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Malus × robusta 5 is a crab apple highly resistant against the disease fire blight attributed to a potential resistance gene at the top of linkage group three. The resistance gene was identified on a 162 kb BAC clone spanning the region of a major QTL. The protein sequence of this gene was further analyzed and classified as CC-NBS-LRR (CNL). By comparing the previously described CNLs of other plants targeting bacterial pathogens and the presence or absence of the interacting proteins, the hypothetical mode of function was suggested to be homolog to the resistance protein RPS2 in Arabidopsis. The remaining predicted genes on the BAC sequence were annotated. Additional potential candidate resistance genes were analyzed in detail, but their predicted function could not be verified.

Fire blight is one of the most damaging diseases in the majority of apple (Malus x domestica) producing countries worldwide. The region of the quantitative trait for resistance to Erwinia amylovora and the same data mapped as single gene in the wild apple Malus x robusta 5 was enriched with additional molecular markers. Generated phenotypic data of individuals that carry a crossing over in the region of interest allowed to position the locus coding for resistance into a short section at the top of linkage group 3 flanked by molecular markers.

Fire blight (FB) is the most important bacterial disease in apple and pear orchards. The disease is caused by Erwinia amylovora and difficult to control because the application of antibiotics is either banned or undesired in many apple production areas. Current breeding programs introgress strong resistance from wild sources such as Malus × robusta 5 (MR5). Previous work attributed the resistance derived from this accession to a QTL located on top of linkage group 3. We enriched this region with molecular markers and isolated the region by means of a genomic library. The genomic sequence encompassed by two flanking markers was analysed in silico and led to the identification of a gene with structural homology to known bacterial resistance genes in model plants. It is differentially expressed in fire blight resistant and susceptible plants, respectively. We therefore indicate it as candidate fire blight resistance gene MR5.

The rapid and effective activation of disease resistance responses is essential for plant defense against pathogen attack. These responses are initiated when pathogen-derived molecules (elicitors) are recognized by the host. In order to create novel mechanisms for fire blight resistance in pear, we have generated transgenic pears expressing the elicitor harpin Nea from Erwinia amylovora under the control of the constitutive promoter CaMV35S. The transient expression of hrpN Ea in pear cells did not provoke any apparent damage. Therefore, stable constitutive expression of hrpN Ea was studied in seventeen transgenic clones of the very susceptible cultivar "Passe Crassane.” Most transgenic clones displayed significant reduction of susceptibility to fire blight in vitro when inoculated by E. amylovora, which was positively correlated to their degree of expression of the transgene hrpN Ea . These results indicate that ectopic expression of a bacterial elicitor such as harpin Nea is a promising way to improve pear resistance to fire blight

Powdery mildew (PM) is a major fungal disease that threatens thousands of plant species. PM is caused by Podosphaera leucotricha in apple and Erysiphe necator in grapevine. Powdery mildew is controlled by frequent applications of fungicides, having negative effects on the environment, and leading to additional costs for growers. To reduce the amount of chemicals required to control this pathogen, the development of resistant apple and grapevine varieties should become a priority. PM pathogenesis is associated with up-regulation of specific MLO genes during early stages of infection, causing down-regulation of plant defense pathways. These up-regulated genes are responsible for PM susceptibility (S-genes) and their knock-out causes durable and broad-spectrum resistance. All MLO S-genes of dicots belong to the phylogenetic clade V. In grapevine, four genes belong to clade V. VvMLO7, 11 and 13 are up-regulated during PM infection, while VvMLO6 is not. Chapter 2 reports the genome-wide characterization and sequence analysis of the MLO gene family in apple, peach and woodland strawberry, and the isolation of apricot MLO homologs. Twenty-one homologues were found in apple, 19 in peach and 17 in woodland strawberry. Evolutionary relationships between MLO homologs were studied and syntenic blocks constructed. Candidate genes for causing PM susceptibility were inferred by phylogenetic relationships with functionally characterized MLO genes and, in apple, by monitoring their expression following inoculation with the PM causal pathogen P. leucotricha. In apple, clade V genes MdMLO11 and 19 were up-regulated, whereas the two other members of clade V, MdMLO5 and 7, were not up-regulated. The clade VII gene MdMLO18 was also up-regulated upon P. leucotricha infection. Chapter 3 reports the knock-down, through RNA interference, of MdMLO11 and 19, as well as complementation of the mutant phenotype by expression of the MdMLO18 gene in the Arabidopsis thaliana triple mlo mutant Atmlo2

Background Powdery mildew (PM) is a major fungal disease of thousands of plant species, including many cultivated Rosaceae. PM pathogenesis is associated with up-regulation of MLO genes during early stages of infection, causing down-regulation of plant defense pathways. Specific members of the MLO gene family act as PM-susceptibility genes, as their loss-of-function mutations grant durable and broad-spectrum resistance. Results We carried out a genome-wide characterization of the MLO gene family in apple, peach and strawberry, and we isolated apricot MLO homologs through a PCR-approach. Evolutionary relationships between MLO homologs were studied and syntenic blocks constructed. Homologs that are candidates for being PM susceptibility genes were inferred by phylogenetic relationships with functionally characterized MLO genes and, in apple, by monitoring their expression following inoculation with the PM causal pathogen Podosphaera leucotricha. Conclusions Genomic tools available for Rosaceae were exploited in order to characterize the MLO gene family. Candidate MLO susceptibility genes were identified. In follow-up studies it can be investigated whether silencing or a loss-of-function mutations in one or more of these candidate genes leads to PM resistance.