Processus et mécanismes centrés sur l'inoculum : une approche fonctionnelle et expérimentale de l'épidémiologie végétale

Mon mémoire d'HDR se decompose en cinq parties : la description de mon parcours scientifique ; une synthèse de mes travaux de recherche menés de 2000 à 2015, déclinés en quatre chapitres selon les thématiques et les questions abordées ; mon projet de recherche et d'animation scientifique collective pour les années à venir ; les références bibliographiques ; une sélection de cinq publications majeures en annexe. Au sein de l'équipe d'épidémiologie de l'UMR BIOGER, mes recherches se déclinent actuellement en trois axes : (1) les processus épidémiques, l'initiation et la récurrence des épidémies de septoriose du blé ; (2) la réponse à la temperature de populations de l'agent pathogène Zymospetoria tritici responsable de la septoriose, ses consequences épidémiologiques et le potential d'adaptation au changement climatique : (3) l'épidémiologie fonctionnelle appliquée à la biosécurité, à l'analyse de risques et à la gestion de crises sanitaires en santé végétale

Spatially explicit ecological modeling improves empirical characterization of plant pathogen dispersal

Dispersal is a key ecological process, but it remains difficult to measure. By recording numbers of dispersed individuals at different distances from the source, one acquires a dispersal gradient. Dispersal gradients contain information on dispersal, but they are influenced by the spatial extent of the source. How can we separate the two contributions to extract knowledge about dispersal? One could use a small, point-like source for which a dispersal gradient represents a dispersal kernel, which quantifies the probability of an individual dispersal event from a source to a destination. However, the validity of this approximation cannot be established before conducting measurements. This represents a key challenge hindering progress in characterization of dispersal. To overcome it, we formulated a theory that incorporates the spatial extent of sources to estimate dispersal kernels from dispersal gradients. Using this theory, we re-analyzed published dispersal gradients for three major plant pathogens. We demonstrated that the three pathogens disperse over substantially shorter distances compared to conventional estimates. This method will allow the researchers to re-analyze a vast number of existing dispersal gradients to improve our knowledge about dispersal. The improved knowledge has potential to advance our understanding of species' range expansions and shifts, and inform management of weeds and diseases in crops

Measuring splash-dispersal of a major wheat pathogen in the field

Capacity for dispersal is a fundamental fitness component of plant pathogens. Characterization of plant pathogen dispersal is important for understanding how pathogen populations change in time and space. We devised a systematic approach to measure and analyze rain splash-driven dispersal of plant pathogens in field conditions, using the major fungal wheat pathogen Zymoseptoria tritici as a case study. We inoculated field plots of wheat (Triticum aestivum) with two distinct Z. tritici strains. Next, we measured disease intensity as counts of fruiting bodies (pycnidia) using automated image analysis. These measurements characterized primary disease gradients, which we used to estimate effective dispersal of the pathogen population. Genotyping of re-isolated pathogen strains with strain-specific PCR-reaction confirmed the conclusions drawn from phenotypic data. Consistently with controlled environment studies, we found that the characteristic scale of dispersal is tens of centimeters. We analyzed the data using a spatially-explicit mathematical model that incorporates the spatial extent of the source, rather than assuming a point source, which allows for a more accurate estimation of dispersal kernels. We employed bootstrapping methods for statistical testing and adopted a two-dimensional hypotheses test based on kernel density estimation, enabling more robust inference compared to standard methods. We report the first estimates of dispersal kernels of the pathogen in field conditions. However, repeating the experiment in other environments would lead to more robust conclusions. We put forward advanced methodology that paves the way to further measurements of plant pathogen dispersal in field conditions, which can inform spatially targeted plant disease management

Evolution within a given virulence phenotype (pathotype) is driven by changes in aggressiveness: a case study of French wheat leaf rust populations

