Lucio Brancadoro

The Influence of Genotype and Environment on Small RNA Profiles in Grapevine Berry

Understanding the molecular mechanisms involved in the interaction between the genetic composition and the environment is crucial for modern viticulture. We approached this issue by focusing on the small RNA transcriptome in grapevine berries of the two varieties Cabernet Sauvignon and Sangiovese, growing in adjacent vineyards in three different environments. Four different developmental stages were studied and a total of 48 libraries of small RNAs were produced and sequenced. Using a proximity-based pipeline, we determined the general landscape of small RNAs accumulation in grapevine berries. We also investigated the presence of known and novel miRNAs and analyzed their accumulation profile. The results showed that the distribution of small RNA-producing loci is variable between the two cultivars, and that the level of variation depends on the vineyard. Differently, the profile of miRNA accumulation mainly depends on the developmental stage. The vineyard in Riccione maximizes the differences between the varieties, promoting the production of more than 1000 specific small RNA loci and modulating their expression depending on the cultivar and the maturation stage. In total, 89 known vvi-miRNAs and 33 novel vvi-miRNA candidates were identified in our samples, many of them showing the accumulation profile modulated by at least one of the factors studied. The in silico prediction of miRNA targets suggests their involvement in berry development and in secondary metabolites accumulation such as anthocyanins and polyphenols.

High-throughput 18K SNP array to assess genetic variability of the main grapevine cultivars from Sicily

The viticulture of Sicily, for its vocation, is one of the most important and ancient forms in Italy. Autochthonous grapevine cultivars, many of which known throughout the world, have always been cultivated in the island from many centuries. With the aim to preserve this large grapevine diversity, previous studies have already started to assess the genetic variability among the Sicilian cultivars by using morphological and microsatellite markers. In this study, simple sequence repeat (SSR) were utilized to verify the true-to-typeness of a large clone collection (101) belonging to 21 biotypes of the most 10 cultivated Sicilian cultivars. Afterwards, 42 Organization Internationale de la Vigne et du Vin (OIV) descriptors and a high-throughput single nucleotide polymorphism (SNP) genotyping array (Vitis18kSNP) were applied to assess genetic variability among cultivars and biotypes of the same cultivar. Ampelographic traits and high-throughput SNP genotyping platforms provided an accuracy estimation of genetic diversity in the Sicilian germplasm, showing the relationships among cultivars by cluster and multivariate analyses. The large SNP panel defined sub-clusters unable to discern among biotypes, previously classified by ampelographic analysis, belonging to each cultivar. These results suggested that a very large number of SNP did not cover the genome regions harboring few morphological traits. Genetic structure of the collection revealed a clear optimum number of groups for K = 3, clustering in the same group a significant portion of family-related genotypes. Parentage analysis highlighted significant relationships among Sicilian grape cultivars and Sangiovese, as already reported, but also the first evidences of the relationships between Nero d’Avola and both Inzolia and Catarratto. Finally, a small panel of highly informative markers (12 SNPs) allowed us to isolate a private profile for each Sicilian cultivar, providing a new tool for cultivar identification.

Stem Xylem Characterization for Vitis Drought Tolerance

Together with stomatal conductance and root conductivity, the stem water reserve and transport systems could be regulatory mechanisms able to participate in the regulation of the plant water status. Lianas, such as Vitis spp., minimize the trunk support role, and stems have evolved to improve their ability in water transport. In this work, stems of 10 different Vitis species were studied in relation to their expected drought tolerance using reflectance spectroscopy. Spectra were measured before (T0) and after coloration with Sudan IV dye. The T0 spectral signature showed characteristic species features. The partial least squares (PLS) regression and the self-organizing map (SOM) neural network analysis were able to predict the expected drought tolerance score; thus, reflectance spectroscopy was demonstrated to be a useful technique for drought tolerance phenotyping. These methods could be applied for the preliminary selection of new rootstocks/cultivars. Wood composition variation appeared to be correlated with the water stress susceptibility. To clarify this relationship, the attention was focused on the wood hydrophobicity. Sudan IV is a microscopy dye traditionally used to underline suberin, waxes, and, in general, hydrophobic substances. Differences between rough and colored spectra evidenced the absorption band of Sudan IV with a maximum at 539 nm. The coloration intensity was used to develop a hydrophobicity index. The obtained values were correlated with the expected drought tolerance score. Therefore, hydrophobic compounds seem to play an important role in water use efficiency, and an hydrophobic barrier in the xylem tissue appears to be a protective mechanism against water stress.

