Grapevine (Vitis vinifera L.), a globally significant crop, is highly susceptible to Botrytis cinerea, the causative agent of gray mold disease. This study investigates transcriptomic responses to B. cinerea in tolerant and susceptible grapevine genotypes using RNA sequencing (RNA-seq). Differentially expressed genes (DEGs) were identified at three time points (T1, T2, T3), highlighting both genotype-independent and genotype-specific responses. Early-stage infection (T1) revealed rapid and robust activation of defense pathways in both genotypes, though the tolerant genotype showed enhanced modulation of metabolic processes by T2, prioritizing secondary metabolism and stress adaptation over growth. In contrast, the susceptible genotype exhibited less coordinated metabolic reprogramming, with delayed or weaker activation of key defense mechanisms. Gene Ontology and KEGG analyses identified critical pathways, including phenylpropanoid biosynthesis-like lignin metabolism, MAPK signaling, as well as candidate genes such as WRKY transcription factors and enzymes involved in cell wall fortification and antifungal compound biosynthesis. Genotype-specific responses emphasized metabolic flexibility as a determinant of resistance, with the tolerant genotype exhibiting superior resource allocation to defense pathways. These findings provide insights into the molecular basis of grapevine resistance to B. cinerea, offering potential targets for breeding or genetic engineering to enhance resilience and reduce fungicide dependency.

Extracting high-quality RNA from grape (Vitis Vinifera) berry skins is challenging due to their high levels of polysaccharides, phenolic compounds, sugars, and organic acids, which can negatively impact RNA purity and yield. Indeed, polyphenols can bind to RNA, polysaccharides may co-precipitate, and sugars and organic acids can interfere with the pH and ionic properties of the extraction buffer. Commercial kits offer a quick extraction method but are often ineffective for grape berry skins. Similarly, protocols that work well for other vegetal tissues are also inefficient and time-consuming for this tissue. To overcome these limitations, we optimized the RNA isolation by adding a sorbitol pre-wash step to both a non-commercial protocol and a commercial kit. Our results show that it significantly improves the RNA yield and quality from grape berry skins, increasing the RNA purity and integrity, as evidenced by higher RIN (RNA Integrity Number) values and better Qubit and Nanodrop measurements. The strategy’s efficacy was further validated through RNA sequencing, yielding high-quality reads with low error rates, suitable for gene expression studies. Thus, incorporating a sorbitol pre-wash step improves the RNA yield and quality from grape berry skins making it suitable for high-throughput sequencing, and provides a reliable tool for advancing grapevine research.

It is known that plant-associated microorganisms must be considered as key drivers for plant health, productivity, community composition, and ecosystem functioning. Much attention has been paid to the exploration of the plant-associated microbiome in search of new microorganisms with antagonistic activity against various phytopathogens. P. viticola represents one of the main fungal agents of grape yield losses during the pre-harvest stage. To date, the use of chemical pesticides is the main method to control downy mildew infection in vineyards. In recent years, innovative and more environmentally suitable control strategies, such as Biological Control Agents (BCAs), have been greatly improved. In this study, 16 new bacterial strains, isolated from the leaves of three table grape cultivars, were tested for their in vivo effectiveness against downy mildew of grape, resulting in one B. velenzensis (‘BLG_B4), one P. pleuroti (‘BLG_B5), one P. psychrotolerans (‘BLG_B1.3’) and one B. subtilis (‘BLG_B1.1.1’) bacterial strains that were marked as good BCAs. As possible mechanisms of action, a genetic analysis was performed to evaluate the ability of selected bacterial strains to reduce the expression of two P. viticola effectors. Finally, their effectiveness against a wide range of fungal pathogens and their safety for human health was also evaluated.