Kurzbeschreibung
(Englisch)
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Fungal diseases of grapevine are responsible for important economical losses. This project, divided in three part, is dedicated to the investigation of the interaction of the fungi causing Esca, a very important trunk wood disease, the toxins production, the resistance mechanism and the biological control of the disease. The identification of the resistance mechanisms and factors affecting the level of the resistance against Plasmopara viticola, the agent of downy mildew , a leave and berry disease, will also be investigated.
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Partner und Internationale Organisationen
(Englisch)
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ETH Zürich Agroscope RAC Changins EIL, HES-SO, Jussy AT, CH, CY, CZ, DE, ES, FR, GR, HU, IL, IT, LU, PL, PT, RS, SE, SI
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Abstract
(Englisch)
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Project A: Our research aims to understand the multiple factors involved in the sudden apoplectic death of grape vine plants, also known as esca. Worldwide, the disease complex of esca is increasing and no direct treatments can be applied to control the disease. In a first approach all the fungi found in diseased grapevine trunks were isolated, identified by morphological and molecular methods and conserved in a collection. From seventy apoplectic trunks, 339 fungal colonies were isolated and sub-cultured. Among them 42 different fungal species could be identified morphologically and by blast of the rDNA sequences. The association of E. lata with other fungi and the complexity of the microflora present in black dead arm-, young esca-, and esca-infected grapevine wood can play a crucial role in the expression of the diseases. Taking in account that a considerable variations in aggressiveness were observed in the genetically diverse populations of Eutypa lata from Europe and Australia, or from a single vineyard, and similarly in the populations of Phaeoacremonium, and the fact that the microorganisms have to co-habit the host tissues should also have an effect on the virulence of the protagonists: increased competition. In this study, we consider that an increase in virulence could be translated into an increase in the synthesis/ secretion of toxic compounds. We consider that fungi can synthesis incidental phytotoxins, as an antifungal agent to win the race against other colonisers. Project B : Eutypa dieback and esca disease are grapevine trunk diseases leading after long latency times to the decline and eventually to the death of the vines by congesting the xylem vessels. It is thought that the aeral symptoms are due to fungal substances transported fom the discoloured wood to the leaves through the transpiration stream. Some toxins have already been identified. No Vitis cultivar is resistant to trunk diseases, but Merlot is rather tolerant and is thought to possess a specific aldehyde reductase responsible for detoxification. First, we have developped a non-invasive field test for the early detection of esca disease before the manifestation of the foliar symptoms. The characteristics of fast chlorophyll a fluorescence transient were investigated in attached leaves by using the so-called JIP-test. The fluorescence transient was analyzed and plants with or without visible esca foliar symptoms were compared (visual assessment, necrosis and white rot indexes). In the susceptible C. sauvignon, an alteration of the photosynthetic apparatus could be detected two months before the appearance of foliar symptoms in autumn. In the tolerant Merlot, only one parameter was affected. A comparison of the relationship of the performance index versus the calculated rate of the electron transfer allowed us to separate the tolerant cultivar from the susceptible one. Also, the method detected modifications in greenhouse-grown Riesling x Sylvaner after drought stress. A comparison drought-stressed with esca-infected grapevine suggests that esca infection cannot simply be interpreted as a water transport deficit, but that other reaction mechanisms in the plants must be considered. Second we have looked at the role played by the natural host defences against esca and eutypa dieback pathogen toxins. The cytotoxicity of eutypine, 4-hydroxybenzaldehyde, juglone, and 3-phenyllactic acid was tested on grapevine cell suspensions and on detached leaves. On leaves, all toxins reproduced the symptoms observed in the field (necrosis and chlorosis), as well as a decrease of the water uptake, and an inhibition of the root formation. On cell suspensions, the expression of genes involved in defence, signalling, detoxification was quantified by real-Time RT-PCR. Our results highlight a possible involvement of glutathione-S-transferase in the detoxification of the toxins, of the jasmonic acid signalling pathway, and of several effector genes underlying a more general response where the toxin could be recognised as an elicitor for the trunk pathogens. Further, we tested whether low concentration of toxins can stimulate grapevine defences, and protect them against higher concentrations. On cells, death rate after the second treatment was significantly reduced. At the transcriptomic level, a differential expression was observed in pre- and non-pretreated samples. On leaves, the esca-like and physiological symptoms were reduced if the plant received first low dosages of toxin. Moreover, the sensitivity of the leaves to other biotic stresses such as downy mildew infection was strongly reduced. Altogether these results unveil the induction of resistance by and to the toxins themselves. Project C: Project C,The development of techniques to test the functional importance of single genes in the defense response is crucial. Since genetic transformation is a tedious and time-consuming process, many groups are testing possible methods of gene silencing (RNA interference). We have adapted to grapevine and tested a method of infiltration of double-stranded RNA into leaves which would allow for an easy testing of genes. However the preliminary results showed a limited success with insufficient spreading of the silencing effect to allow the testing of defense reactions in leaves. The work is continuing now with a virus-based approach (VIGS, virus-induced gene silencing).
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