Alessandro Spagnolo

Physiological changes in green stems of Vitis vinifera L. cv. Chardonnay in response to esca proper and apoplexy revealed by proteomic and transcriptomic analyses.

Among grapevine trunk diseases, esca proper and apoplexy commonly represent a threat for viticulture worldwide. To retrieve further information about the mechanisms activated in apoplectic and esca proper-affected plants, a two-dimensional gel electrophoresis (2-DE) based analysis was conducted on green stems from 26-year-old standing vines. Symptomatic and asymptomatic stems from both apoplectic (A) and esca proper-affected (E) plants compared to control (without visual symptom since 10 years) stems were studied. Thirty-three differentially expressed proteins were identified by nanoLC-MS/MS and included into three groups conceptually defined as proteins involved in (i) metabolism and energy, (ii) stress tolerance, and (iii) defense response. For nine of them, expression of the relative mRNAs was also monitored by qRT-PCR. Proteome variations were specifically related to apoplexy and esca proper but were more similar in asymptomatic stems than in the symptomatic ones. Remarkable quantitative differences were noted for several proteins in symptomatic stems according to the expressed form, A and E. Results further indicate that similar responses are likely activated in asymptomatic stems but a various quantitative expression is triggered upon onset of apoplexy or esca proper symptoms while both kind of plants are infected by the same pathogenic fungi.

Leaf stripe form of esca induces alteration of photosynthesis and defence reactions in presymptomatic leaves

Esca is a destructive disease in grapevines (Vitis vinifera L.) caused by at least three fungi and characterised by two different external symptoms, the apoplectic and leaf stripe form. This latter form can be discerned as soon as symptoms become visible, but the preceding discrete signs during incubation are poorly or not understood. To further understand the development of the leaf stripe form, the period preceding and following the appearance of symptoms was investigated by studying physiological and molecular markers associated with photosynthetic mechanisms and stress response. No perturbation of any targeted metabolism was observed in asymptomatic leaves of asymptomatic canes from vines showing the leaf stripe form of esca. Conversely, drastic alterations of photosynthesis functions were registered in presymptomatic leaves, as revealed by the decrease of gas exchange and chlorophyll fluorescence, and the repression of photosynthesis-related genes. These alterations were amplified during symptom development. Expression of defence-related genes was affected and detected early in presymptomatic leaves and amplified during symptom expression. Our results suggest that grapevines may react precociously by reducing photosynthesis and triggering defence mechanisms in response to the leaf stripe form of esca.

Flowering as the Most Highly Sensitive Period of Grapevine (Vitis vinifera L. cv Mourvèdre) to the Botryosphaeria Dieback Agents Neofusicoccum parvum and Diplodia seriata Infection

Botryosphaeria dieback is a fungal grapevine trunk disease that currently represents a threat for viticulture worldwide because of the important economical losses due to reduced yield of affected plants and their premature death. Neofusicoccum parvum and Diplodia seriata are among the causal agents. Vine green stems were artificially infected with N. parvum or D. seriata at the onset of three different phenological stages (G stage (separated clusters), flowering and veraison). Highest mean lesion lengths were recorded at flowering. Major proteome changes associated to artificial infections during the three different phenological stages were also reported using two dimensional gel electrophoresis (2D)-based analysis. Twenty (G stage), 15 (flowering) and 13 (veraison) differentially expressed protein spots were subjected to nanoLC-MS/MS and a total of 247, 54 and 25 proteins were respectively identified. At flowering, a weaker response to the infection was likely activated as compared to the other stages, and some defense-related proteins were even down regulated (e.g., superoxide dismutase, major latex-like protein, and pathogenesis related protein 10). Globally, the flowering period seemed to represent the period of highest sensitivity of grapevine to Botryosphaeria dieback agent infection, possibly being related to the high metabolic activity in the inflorescences.

The effects of grapevine trunk diseases (GTDs) on vine physiology

Esca disease as well as Botryosphaeria and Eutypa dieback cause considerable economic problems for vineyards worldwide, and currently, no efficient treatment is available to control these diseases. For these three grapevine trunk diseases (GTDs), the main physiological effects reported concern carbohydrate metabolism and defence responses in the different organs of vine. In the trunk, a depletion of starch reserves in woody tissues is associated with fungal colonization; in the leaves, where pathogens are not present, the carbohydrate metabolism is also affected as revealed by a decline of the photosynthetic rate. A consequence of these disturbances is a lower pool of carbon reserves that might contribute to a decrease of plant development and vigour during the subsequent year. Other metabolic activities such as lipid and amino acid metabolism are down regulated. The perturbation of these primary metabolisms is often associated with the induction of defence responses. The development of biochemical barriers resulting from the accumulation of both tyloses and gummosis is observed during the infection of the wood causing blockage of the xylem vessels and thus limiting the fungal invasion. Their progression in the wood is also inhibited by the formation of polyphenol-rich reaction zones and by the accumulation of pathogenesis-related proteins, and the oxidative burst and the production of reactive oxygen species. Additionally, detoxification processes of the vine are involved; this reaction could be linked to the production of extracellular compounds by GTD agents some of which are phytotoxic. As a consequence, the sensory quality of berries and probably the wine made from these berries decrease. This review presents an overview of the physiological modifications described in vines affected by GTDs.

Changes in plant metabolism and accumulation of fungal metabolites in response to Esca proper and apoplexy expression in the whole grapevine

Trunk diseases have become among the most important grapevine diseases worldwide. They are caused by fungal pathogens that attack the permanent woody structure of the vines and cause various symptoms in woody and annual organs. This study examined modifications of plant responses in green stem, cordon, and trunk of grapevines expressing Esca proper (E) or apoplexy (A) event, which are the most frequent grapevine trunk disease symptoms observed in Europe. Transcript expression of a set of plant defense- and stress-related genes was monitored by quantitative reverse-transcription polymerase chain reaction while plant phytoalexins and fungal metabolites were quantified by high-performance liquid chromatography-mass spectrometry in order to characterize the interaction between the grapevine and trunk disease agents. Expression of genes encoding enzymes of the phenylpropanoid pathway and trans-resveratrol content were altered in the three organs of diseased plants, especially in the young tissues of A plants. Pathogenesis-related proteins and the antioxidant system were severely modulated in A plants, which indicates a drastic stress effect. In the meantime, fungal polyketides 6-MSA, (R)-mellein, and (3R,4R)-4-hydroxymellein, were accumulated in A plants, which suggests their potential effect on plant metabolism during the appearance of foliar symptoms.

Alterations in Grapevine Leaf Metabolism Occur Prior to Esca Apoplexy Appearance

Esca disease is one of the major grapevine trunk diseases in Europe and the etiology is complex, since several inhabiting fungi are identified to be associated with this disease. Among the foliar symptom expressions, the apoplectic form may be distinguished and characterized by sudden dieback of shoots, leaf drop, and shriveling of grape clusters in a few days that can ultimately induce the plant death. To further understand this drastic event, we conducted transcriptomic and metabolomic analyses to characterize responses of leaves during the period preceding symptom appearance (20 and 7 days before foliar symptom expression) and at the day of apoplexy expression. Transcriptomic and metabolomic analyses provide signatures for the apoplectic leaves and most changes concerning the metabolism of carbohydrates, amino acids, and phenylpropanoids. In deciphering glutathione-S-transferase (GST), its preferential location in phloem, correlated with the upregulation of GST genes and a decrease of the glutathione level, offers further support to the putative role of glutathione during apoplexy expression.