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No description available.

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No description available.

No description available.

Latent B. cinerea was detected in nine symptomless wild host species from the families Asteraceae and Brassicaceae, in addition to greenhouse grown lettuce. Conventional testing methods revealed that latent B. cinerea was equally prevalent in the root system as the above ground parts. Incidence of latent infection was moderate in some species (Achillea milleforlium, Arabidopsis thaliana, Centraurea nigra, Cirsium vulgare, Senecio jacobaea, Senecio vulgaris and Taraaxacum agg.) and rare in others (Tussilago farfara and Bellis perennis). In greenhouse lettuce, latent infection was activated by prolonged water stress and artificial inoculation. Despite inoculation, unstressed, vigorously growing lettuce and Arabidopsis plants remained asymptomatic throughout the growing period. Fungicide seed treatment did not significantly affect the amount of latent B. cinerea recovered from the lettuce plants. Introduction of antagonistic micro-organism Trichoderma harzianum T-39 into the soil decreased the amount of latent infection recovered from lettuce leaves but increased it in the stem. A weak negative correlation was found between photosynthesis and the amount of B. cinerea recovered from the leaves. Weight of the plants was reduced due to inoculation of B. cinerea even though latent infection was unaltered. There was no relation between plant weight and total endophytic B. cinerea. A marginal increase of the phenolic contents of the leaf was observed due to inoculation, but no changes to the antioxidant activity, chlorophyll content or carotenoids were found. The high incidence of latent infection found in greenhouse grown lettuce plants with or without successful inoculation may have been due to the presence of several genetically distinct isolates of B. cinerea. Eight different haplotypes were identified among the 32 isolates assessed. A single very common haplotype presumably originated from seed borne infection, because it was rare in plants grown from fungicide treated seed. Latency may be attributed to a mild strain defence response by the presence of several genetically different strains of the pathogen present within the plant as endophytes.

Understanding the molecular mechanisms underpinning the ecological success of plant pathogens is critical to develop strategies for controlling diseases and protecting of crops. Recent observations have shown that plant pathogenic bacteria are not just interacting with the plant itself, but with a variety of potential predators present in the soil, such as nematodes, aphids, larvae and amoebae. Bacteria are therefore under strong selective pressure to avoid or survive predation and I hypothesise that bacteria have evolved mechanisms to escape predation. To identify the gene systems that contribute to the ecological success of P. syringae, with a specific focus on anti-predation and pathogenicity mechanisms I have used a Rapid Virulence Annotation (RVA) screening. Three cosmid libraries for P. syringae pv. aesculi, pv. tomato and pv. phaseolicola comprising approximately 2000 cosmids each were screened and a number of potential genes involved in bacterial survival identified. These included also genes encoding for Type 6 secretion system, hemolysins and biofilm formation, motility and adhesion. A comparison of the genes conferring resistance within other P. syringae strains showed differences in the organization of these clusters, indicating evolutionary changes. These data provide an important understanding of how bacteria cope with different biotic pressures and to give insight to the function of the genes and their role in ecological success. Objective of this thesis was also to characterize virulence mechanisms of P. syringae pv. aesculi the agent of the bleeding canker disease of Horse Chestnut. In this thesis I characterized the origin of this pathogen, its evolution and the factors that allow it to cause disease, focussing on the Type 3 Secretion System. hopAB1 gene was identified to be the main determinant in virulence both in N. benthamiana and A. hippocastanum. Analysis and expression in Pae of hopAB1 orthologues allowed the definition of two amino acids responsible successful virulence.

The three different platform candidates were antibody microarray, surface Plasmon resonance (SPR) and bead array. Another objective of this project was to explore other applications of these platforms, and the selected example application was to explore other applications of these platforms for hybridoma screening. Since antibodies against plant pathogens have previously been produced, characterized and used in development of the three different platforms, to examine the possibility of applying the platforms to facilitate the screening of hybrid om a, pilhogenic foodbome bacterium Listeria monocytogenes was used as a model study. In summary, antibody array and bead array technologies were demonstrated to be able to detect multiple plant pathogens at the same time. Both were shown to be able to detect the pathogens in the real samples from fields. making them good candidates for plant industries. On the other hand, despite considerable effort to incr¢ase the sensiritivity of SPR, lower limit of detection could not be achieved. This courd be an intrinsic limitation of this method for detecting a whole cell where the size of the analytic is bigger than usual SPR analytes such as small compounds. From these technologies. an antibody array was selected for hybridoma screening and was shown to be effective, albeit with lower sensitivity compared to ELISA. The antibody array proved to be a faster and simpler method to screen hundreds of hybricioma 3 clones. A comnbination of biosensor tcchnology ensures good quality screening for hybridoma clones as demonstrated in this study.

