Basal resistance of barley to adapted and non-adapted forms of Blumeria graminis

In the barley-Blumeria interaction, resistance at penetration stage in association with papilla formation is a commonly occurring mechanism. This mechanism of defense reduces the infection severity by adapted powdery mildew pathogen (basal resistance to Blumeria graminis f.sp. hordei, Bgh) and fully protects the plant against non-adapted powdery mildew pathogens (non-host resistance to non-adapted forms of B. graminis). In this thesis we followed an integrated approach based on QTL mapping and candidate gene analysis. It was our objective to determine which of the known candidate genes have a map position that coincides with one of the QTLs detected in our mapping study. Such candidate genes might determine the natural variation of basal resistance against powdery mildew in barley. We also investigated whether the genotypic variation in level of resistance of barley to the non-adapted wheat powdery mildew, B. graminis f.sp. tritici (Bgt) could be used to develop an experimental line to determine the inheritance of non-host resistance of barley to this pathogen. Finally we aimed to determine whether the basal resistance of barley to powdery mildew is being suppressed by prior attack by a different pathogenic haustorium forming fungus, Puccinia hordei. Chapter 1 presents an introduction about basal resistance of barley to powdery mildew and briefly lists evidence on molecular associations between basal, mlo-mediated and non-host resistance. In Chapter 2, we performed QTL analysis for powdery mildew resistance at seedling and at adult plant stage in six mapping populations of barley. In that analysis quantitative resistance of barley to powdery mildew was found to be based on a large number of genes, with only few detected in more than one mapping population. These QTLs for powdery mildew resistance in barley are mainly plant growth stage dependent. From gene-expression, gene silencing and transient transformation assays 39 genes emerged that increased or decreased basal resistance of barley to the powdery mildew fungus. They are considered candidate genes that might be responsible for natural variation in level of basal resistance. By looking for polymorphism in those genes, we could determine the map position of 23 of these candidate genes. Mapping positions of three more genes, mlo, Ror1 and Ror2, were already available, and these were also considered as candidate genes in our study.To compare the map position of QTLs in different mapping populations with the map positions of the 26 candidate genes, we constructed an improved high-density integrated linkage map of barley. This improved map was based on the marker data of seven mapping populations of barley. More than 43% of the markers on our improved integrated map target ESTs or gene sequences (gene-targeted markers, GTM) and can be used for candidate gene identification. Six out of the 26 candidate genes co-localized with QTLs for basal powdery mildew resistance. They are interesting targets for further studies, since their allelic forms may be responsible for part of the differences in basal resistance between the parents in our mapping study. Chapter 3 reports the development of two experimental barley lines with extremely high and low level of basal resistance to barley powdery mildew. These lines were obtained by convergent crossing between the most resistant and the most susceptible lines, respectively, from four mapping populations of barley studied in Chapter 2. We consider the extremely susceptible and resistant lines developed here as valuable material to be used in further experiments to characterize the molecular basis of basal resistance to powdery mildew. The results suggest that phenotypic selection is sufficiently efficient to achieve high levels of basal resistance. We report some difficulties in the application of marker assisted selection that might make that approach unnecessary and less efficient to achieve a similar high level of basal resistance. In Chapter 4 the development of two more experimental lines is described. These lines have, at the seedling stage an unprecedented level of susceptibility to the non-adapted wheat powdery mildew. A large collection of barley germplasm was screened and some rare barley accessions were identified with rudimentary susceptibility to the wheat powdery mildew, Bgt. Those accessions were intercrossed in two cycles, and resulted in the two exceptional research lines, called SusBgt SC and SusBgt DC. The quantitative variation among barley accessions and in the progenies after convergent crossing suggest a polygenic basis of this non-host resistance. Component analysis of the infection process suggested that non-host resistance factors are Blumeria-form specific. The developed lines will serve to elucidate the genetic basis of non-host resistance in barley to wheat powdery mildew, and are useful tools in gene expression and complementation studies on non-host resistance. In Chapter 5 we reported that a compatible interaction of barley-Puccinia hordei induces increased papilla based resistance to a challenge infection by a compatible powdery mildew isolate. These pathogens differ in strategies to suppress plant cell defense. We showed that the level of rust-induced mildew resistance varies among barley accessions and is not determined by the virulent/avirulent spectra of the challenger isolate. Macroarray gene expression analysis showed that several genes involved in metabolism and photosynthesis were highly down-regulated and a few genes involved in plant defense including a pathogenesis related protein (PR-1) and cysteine synthase were highly up-regulated in the double inoculated treatment. Our histological and gene expression analyses are compatible with the hypothesis that the rust “primes” the basal mildew resistance genes prior to the challenge mildew infection. In Chapter 6, the results obtained in the previous Chapters are being discussed. The perspectives of the experimental barley lines that were developed in this study for the identification of genes that determine basal resistance of barley to adapted and non-adapted Blumeria graminis forms are highlighted.