation (hpi) but also at 168 hpi in the palisade mesophyll using laser capture microdissection. Mlp Secretome analysis, based on annotation, evolution and expression of genes encoding SSPs helped in identifying candidate poplar rust effectors. Some, specifically expressed in planta or showing homologies with known rust effectors were localized around the haustorium. Interestingly, other candidate genes, belonging to multigenic families under diversifying selection are cystein-rich, specifically expressed in planta and harbour a translocation signal potentially involved in effector export inside host cell. This genome-wide analysis of putative fungal effectors will contribute to the general knowledge of rust biology and will help to set new approaches to prevent and control the diseas

Contribution à l'étude des déterminants génétiques impliqués dans le processus infectieux de Melampsora larici-populina, l'agent de la rouille foliaire du peuplier

La maladie de la rouille foliaire du peuplier, causée par le basidiomycète Melampsora larici-populina (Mlp) cause des dégâts importants dans les peupleraies européennes. Le séquençage du génome de la souche 98AG31 de Mlp a ouvert de nouvelles perspectives pour l'identification de déterminants géniques impliqués dans le processus infectieux du champignon et notamment ceux codant des effecteurs fongiques capables de manipuler la structure et le fonctionnement de la cellule hôte pour assurer le succès de l'infection. L'analyse du transcriptome du champignon au cours des différentes phases du processus infectieux, basée sur l'utilisation de puces à oligonucléotides NimbleGen ou le séquençage massifs d'ESTs, a permis d?identifier des gènes marqueurs de la germination, de la phase de croissance biotrophe et de la sporulation du champignon. Nous avons notamment pu montrer l'induction importante de nombreux gènes codant des petites protéines sécrétées (SSPs) au cours de la phase biotrophe à 96 hpi heures post-inoculation (hpi) ainsi qu'au sein du parenchyme lacuneux à 168 hpi par microdissection à capture laser. L?analyse fine du sécrétome de Mlp, basée sur l'annotation, l'évolution et l'expression des gènes codant des SSPs a permis de mettre à jour des effecteurs candidats. Certains, spécifiquement exprimés in planta ou présentant des homologies de séquence avec des effecteurs de rouilles ont été localisés au niveau de l'haustorium. De manière intéressante, d'autre gènes candidats appartenant à des familles multigéniques sous pression de sélection positive, sont riches en cystéines, spécifiquement exprimés in planta et possèdent un motif de translocation potentiellement impliqué dans l'export de l'effecteur dans la cellule hôte. Ce travail d'analyse fine des effecteurs potentiels d'un agent de rouille à l'échelle génomique va contribuer à l'amélioration des connaissances sur la biologie de ces champignons biotrophes et contribuera à faciliter la recherche de nouvelles méthodes de lutte contre la maladieThe leaf rust disease caused by Melampsora larici-populina (Mlp) is the main disease affecting poplar plantations in Europe with severe economic losses. The recent sequencing of the genome of Mlp (strain 98AG31) opens new perspectives to identify key genes involved in the fungal infection process and particularly those encoding fungal effectors that could manipulate host cell structure and function to facilitate host colonization. Analysis of the rust transcriptome during time course infection of poplar leaves, based on NimbleGen oligoarrays and massive EST sequencing led to the identification of genes related to fungal germination, biotrophy and sporulation. A consistant induction of genes encoding small-secreted proteins (SSPs) was observed during the biotrophic growth at 96 hours post-inoculation (hpi) but also at 168 hpi in the palisade mesophyll using laser capture microdissection. Mlp Secretome analysis, based on annotation, evolution and expression of genes encoding SSPs helped in identifying candidate poplar rust effectors. Some, specifically expressed in planta or showing homologies with known rust effectors were localized around the haustorium. Interestingly, other candidate genes, belonging to multigenic families under diversifying selection are cystein-rich, specifically expressed in planta and harbour a translocation signal potentially involved in effector export inside host cell. This genome-wide analysis of putative fungal effectors will contribute to the general knowledge of rust biology and will help to set new approaches to prevent and control the diseaseNANCY1-Bib. numérique (543959902) / SudocSudocFranceF

Microbial interactions within the plant holobiont

Abstract Since the colonization of land by ancestral plant lineages 450 million years ago, plants and their associated microbes have been interacting with each other, forming an assemblage of species that is often referred to as a “holobiont.” Selective pressure acting on holobiont components has likely shaped plant-associated microbial communities and selected for host-adapted microorganisms that impact plant fitness. However, the high microbial densities detected on plant tissues, together with the fast generation time of microbes and their more ancient origin compared to their host, suggest that microbe-microbe interactions are also important selective forces sculpting complex microbial assemblages in the phyllosphere, rhizosphere, and plant endosphere compartments. Reductionist approaches conducted under laboratory conditions have been critical to decipher the strategies used by specific microbes to cooperate and compete within or outside plant tissues. Nonetheless, our understanding of these microbial interactions in shaping more complex plant-associated microbial communities, along with their relevance for host health in a more natural context, remains sparse. Using examples obtained from reductionist and community-level approaches, we discuss the fundamental role of microbe-microbe interactions (prokaryotes and micro-eukaryotes) for microbial community structure and plant health. We provide a conceptual framework illustrating that interactions among microbiota members are critical for the establishment and the maintenance of host-microbial homeostasis

Multi-genome metabolic modeling predicts functional inter-dependencies in the Arabidopsis root microbiome

International audienceBackground: From a theoretical ecology point of view, microbiomes are far more complex than expected. Besides competition and competitive exclusion, cooperative microbe-microbe interactions have to be carefully considered. Metabolic dependencies among microbes likely explain co-existence in microbiota. Methodology: In this in silico study, we explored genome-scale metabolic models (GEMs) of 193 bacteria isolated from Arabidopsis thaliana roots. We analyzed their predicted producible metabolites under simulated nutritional constraints including "root exudate-mimicking growth media " and assessed the potential of putative metabolic exchanges of by- and end-products to avoid those constraints. Results: We found that the genome-encoded metabolic potential is quantitatively and qualitatively clustered by phylogeny, highlighting metabolic differentiation between taxonomic groups. Random, synthetic combinations of increasing numbers of strains (SynComs) indicated that the number of producible compounds by GEMs increased with average phylogenetic distance, but that most SynComs were centered around an optimal phylogenetic distance. Moreover, relatively small SynComs could reflect the capacity of the whole community due to metabolic redundancy. Inspection of 30 specific end-product metabolites (i.e., target metabolites: amino acids, vitamins, phytohormones) indicated that the majority of the strains had the genetic potential to produce almost all the targeted compounds. Their production was predicted (1) to depend on external nutritional constraints and (2) to be facilitated by nutritional constraints mimicking root exudates, suggesting nutrient availability and root exudates play a key role in determining the number of producible metabolites. An answer set programming solver enabled the identification of numerous combinations of strains predicted to depend on each other to produce these targeted compounds under severe nutritional constraints thus indicating a putative sub-community level of functional redundancy. Conclusions: This study predicts metabolic restrictions caused by available nutrients in the environment. By extension, it highlights the importance of the environment for niche potential, realization, partitioning, and overlap. Our results also suggest that metabolic dependencies and cooperation among root microbiota members compensate for environmental constraints and help maintain co-existence in complex microbial communities