Bernadette Dubos’s research while affiliated with Chouaib Doukkali University and other places

Effects of Eutypa lata, the fungal agent of eutypiosis, on polyamine (putrescine (Put), spermidine (Spd), spermine (Spm), diaminopropane (Dap)) and tyramine (Tyr) contents in grapevine leaves were studied to determine the relationship between these components and disease development. The external symptoms of eutypiosis are small and necrotic leaves, which could be observed on only one arm or on two arms of the vine stock. This study was carried out on healthy leaves (S, borne by healthy vine stock), apparently healthy leaves (Aps, borne by one arm without symptoms, whereas the other arm was diseased), and moderately (M. mal.) and strongly (F. mal.) diseased leaves. We analysed free polyamines (free PAs, chlorhydric acid soluble fraction), conjugated polyamines (PAs-fpm, PAs covalently linked to molecules with low molecular mass like hydroxycinnamic acids), and bound polyamines (PAs-mm, covalently linked to macromolecules) in Vitis vinifera L. 'Cabernet sauvignon', a variety sensitive to eutypiosis. In diseased leaves, we noted a decrease of free PAs, PAs-fpm, free Tyr, and Tyr-fpm levels. By contrast, the Dap-fpm level was increased by the disease. Eutypiosis also induced an increase of Spm-mm and Dap-mm levels, whereas the Put-mm and Spd-mm levels were decreased. Free Put, free Spd, PAs-fpm, free Tyr, and Tyr-fpm levels were higher in apparently healthy leaves than in healthy leaves. The PAs-mm levels were slightly affected by eutypiosis in apparently healthy leaves. The specific role of each type of polyamines and tyramine is discussed with regard to the host-parasite relationship.Key words: polyamines, leaf, Eutypa lata, Vitis vinifera.

Résumé: Des dosages d'acide abscissique libre (ABA) et conjugué (abscissate de β-D-glucopyranose : ABA-GE) ont été réa-lisés dans les feuilles, entre-noeuds, boutons floraux, fleurs et jeunes baies de vigne (Vitis vinifera L. cv. Cabernet-Sauvignon) issus de vignes saine et infectée par Eutypa lata, l'agent causal de l'eutypiose de la vigne. En général, la croissance, le déve-loppement et les teneurs en eau et en ABA des organes ne présentant pas de symptômes d'eutypiose, mais portés par des pieds de vigne infectés par E. lata ne sont pas ou peu différents de ceux des organes portés par des pieds sains. Les organes présen-tant des symptômes de la maladie (réduction de la croissance) sont déshydratés par rapport aux organes sains. Pendant toute la période étudiée pour les feuilles et à partir du stade fécondation pour les entre-noeuds, les teneurs en ABA et en ABA-GE sont augmentées par l'eutypiose. Cette maladie retarde aussi l'apparition du maximum d'ABA caractérisant les fleurs et réduit sa valeur. De même, elle provoque un appauvrissement en ABA-GE dans les boutons floraux, les fleurs et les jeunes baies. Ces différents dérèglements sont d'autant plus intenses que les symptômes sont plus marqués. L'étude du contenu en ABA dans le mycélium et le filtrat de culture du champignon montre qu'E. lata est capable de synthétiser ce régulateur de croissance et qu'il est capable d'en libérer dans le milieu extérieur. L'origine de l'ABA accumulé dans les organes malades et l'implication de cet enrichissement dans l'apparition des symptômes sont discutées. Abstract: In grapevine, eutypa dieback is a disease induced by ascomyceta fungus, Eutypa lata. At present, eutypa dieback is considered as the most serious deterioration disease of grapevine. The external symptoms of this disease are most conspi-cuous during the first months of the annual growth cycle and include dwarfed shoots with smaller and necrotic leaves and unfa-vourable development of grapes. All these symptoms were still suggested an hormonal perturbation in grapevine organs. The aim of this work was to determine the effects of eutypa dieback on abscisic acid contents in flower buds, flowers, young ber-ries, leaves and internodes of grapevine. This study was also performed to determine the relationship between this growth regu-lator and disease development. The study was performed using organs (1) from healthy vines, (2) healthy appearing organs from vines with one symptomless arm and one diseased arm, and (3) moderately and (4) strongly diseased organs. From flo-wers buds separated to bunch of grapes closed stages, free abscisic acid (ABA) and the glucose esters of this acid (ABA-GE) were analysed in Vitis vinifera L. cv. Cabernet Sauvignon which is sensitive to E. lata. In the healthy appearing organs, growth and development as well as water and ABA contents were not changed by eutypa dieback fungi. Whereas eutypa dieback induced a decrease of water content and the rate of growth increase in diseased organs. During all the period studied in leaves and since fruit set stage in the internodes, the ABA and ABA-GE contents were increa-sed by eutypa dieback proportionally to symptoms. The ABA levels increase in leaves and internodes were not seem to be pro-duced by ABA-GE hydrolysis since increase in ABA-GE levels were also observed in response to eutypa dieback. The maximum of ABA characterising grapevine flowers was delayed by eutypa dieback and their level was reduced with increasing symp-toms severity. In flower buds, flowers and young berries, this disease induced a decrease of ABA-GE contents during all period studied proportionally to symptoms severity. The mycelium of two E. lata isolates cultured in Errikson and Petersson medium culture can synthesize ABA. A part of this growth regulator was diffused in medium culture. It was suggested that increase of ABA levels in diseased organs must have many origins: perturbation of (1) ABA-GE translation, (2) synthesis of ABA by grapevine caused by water stress and (3) synthesis of ABA by the fungi. The possible relationship between increa-sed ABA content in the diseased organs and expression of eutypa dieback symptoms is discussed.