Effects of nutrient spray applications on malic and tartaric acid levels in grapevine berry

Abstract The levels of tartaric and malic acids in grape berries sprayed with either CaCl2, KCl, KNO3, NH4Cl, or Ca(NO3)2 in order to increase potassium (K), calcium (Ca), ammonium (NH4), or nitrate (NO3) availability during the fruit set‐veraison period did not show a direct effect on the synthesis of malic and tartaric acids in the berries during the pre‐veraison period; however at veraison, a relationship was established between K and malic acid, a relationship that did not last until the ripening of the berry.

Water deficit effects on grapevine woody tissue pigmentations

Grossi D., Rustioni L., Simone Di Lorenzo G., Failla O., Brancadoro L. (2016): Water deficit effects on grapevine woody tissue pigmentations. Hort. Sci. (Prague): 43: 188–194. Water availability is an important environmental factor in viticulture. In a climate change context, vineyard management should be adapted to the new conditions. Drought-resistant rootstocks need to be selected. In this paper, reflectance spectroscopy is proposed as a new method to characterize the water stress effects on woody section pigmentations. Cabernet Sauvignon grafted on 4 different rootstocks (140Ru, 420A, M2 and M3) represented the plant material. Greenhouse controlled conditions allowed the comparison of well-watered (WW) and water-stressed (WS) plants. The physiological responses were characterized concerning daily water consumption, stem water potential, gas exchange, and plant growth. The water use efficiency was calculated and discussed as well. Spectroscopy analyses of woody sections indicated a major absorption band probably related to phenolic derivatives. Water stress produced characteristic spectrum modifications both in the Cabernet Sauvignon stem and in the rootstock xylem. These preliminary results encourage further studies addressed at the evaluation of drought-resistant genotypes, to distinguish their stress responses and to characterize the compositional aspects linked to drought tolerance.

Study of intra-varietal diversity in biotypes of Aglianico and Muscat of Alexandria (Vitis vinifera L.) cultivars

Background and Aims Somatic mutations are the major source of intra-varietal variability in some species such as grapevine (Vitis vinifera L.) that are propagated vegetatively. Aglianico and Muscat of Alexandria are two cultivars mostly cultivated in the south of Italy, showing great intra-varietal diversity. The variability of 15 clones of Aglianico belonging to three biotypes, Taburno, Taurasi and Vulture, and 21 clones of Muscat of Alexandria, coming from France, Greece, Italy and Spain, was investigated. Methods and Results The intra-varietal diversity was investigated by ampelographic and eno-carpological (berry characteristics of oenological interest) methods (nine OIV descriptors, related to bunch and berry morphology, sugar content, total acid content and pH of must), chemical (anthocyanins, aromatic compounds and phenolic substances) and genetic analysis [18 K SNPs, single nucleotide polymorphisms (SNP)]. Intra-varietal diversity has been detected in both cultivars, mainly based on differences in the accumulation of secondary metabolites. The Taurasi biotype was discriminated by linear discrimination analysis mainly for the proportion of malvidin-3-O-glucoside acetate and methoxylated anthocyanins, while the Spanish Muscat biotype was discriminated based on the proportion of flavanols and hydroxycinnamic acids. Conclusions Ampelography and the concentration of secondary metabolites have proved to be the best tools for the characterisation of the intra-varietal variability of grapevine germplasm. The high-throughput genetic analysis based on 18 K SNP loci did not highlight a polymorphism sufficient to distinguish among biotypes of the same cultivar, suggesting that a different molecular approach has to be adopted. Significance of the Study This study confirms that intra-varietal diversity is better examined with a multidisciplinary approach.

Grapevine field experiments reveal the contribution of genotype, the influence of environment and the effect of their interaction (GxE) on berry transcriptome

Changes in the performance of genotypes in different environments are defined as genotype × environment (G×E) interactions. In grapevine (Vitis vinifera), complex interactions between different genotypes and climate, soil and farming practices yield unique berry qualities. However, the molecular basis of this phenomenon remains unclear. To dissect the basis of grapevine G×E interactions we characterized berry transcriptome plasticity, the genome methylation landscape and within-genotype allelic diversity in two genotypes cultivated in three different environments over two vintages. We identified, through a novel data-mining pipeline, genes with expression profiles that were: unaffected by genotype or environment, genotype-dependent but unaffected by the environment, environmentally-dependent regardless of genotype, and G×E-related. The G×E-related genes showed different degrees of within-cultivar allelic diversity in the two genotypes and were enriched for stress responses, signal transduction and secondary metabolism categories. Our study unraveled the mutual relationships between genotypic and environmental variables during G×E interaction in a woody perennial species, providing a reference model to explore how cultivated fruit crops respond to diverse environments. Also, the pivotal role of vineyard location in determining the performance of different varieties, by enhancing berry quality traits, was unraveled.

