Franco Mannini

Novel functional microRNAs from virus-free and infected Vitis vinifera plants under water stress

MicroRNAs (miRNAs) are small non-coding RNAs that regulate the post-transcriptional control of several pathway intermediates, thus playing pivotal roles in plant growth, development and response to biotic and abiotic stresses. In recent years, the grapevine genome release, small(s)-RNAseq and degradome-RNAseq together has allowed the discovery and characterisation of many miRNA species, thus rendering the discovery of additional miRNAs difficult and uncertain. Taking advantage of the miRNA responsiveness to stresses and the availability of virus-free Vitis vinifera plants and those infected only by a latent virus, we have analysed grapevines subjected to drought in greenhouse conditions. The sRNA-seq and other sequence-specific molecular analyses have allowed us to characterise conserved miRNA expression profiles in association with specific eco-physiological parameters. In addition, we here report 12 novel grapevine-specific miRNA candidates and describe their expression profile. We show that latent viral infection can influence the miRNA profiles of V. vinifera in response to drought. Moreover, study of eco-physiological parameters showed that photosynthetic rate, stomatal conductance and hydraulic resistance to water transport were significantly influenced by drought and viral infection. Although no unequivocal cause–effect explanation could be attributed to each miRNA target, their contribution to the drought response is discussed.

A multidisciplinary study on the effects of phloem-limited viruses on the agronomical performance and berry quality of Vitis vinifera cv. Nebbiolo

Viral infections are known to have a detrimental effect on grapevine yield and performance, but there is still a lack of knowledge about their effect on the quality and safety of end products. Vines of Vitis vinifera cv. Nebbiolo clone 308, affected simultaneously by Grapevine leafroll-associated virus 1 (GLRaV-1), Grapevine virus A (GVA), and Rupestris stem pitting associated virus (RSPaV), were subjected to integrated analyses of agronomical performance, grape berry characteristics, instrumental texture profile, and proteome profiling. The comparison of performance and grape quality of healthy and infected vines cultivated in a commercial vineyard revealed similar shoot fertility, number of clusters, total yield, with significant differences in titratable acidity, and resveratrol content. Also some texture parameters such as cohesiveness and resilience were altered in berries of infected plants. The proteomic analysis of skin and pulp visualized about 400 spots. The ANOVA analysis on 2D gels revealed significant differences among healthy and virus-infected grape berries for 12 pulp spots and 7 skin spots. Virus infection mainly influenced proteins involved in the response to oxidative stress in the berry skin, and proteins involved in cell structure metabolism in the pulp.

Field Performance and Wine Quality Modification in a Clone of Vitis vinifera cv. Dolcetto after GLRaV-3 Elimination

Grapevine leafroll is one of the most widespread virus diseases of grapevine and Grapevine leafroll-associated virus 3 (GLRaV-3) is one of its major causal agents. The effect of GLRaV-3 elimination was studied under vineyard conditions for two years by comparing the virus-free and infected lines of the same clone of Vitis vinifera cv. Dolcetto, an important red wine cultivar grown in northwestern Italy. The results confirmed the advantages of using virus-free vines in terms of field performance (vigor, yield, leaf photosynthesis, and juice soluble solids), whereas the advantages in wine quality were less evident. Sensory analysis clarified that the wine from virus-free vines was slightly different from that of vines infected by GLRaV-3, but did not indicate a significant preference for one or the other.

Modifications in chemical, physical and mechanical properties of Nebbiolo (Vitis vinifera L.) grape berries induced by mixed virus infection.

Modifications in grape quality parameters induced by mixed infection with GFLV and GFkV, GLRaV-1 and GVA, and GLRaV-3 and GVA in three Nebbiolo clones were compared against healthy plants of the same clones in two experimental vineyards in Piemonte, northwest Italy. The aim of the study was to evaluate the effect of virus infection on the mechanical properties of the berry skin and the whole berry as assessed by texture analysis tests, and on the amount and quality of berry skin phenols. Differences were observed in grapevine vigour, yield and juice composition, depending on the viral status of the plants. The anthocyanin profile of the vines infected with GFV and GFkV and those infected with GLRaV-1 and GVA showed a lower percentage of the more stable tri-substituted malvidin-3-glucoside and a higher percentage of cyanidin and peonidin-3-glucosides. Texture analysis showed that the viruses may increase berry-skin thickness and reduce phenol extractability. These effects carry practical implications for wine quality.

Whole-genome sequencing and SNV genotyping of ‘Nebbiolo’ ( Vitis vinifera L.) clones

Abstract‘Nebbiolo’ (Vitis vinifera) is among the most ancient and prestigious wine grape varieties characterised by a wide genetic variability exhibited by a high number of clones (vegetatively propagated lines of selected mother plants). However, limited information is available for this cultivar at the molecular and genomic levels. The whole-genomes of three ‘Nebbiolo’ clones (CVT 71, CVT 185 and CVT 423) were re-sequenced and a de novo transcriptome assembly was produced. Important remarks about the genetic peculiarities of ‘Nebbiolo’ and its intra-varietal variability useful for clonal identification were reported. In particular, several varietal transcripts identified for the first time in ‘Nebbiolo’ were disease resistance genes and single-nucleotide variants (SNVs) identified in ‘Nebbiolo’, but not in other cultivars, were associated with genes involved in the stress response. Ten newly discovered SNVs were successfully employed to identify some periclinal chimeras and to classify 98 ‘Nebbiolo’ clones in seven main genotypes, which resulted to be linked to the geographical origin of accessions. In addition, for the first time it was possible to discriminate some ‘Nebbiolo’ clones from the others.

Grapevine Grafting: Scion Transcript Profiling and Defense-Related Metabolites Induced by Rootstocks

Rootstocks are among the main factors that influence grape development as well as fruit and wine composition. In this work, rootstock/scion interactions were studied using transcriptomic and metabolic approaches on leaves of the “Gaglioppo” variety, grafted onto 13 different rootstocks growing in the same vineyard. The whole leaf transcriptome of “Gaglioppo” grafted onto five selected rootstocks showed high variability in gene expression. In particular, significant modulation of transcripts linked to primary and secondary metabolism was observed. Interestingly, genes and metabolites involved in defense responses (e.g., stilbenes and defense genes) were strongly activated particularly in the GAG-41B combination, characterized in addition by the down-regulation of abscisic acid (ABA) metabolism. On the contrary, the leaves of “Gaglioppo” grafted onto 1103 Paulsen showed an opposite regulations of those transcripts and metabolites, together with the greater sensitivity to downy mildew in a preliminary in vitro assay. This study carried out an extensive transcriptomic analysis of rootstock effects on scion leaves, helping to unravel this complex interaction, and suggesting an interesting correlation among constitutive stilbenes, ABA compound, and disease susceptibility to a fungal pathogen.