p>Development of polyamines levels [Putrescine (Put), Spermidine (Spd), and Spermine (Spm)] was investigated in internodes of grapevine infected by eutypiosis fungus, Eutypa lata . The study was performed using (1) internodes from healthy vines, (2) apparently healthy internodes from vines with one arm symptomless and one diseased arm and (3) diseased internodes. Free polyamines (PAs: hydrochloric acid soluble fraction), conjugated polyamines (PAs-fpm: PAs covalently linked to molecules with low molecular weight like hydroxycinnamic acids) and bound polyamines (PAs-mm: covalently linked to macromolecules) were analysed in Vitis vinifera L. cv. Cabernet sauvignon, which is sensitive to eutypiosis. The internodes stunting induced by eutypiosis was accompanied by a decrease of free Put and free Spm contents between flower buds separated and fruit set stages. The same results were observed for PAs-mm but more slightly than free PAs. The Put-fpm and Spd-fpm levels increased in the apparently healthy internodes and decreased in diseased organs by Eutypa lata effect. The relationship between free PAs contents in the diseased internodes and their growth reduction, as well as the involvement of PAs-fpm on the grapevine response to Eutypa lata infection are discussed.</p

Evolution des teneurs en polyamines dans les boutons floraux, les fleurs et les jeunes baies de Vitis vinifera L. (cv. Cabernet Sauvignon) atteints d'eutypiose Résumé L'influence de l'eutypiose sur le contenu en polyami-nes (PAs) et en tyramine des boutons floraux des fleurs et des jeunes baies de la vigne a été déterminée afin de préci-ser l'importance de ces composés dans le développement de cette maladie, induite par Eutypa lata. Ce travail a été réa-lisé sur des organes sains (portés par des pieds sains), sur des organes apparemment sains (portés par un bras ne pré-sentant pas de symptômes alors que l'autre bras est ma-lade) et sur des organes malades (présentant des symptô-mes de la maladie). Cette étude a porté sur les PAs libres, conjuguées aux molécules de faibles poids moléculaires, (PAs-fpm) et liées aux macromolécules (PAs-mm) chez un cépage de Vitis vinifera L. sensible à cette maladie, le Ca-bernet Sauvignon. Dans les organes malades, l'eutypiose provoque une augmentation des PAs-mm et une diminution des PAs libres, perturbation qui concerne essentiellement le diaminopropane et la putrescine. Les teneurs de ces deux dernières formes de PAs ne sont pas influencées par l'eutypiose dans les organes apparemment sains. Le con-tenu en PAs-fpm des différents organes étudiés a égale-ment été perturbé par la maladie. Ces derniers composés, qui auraient un rôle important dans la réaction de la plante contre les agressions parasitaires, sont faiblement pré-sents dans les organes malades par rapport aux organes sains. Paradoxalement, les organes apparemment sains contiennent plus de PAs-fpm que les organes sains. Il est suggéré que les perturbations du contenu en PAs et en PAs-mm dans les organes malades seraient en relation avec le mauvais déroulement de la floraison. L'accumulation des PAs-fpm dans les organes apparemment sains serait en revanche l'un des processus utilisés par la plante pour faire face à l'agression induite par la présence d'E. lata dans le pied de vigne afin de freiner l'extension de la maladie et d'éviter l'apparition des symptômes. La tyramine conju-guée, en s'accumulant dans les organes apparemment sains et malades, pourrait constituer un marqueur de la présence du champignon dans les pieds de vigne même en l'absence de symptômes. M o t s c l é s : Polyamines, boutons floraux, fleurs, baies, Eutypa lata, Vitis vinifera. Development of polyamine levels in flower buds, flowers and young berries of Vitis vinifera L. (cv. Cabernet Sauvignon) infected by eutypiosis fungus, Eutypa lata Effects of eutypiosis on polyamines (PAs) and tyramine contents in