La vigne (Vitis vinifera) est sensible à un grand nombre de maladies. Les politiques de limitation des traitements phytosanitaires font qu'aujourd'hui, les viticulteurs ne disposent d'aucun moyen de lutte contre certains bioagresseurs. C'est le cas des maladies du bois causées par des complexes fongiques nécrosant les ceps et du court noué, maladie virale transmise par des nématodes. La vigne est une espèce végétale connue pour sa production particulière de métabolites secondaires en réponse à des infections. Dans le cadre de cette thèse, nous nous sommes intéressés aux phytoanticipines, préformées dans les plantes et dont le potentiel de toxicité envers des agents pathogènes est intéressant, c’est le cas des triterpénoïdes. Les objectifs de cette thèse ont donc été dans un premier temps d'estimer la capacité de la vigne à produire de tels composés. Pour cela, la composition en triterpénoïdes de différents organes et de différents cépages a été analysée. Nous avons pu montrer que la composition générale en triterpénoïdes est caractéristique de chaque cépage et de chaque organe. Dans une deuxième partie, 9 gènes impliqués dans la synthèse de triterpènes chez la vigne ont été identifiés et leur expression a été évaluée dans différents organes, chez différents cépages et sous différentes conditions de stress biotique et abiotique. Cette étude exploratoire nous donne des pistes pour mettre en corrélation l'expression de certaines triterpène synthases avec la production différentielle de certains triterpènes à la surface des feuilles de différents cépages. Enfin, nous nous sommes intéressés aux triterpènes glycosylés, les saponines, afin d'évaluer leur potentiel dans la lutte contre certaines maladies majeures de la vigne pour lesquelles aucun traitement n'est actuellement disponible. Pour cela, l'efficacité de saponines issues de la gypsophile et du quillaja a été testée contre certains champignons associés aux maladies du bois ainsi que contre les nématodes vecteurs des virus du court noué. Nous avons pu mettre en évidence que les souches de champignons testées étaient capables de contourner la toxicité des saponines, tandis qu'un tel traitement était rapidement efficace pour lutter contre les nématodes. Afin de vérifier l'innocuité de ce traitement pour l'environnement, les doses efficaces ont été testées et n'ont pas eu d'impact significatif sur différents bioindicateurs. / Vitis vinifera is susceptible to many pathogens. These past few years, treatment policies led to the withdrawal of many pesticides. Renee, no chemical treatments are available to treat some grapevine diseases such as the grapevine trunk diseases caused by fungi complexes and the grapevine fanleaf degeneration, a viral disease transmitted from grapevine to grapevine by vector nematodes. Grapevine is known for the production of secondary metabolites as a response to pathogen infections. In this work, we focused on phytoanticipins such as triterpenoids, that are found as preformed compounds and that confer a basal resistance level to plants. First, a chemical analysis was made on the triterpenoid composition of some grapevine cultivars and organs. This study revealed that the triterpenoid composition is specific to the V. vinifera cultivar and the organ. In a genomic approach, 9 candidate genes involved in the triterpene biosynthesis were identified and their expression was studied in different organs, varieties and biotic or abiotic stress conditions. This explorative study shows correlations between gene expression and differential triterpene production at the leaf surface of the different varieties. In the last part of this study, the use of glycosylated triterpenes, also called saponins, as a substitution solution to withdrawed treattnents against major grapevine diseases was tested. Therefore, the efficiency of saponins extracted from gypsophila and quillaja was tested against fungi associated to grapevine trunk diseases and some nematodes vector of the grapevine fanleaf degeneration. These tests evidenced that the fungi were able to avoid saponins toxicity, whereas such treatment was efficient to kill nematodes. In order to evaluate the effect of the treatment on the environment, the efficient doses were tested and bad no significant impact on some bioindicators.