Plant pathogens are constantly evolving and adapting to their environment, including their host. Virulence alleles emerge, and then increase, and sometimes decrease in frequency within pathogen populations in response to the fluctuating selection pressures imposed by the deployment of resistance genes. In some cases, these strong selection pressures cannot fully explain the evolution observed in pathogen populations. A previous study on the French population of Puccinia triticina, the causal agent of wheat leaf rust, showed that two major pathotypes — groups of isolates with a particular combination of virulences — predominated but then declined over the 2005-2016 period. The relative dynamics and the domination of these two pathotypes — 166 317 0 and 106 314 0 —, relative to the other pathotypes present in the population at a low frequency although compatible, i.e. virulent on several varieties deployed, could not be explained solely by the frequency of Lr genes in the landscape. Within these two pathotypes, we identified two main genotypes that emerged in succession. We assessed three components of aggressiveness — infection efficiency, latency period and sporulation capacity — for 44 isolates representative of the four P. triticina pathotype-genotype combinations. We showed, for both pathotypes, that the more recent genotypes were more aggressive than the older ones. Our findings were highly consistent for the various components of aggressiveness for pathotype 166 317 0 grown on Michigan Amber — a ‘naive’ cultivar never grown in the landscape — or on Apache — a ‘neutral’ cultivar, which does not affect the pathotype frequency in the landscape and therefore was postulated to have no or minor selection effect on the population composition. For pathotype 106 314 0, the most recent genotype had a shorter latency period on several of the cultivars most frequently grown in the landscape, but not on ‘neutral’ and ‘naive’ cultivars. We conclude that the quantitative components of aggressiveness can be significant drivers of evolution in pathogen populations. A gain in aggressiveness stopped the decline in frequency of a pathotype, and subsequently allowed an increase in frequency of this pathotype in the pathogen population, providing evidence that adaptation to a changing varietal landscape not only affects virulence but can also lead to changes in aggressiveness

PLoS Pathog

Kaposi's sarcoma herpesvirus (KSHV) encodes a cluster of twelve micro (mi)RNAs, which are abundantly expressed during both latent and lytic infection. Previous studies reported that KSHV is able to inhibit apoptosis during latent infection; we thus tested the involvement of viral miRNAs in this process. We found that both HEK293 epithelial cells and DG75 cells stably expressing KSHV miRNAs were protected from apoptosis. Potential cellular targets that were significantly down-regulated upon KSHV miRNAs expression were identified by microarray profiling. Among them, we validated by luciferase reporter assays, quantitative PCR and western blotting caspase 3 (Casp3), a critical factor for the control of apoptosis. Using site-directed mutagenesis, we found that three KSHV miRNAs, miR-K12-1, 3 and 4-3p, were responsible for the targeting of Casp3. Specific inhibition of these miRNAs in KSHV-infected cells resulted in increased expression levels of endogenous Casp3 and enhanced apoptosis. Altogether, our results suggest that KSHV miRNAs directly participate in the previously reported inhibition of apoptosis by the virus, and are thus likely to play a role in KSHV-induced oncogenesis

Kaposi's Sarcoma Herpesvirus microRNAs Target Caspase 3 and Regulate Apoptosis

Kaposi's sarcoma herpesvirus (KSHV) encodes a cluster of twelve micro (mi)RNAs, which are abundantly expressed during both latent and lytic infection. Previous studies reported that KSHV is able to inhibit apoptosis during latent infection; we thus tested the involvement of viral miRNAs in this process. We found that both HEK293 epithelial cells and DG75 cells stably expressing KSHV miRNAs were protected from apoptosis. Potential cellular targets that were significantly down-regulated upon KSHV miRNAs expression were identified by microarray profiling. Among them, we validated by luciferase reporter assays, quantitative PCR and western blotting caspase 3 (Casp3), a critical factor for the control of apoptosis. Using site-directed mutagenesis, we found that three KSHV miRNAs, miR-K12-1, 3 and 4-3p, were responsible for the targeting of Casp3. Specific inhibition of these miRNAs in KSHV-infected cells resulted in increased expression levels of endogenous Casp3 and enhanced apoptosis. Altogether, our results suggest that KSHV miRNAs directly participate in the previously reported inhibition of apoptosis by the virus, and are thus likely to play a role in KSHV-induced oncogenesis