The Constitutive Endopolygalacturonase TvPG2 Regulates the Induction of Plant Systemic Resistance by Trichoderma virens.

Trichoderma spp. are opportunistic fungi some of which are commonly present in the rhizosphere. Several species, such as T. virens, are also efficient biocontrol agents against phytopathogenic fungi and exert beneficial effects on plants. These effects are the consequence of interactions between Trichoderma and plant roots, which trigger enhanced plant growth and induce plant resistance. We have previously shown that T. virens I10 expresses two endopolygalacturonase genes, tvpg1 and tvpg2, during the interaction with plant roots; tvpg1 is inducible while tvpg2 is constitutively transcribed. Using the same system, the tomato polygalacturonase-inhibitor gene Lepgip1 was induced at the same time as tvpg1. Here we show by gene disruption that TvPG2 performs a regulatory role on the inducible tvpg1 gene and in triggering the plant immune response. A tvpg2-knockout strain fails to transcribe the inducible tvpg1 gene in neither in vitro in inducing media containing pectin or plant cell walls, nor during the in vivo interaction with tomato roots. Likewise, the in vivo induction of Lepgip1 does not occur, and its defense against the pathogen Botrytis cinerea is significantly reduced. Our data prove the importance of a T. virens constitutively produced endopolygalacturonase in eliciting plant induced systemic resistance against pathogenic fungi.

Study of the terpene profile at harvest and during berry development of Vitis vinifera L. aromatic varieties Aleatico, Brachetto, Malvasia di Candia aromatica and Moscato bianco.

BACKGROUND: In this work, we evaluated the aromatic composition of berries at harvest and during the whole berry development of Aleatico, Brachetto, Malvasia di Candia aromatica and Moscato bianco grapevine varieties, which were cultivated in the same vineyard and under the same agricultural conditions. RESULTS: Malvasia had a total concentration of aroma which was significantly higher than the other grapes that showed comparable amounts with each other. The class of monoterpenes was quantitatively predominant for all four grapes, in the free and in the most plentiful bound form. In Malvasia and Aleatico there was a high prevalence of geraniol derivatives throughout the entire berry development while in Brachetto and in Moscato there was a prevalence of linalool derivatives during the green phase and geraniol derivatives during ripening. CONCLUSION: The study of the monoterpene profile allowed us to highlight similarities and differences among the four aromatic varieties and to present a hypothesis about their biosynthetic dynamics. This information is useful for further studies on gene functional characterisation and the regulation of these important berry pathways. © 2016 Society of Chemical Industry.

Influence of vineyard altitude on Glera grape ripening (Vitis vinifera L.): effects on aroma evolution and wine sensory profile.

BACKGROUND: Environmental factors have been acknowledged to greatly influence grape and wine aromas. Among them, the effect of altitude on grape aroma compounds has scarcely been debated in literature available to date. In the present study, we investigated the influence of altitude on grape composition and aroma evolution during ripening of Vitis vinifera L. cultivar Glera grown in Conegliano-Valdobbiadene DOCG area (Italy). RESULTS: The site at highest altitude (380 m above sea level) was warmer than the lowest site (200 m above sea level) and, even with differences in temperature in the range 1.5-2 °C, the impact of the cultivation site on grape ripening and aroma accumulation and preservation was significant. The lowest site demonstrated slower grape ripening, and grapes at harvest accumulated lower amounts of all of the main classes of aroma compounds typical of the Glera variety. Wines produced from the highest site were preferred in tasting trials for their more patent floral notes and elegance. CONCLUSION: Altitude strongly influences grape ripening evolution and flavour accumulation in the Glera grape, and this result accounts for the different styles in the sparkling wines subsequently produced. Moreover, the present study shows that aroma compound biosynthesis, particularly that of benzenoides, starts before véraison in Glera. © 2016 Society of Chemical Industry.

Distinct transcriptome responses to water limitation in isohydric and anisohydric grapevine cultivars.