Characterization of iron deficiency symptoms in grapevine (Vitis spp.) leaves by reflectance spectroscopy

The work aims at the description of the iron deficiency symptoms in grapevine leaves by reflectance spectroscopy at the plant and leaf levels. 5 genotypes of Vitis spp. were selected and grown in hydroponic conditions with and without iron supply. 450 spectra were collected among basal, young and apical leaves, as well as veins and interveinal areas. Iron deficiency produced significant and characteristic modifications in the pigment accumulation, proportion and distribution in plants. Basal leaves resulted to have higher concentrations of photosynthetic pigments in stressed plants with respect to the control, probably due to compensation effects. Iron deficient plants had lower chlorophyll concentrations in young and apical leaves. In the apical zone, also the relative composition of pigments appeared to be modified, explaining the reddish-yellowish apex appearance of iron deficient vines. Finally, the pigment distribution along the shoot characterized the symptoms, as well as the spectral variations among veins and interveinal areas. These results could support future applications in vineyard management (e.g.: symptom identification and detection; precision fertilization) as well as breeding programs for new rootstock selections (e.g.: fast screenings of seedlings).

Evidence for a Sympatric Origin of Ribolla gialla, Gouais Blanc and Schiava cultivars (V. vinifera L.)

Ribolla gialla is an autochthonous grape variety cultivated in Friuli Venezia Giulia (Italy) and in Slovenia, and probably originated in Eastern Central Europe. Just like Ribolla gialla, the Gouais blanc and Schiava cultivars also appear to have originated in the same geographical area. To verify this hypothesis, a pool of varieties probably sharing the same historical and geographical origin were taken into account. The fingerprinting, by 35 SSR loci, of Gouais blanc, Ribolla gialla, Schiava grossa and Schiava lombarda (synonyms of Schiava bresciana), is presented and the correlation among historical, geographical and genetic information of these cultivars was investigated. Gouais blanc and Traminer, already suggested as key varieties in the development of European grape diversity and as parents for some French varieties, interestingly enough seems to be linked to Ribolla gialla. The putative parentage was verified using 58 microsatellite markers. The genetic results suggest a common geographical origin for Gouais blanc, Ribolla gialla and the Schiava group: these cultivars appear to be related through sympatric origin. The hypothesis of Ribolla gialla as a progeny of Gouais blanc and Traminer was ruled out. The data proved a second-degree relationship between Gouais blanc and Ribolla gialla and a third or more distant degree of relationships between Ribolla gialla and Traminer.

Estimating Leaf Area Index (LAI) in Vineyards Using the PocketLAI Smart-App

Estimating leaf area index (LAI) of Vitis vinifera using indirect methods involves some critical issues, related to its discontinuous and non-homogeneous canopy. This study evaluates the smart app PocketLAI and hemispherical photography in vineyards against destructive LAI measurements. Data were collected during six surveys in an experimental site characterized by a high level of heterogeneity among plants, allowing us to explore a wide range of LAI values. During the last survey, the possibility to combine remote sensing data and in-situ PocketLAI estimates (smart scouting) was evaluated. Results showed a good agreement between PocketLAI data and direct measurements, especially for LAI ranging from 0.13 to 1.41 (R2 = 0.94, RRMSE = 17.27%), whereas the accuracy decreased when an outlying value (vineyard LAI = 2.84) was included (R2 = 0.77, RRMSE = 43.00%), due to the saturation effect in case of very dense canopies arising from lack of green pruning. The hemispherical photography showed very high values of R2, even in presence of the outlying value (R2 = 0.94), although it showed a marked and quite constant overestimation error (RRMSE = 99.46%), suggesting the need to introduce a correction factor specific for vineyards. During the smart scouting, PocketLAI showed its reliability to monitor the spatial-temporal variability of vine vigor in cordon-trained systems, and showed a potential for a wide range of applications, also in combination with remote sensing.