Browsing by Person "Hammer, Timo"

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Untersuchungen zum asexuellen Gentransfer bei biotrophen Oomyceten anhand der Fallbeispiele Plasmopara halstedii und Peronospora tabacina
(2009) Hammer, Timo; Spring, Otmar
Evidence for gene transfer during the asexual life cycle of certain biotrophic oomycetes was searched for in this study in order to evaluate the potential impact of such parasexual recombination on the variability of these important plant pathogens. Therefore, two case studies with Plasmopara halstedii, the causal agent of sunflower downy mildew, and with the tobacco blue mould pathogen, Peronospora tabacina, were conducted. Although the life cycles of both pathogens lack the possibility of genetic recombination, the organisms differ significantly in their variability. Using molecular methods, several indications for interspecific parasexual recombination between the near relatives Plasmopara halstedii and Plasmopara angustiterminalis were found, giving a possible explanation for the unexpected variability of these pathogens. Asexually formed offspring from dual infection experiments with the two Plasmopara species showed pheno- and genotypic parental traits in new combination and could be cultivated under double selective pressure. The recombinant strain ?R? was studied over 30 generations. Up to the 9th generation and after single sporangium infection, nuclear and mitochondrial traits of both parents were detected in ?R?, indicating a heterokaryon with nuclei and mitochondria of both parental strains. Starting with the 10th generation only fungicide-resistance remained as Pl. halstedii-specific trait, whereas all other detected signals were Pl. angustiterminalis-specific, which indicated true genetic recombination. As one possible mechanism for the genetic exchange it was shown that nuclei can be exchanged between neighboured hyphae via anastomoses and that more than one nucleus can be distributed into one developing sporangium. To prove anastomoses between the different Plasmopara species, specific optical labelling of the participating hyphae is a prerequisit. However, only transient expression of an optical marker gene was achieved, yet. Additional experiments for establishing a stable transformation system were conducted, but were not yet successful in selecting transgenic strains. In contrast to the study on Plasmopara, there was no evidence for recombination in Peronospora tabacina. Neither anastomoses nor heterokaryotic sporangia were found throughout the study. The results are concordant with the findings that tobacco blue mould shows very low variability and that only two phenotypes are known so far from studies in Europe. As the two types of the pathogen, which differ in fungicide sensitivity, did not interact during parallel infection of the same host tissue, they were characterized in detail. Several new pheno- and genotypic differences could be revealed, showing the genetic distance between the two types of the pathogen. A simple PCR detection system to differentiate the two genotypes was created and more than 50 European isolates of tobacco blue mould were monitored.