flower buds, flowers and young berries of grapevine were studied to determine the relationship between these components and disease development. The study was performed using organs (1) from healthy vines, (2) healthy appearing organs from vines with one symptomless arm and one diseased arm, and (3) diseased organs. Free PAs, conjugated polyamines (PAs-fpm) and wall-bound polyamines (PAs-mm) were analysed in Vitis vinifera L. cv. Cabernet Sauvignon which is sensitive to eutypiosis. An increase of PAs-mm and a decrease of free PAs were observed in diseased organs; particularly putrescine and diaminopropane were concerned. On the other hand these substances were not affected by eutypiosis in the healthy appearing organs. Alterations of free PAs and PAs-mm in diseased organs were correlated with alteration of the flowering process. In contrast, the PAs-fpm levels decreased in diseased organs in response to eutypiosis whereas they increased in healthy appearing organs. This group of PAs seems to play an important role in the response to Eutypa lata attacks to prevent the ex-pression of symptoms. Accumulation of the conjugated tyramine in diseased and healthy appearing organs as a response to eutypiosis might be used as marker of Eutypa lata infection. K e y w o r d s : Polyamines, flower buds, flower, berries, Eutypa lata, Vitis vinifera.

Effects of eutypiosis on polyamines (PAs) and tyramine contents in flower buds, flowers and young berries of grapevine were studied to determine the relationship between these components and disease development. The study was performed using organs (1) from healthy vines, (2) healthy appearing organs from vines with one symptomless arm and one diseased arm, and (3) diseased organs. Free PAs, conjugated polyamines (PAs-fpm) and wall-bound polyamines (PAs-mm) were analysed in Vitis vinifera L. cv. Cabernet Sauvignon which is sensitive to eutypiosis. An increase of PAs-mm and a decrease of free PAs were observed in diseased organs; particularly putrescine and diaminopropane were concerned. On the other hand these substances were not affected by eutypiosis in the healthy appearing organs. Alterations of free PAs and PAs-mm in diseased organs were correlated with alteration of the flowering process. In contrast, the PAs-fpm levels decreased in diseased organs in response to eutypiosis whereas they increased in healthy appearing organs. This group of PAs seems to play an important role in the response to Eutypa lata attacks to prevent the expression of symptoms. Accumulation of the conjugated tyramine in diseased and healthy appearing organs as a response to eutypiosis might be used as marker of Eutypa lata infection.

Eytypiose of grapevines led to a lowering of the water content and to an accumulation of abscisic acid (ABA) in the leaves of Cabernet Sauvignon known to be susceptible to Eutypiose. The accumulation of ABA in the leaves of infected plants possibly causes changes in the composition of fatty acids, in partiular by reducing their degree of insaturation and by elongating their aliphatic chains. These changes may lower the permeability of membranes and, as a consequence, exchanges with the environment, which possibly intensifies dehydration of infected leaves during their development.

By HPLC analysis, we showed that myceliums of two Eutypa lata isolates cultivated in Enikson and Petersson medium culture synthesized polyamines (Putrescine, Spermidine and Spermine) and abscisic acid (ABA). ln this medium, myceliums of high virulent isolate (BXI-IO) produced fewer polyamines than less virulent isolate (BX 1-5) while inverse phenomenon was observed for ABA content. The two growth regulators were diffused in medium culture in which their levels increase with age of mycelium. In the end, variations of PA and ABA in myceliums show antagonist effect of the two growth regulators in Eutypa lata fungi.