BACKGROUND: Grapevine (Vitis vinifera L.) is an economically important crop with a wide geographical distribution, reflecting its ability to grow successfully in a range of climates. However, many vineyards are located in regions with seasonal drought, and these are often predicted to be global climate change hotspots. Climate change affects the entire physiology of grapevine, with strong effects on yield, wine quality and typicity, making it difficult to produce berries of optimal enological quality and consistent stability over the forthcoming decades. RESULTS: Here we investigated the reactions of two grapevine cultivars to water stress, the isohydric variety Montepulciano and the anisohydric variety Sangiovese, by examining physiological and molecular perturbations in the leaf and berry. A multidisciplinary approach was used to characterize the distinct stomatal behavior of the two cultivars and its impact on leaf and berry gene expression. Positive associations were found among the photosynthetic, physiological and transcriptional modifications, and candidate genes encoding master regulators of the water stress response were identified using an integrated approach based on the analysis of topological co-expression network properties. In particular, the genome-wide transcriptional study indicated that the isohydric behavior relies upon the following responses: i) faster transcriptome response after stress imposition; ii) faster abscisic acid-related gene modulation; iii) more rapid expression of heat shock protein (HSP) genes and iv) reversion of gene-expression profile at rewatering. Conversely, that reactive oxygen species (ROS)-scavenging enzymes, molecular chaperones and abiotic stress-related genes were induced earlier and more strongly in the anisohydric cultivar. CONCLUSIONS: Overall, the present work found original evidence of a molecular basis for the proposed classification between isohydric and anisohydric grapevine genotypes.

Analysis of subgroup C of fungal chitinases containing chitin-binding and LysM modules in the mycoparasite Trichoderma atroviride.

Fungi have a plethora of chitinases, which can be phylogenetically divided into three subgroups (A, B and C). Subgroup C (sgC) chitinases are especially interesting due to their multiple carbohydrate-binding modules, but they have not been investigated in detail yet. In this study, we analyzed sgC chitinases in the mycoparasites Trichoderma atroviride and Trichoderma virens. The expression of sgC chitinase genes in T. atroviride was induced during mycoparasitism of the fungal prey Botrytis cinerea, but not Rhizoctonia solani and correspondingly only by fungal cell walls of the former. Interestingly, only few sgC chitinase genes were inducible by chitin, suggesting that non-chitinous cell wall components can act as inducers. In contrast, the transcriptional profile of the most abundantly expressed sgC chitinase gene tac6 indicated a role of the protein in hyphal network formation. This shows that sgC chitinases have diverse functions and are not only involved in the mycoparasitic attack. However, sequence analysis and 3D modelling revealed that TAC6 and also its ortholog in T. virens have potentially detrimental deletions in the substrate-binding site and are thus probably not catalytically active enzymes. Genomic analysis showed that the genes neighboring sgC chitinases often encode proteins that are solely composed of multiple LysM modules, which were induced by similar stimuli as their neighboring sgC chitinase genes. This study provides first insights into fungal sgC chitinases and their associated LysM proteins.

Effect of methyl jasmonate on the aroma of Sangiovese grapes and wines.

Methyl jasmonate (MeJA) was applied in a vineyard on leaves and grape clusters of cv Sangiovese to test its ability to stimulate the production of aromas and identify the main genes involved in the biosynthetic pathways switched on by the elicitor. MeJA application led to a delay in grape technological maturity and a significant increase in the concentration of several berry aroma classes (about twice the total aroma: from around 3 to 6µg/g of berry). Of these, monoterpenes showed the most significant increase. An analysis of the expression of terpenoid biosynthesis genes confirmed that the MeJA application activated the related biosynthetic pathway. The expression of all the TPS genes analyzedwas higher in samples treated with MeJA. Also the wines produced by microvinification of Sangiovese treated and untreated grapes showed a rise in the aroma concentration as in berries, with an important impact on longevity and sensorial characters of wines.

Physiological parameters and protective energy dissipation mechanisms expressed in the leaves of two Vitis vinifera L. genotypes under multiple summer stresses.

