Erforschung und Entwicklung alternativer Mittelzubereitungen für die Apfelschorfbekämpfung im Falllaub

Abstract

The objective of the study was to achieve a phytosanitary effect on fruiting bodies of V. inaequalis in apple leaf litter by application of nutrient media, potential antagonists and/or fungicidal plant extracts (saponins). The effects of leaf litter treatments on the remaining ascospore potential were assessed in course of the primary season of the pathogen. The principles and mode of action of the applied substances were evaluated by microbiological and biochemical analyses. As an alternative method for the isolation of saponins from plant materials an extraction with chloroform and methanol was developed. The saponin extracts were cleaned with SPE cartridges and for the first time with Hydrophobic Interaction Chromatography. Nearly all ineffective substances could be removed from the extracts. The detection of membranolytic/hemolytic fractions was performed with an agar diffusion assay amended with sheep erythrocytes. Hemolytic fractions were further characterized by HPLC analyses. In vitro all plant extracts showed a fungicidal effect on conidia germination and mycelia growth of V. inaequalis. In seedling assays, a preventative treatment with soap nut and chestnut extract (1%) reduced apple scab symptoms nearly completely. A treatment 6 h after inoculation with conidia reduced scab symptoms by more than 70%. Pathogen sporulation on infected leaves was reduced by 98-100% in comparison to control groups. Ascospore potential could be reduced by leaf litter treatments by up to 90%. The application of in vitro identified antagonists on apple scab litter showed no effect on ascospore amount. Leaf litter treatments with nutrient media in autumn/winter revealed a reduction in the ascospore potential of more than 97% in four following apple scab seasons. The effect did not depend on geographical location of the orchard site. The best results could be achieved with a concentrated LEIBER yeast extract, which reduced the ascospore amount nearly completely. The microbiological analyses of leaf litter showed an increase of the number of organisms, including potential antagonists, from 10 up to 1.000 times. The microbiological activity (biological oxygen demand) was up to three times higher and leaf decay was enhanced compared to untreated litter. Bacterial and fungal leaf colonizers were isolated and identified with MALDI-TOF or sequencing of ITS rDNA. Pseudomonas proved to be the dominant bacterial species in apple leaf litter and contributed 37 to 45% of bacterial isolates. In vitro an inhibition of mycelia growth of V. inaequalis from 48 to 69% was observed. Cladosporium cladosporioides, Aureobasidium pullulans and Epicoccum nigrum reduced mycelial growth by up to 57%. Each one of these species represented more than 10% of the fungal isolates. In the majority of samples the microbial composition was changed mainly quantitatively and a shift within the dominant species was not detectable. Peptides ≤ 3 kDa and amino acids could be identified as the effective constituents of the media. Leaf litter treatments with nutrient media could be established as a new phytosanitary strategy in organic fruit production and contribute to a replacement of coppere fungicides.