Photosynthetic performances and energy dissipation mechanisms were evaluated on the anisohydric cv. Sangiovese and on the isohydric cv. Montepulciano (Vitis vinifera L.) under conditions of multiple summer stresses. Potted vines of both cultivars were maintained at 90% and 40% of maximum water availability from fruit-set to veraison. One week before veraison, at predawn and midday, main gas-exchange and chlorophyll fluorescence parameters, chlorophyll content, xanthophyll pool and cycle and catalase activity were evaluated. Under water deficit and elevated irradiance and temperature, contrary to cv. Montepulciano and despite a significant leaf water potential decrease, Sangiovese's leaves kept their stomata more open and continued to assimilate CO2 while also showing higher water use efficiency. Under these environmental conditions, in comparison with the isohydric cv. Montepulciano, the protective mechanisms of energy dissipation exerted by the anisohydric cv. Sangiovese were: (i) higher stomatal conductance and thermoregulation linked to higher transpiration rate; (ii) greater ability at dissipating more efficiently the excess energy via the xanthophylls cycle activity (thermal dissipation) due to higher VAZ pool and greater increase of de-epoxidation activity.

Expression of terpene synthase genes associated with the formation of volatiles in different organs of Vitis vinifera.

Plants produce a plethora of volatile organic compounds (VOCs) which are important in determining the quality and nutraceutical properties of horticultural food products, including the taste and aroma of wine. Given that some of the most prevalent grape aroma constituents are terpenoids, we investigated the possible variations in the relative expression of terpene synthase (TPS) genes that depend on the organ. We thus analysed mature leaves, young leaves, stems, young stems, roots, rachis, tendrils, peduncles, bud flowers, flowers and berries of cv Moscato bianco in terms of their VOC content and the expression of 23 TPS genes. In terms of the volatile characterization of the organs by SPME/GC-MS analysis, flower buds and open flowers appeared to be clearly distinct from all the other organs analysed in terms of their high VOC concentration. Qualitatively detected VOCs clearly separated all the vegetative organs from flowers and berries, then the roots and rachis from other vegetative organs and flowers from berries, which confirms the specialization in volatile production among different organs. Our real-time RT-PCR results revealed that the majority of TPS genes analysed exhibited detectable transcripts in all the organs investigated, while only some were found to be expressed specifically in one or just a few organs. In most cases, we found that the known products of the in vitro assay of VvTPS enzymes corresponded well to the terpenes found in the organs in which the encoding gene was expressed, as in the case of (E)-ß-caryophyllene synthases, α-terpineol synthase and α-farnesene synthase. In addition, we found groups of homologous TPS genes, such as (E)-ß-caryophyllene and ß-ocimene synthases, expressed distinctively in the various tissues. This thus confirmed the subfunctionalization events and a specialization on the basis of the organs in which they are mostly expressed.

Analysis of the expression of terpene synthase genes in relation to aroma content in two aromatic Vitis vinifera varieties.

Grape (Vitis vinifera L.) flavour management in the vineyard requires knowledge of the derivation of individual flavour and aroma characteristics. Some of the most prevalent wine grape aroma constituents are terpenoids and this study represents a wide report about grape terpene synthase (TPS) gene transcript profiling in different tissues of two aromatic grapevine varieties, particularly flowers and developing berries, correlated with the accumulation patterns of free aroma compounds. All investigated genes belonging to the TPS-a and TPS-b subfamilies reached the highest expression in accordance with the peak of accumulation of the respective compounds. In the TPS-g subfamily, only one of the genes characterised for linalool synthases showed major transcript abundance in ripening berries, whereas the only geraniol synthase had a peak of expression in green berries and at the beginning of ripening, when geraniol concentration started to increase and overcome the linalool concentration. The genes identified in this study as being mainly responsible for linalool and geraniol synthesis during berry development, and the phenological phases in which they are mostly expressed, should be of interest to viticulturists and wine makers to improve decision making along the chain of production.