Different approaches to identify genetic traits responsible for susceptibility and resistance to Flavescence dorée in grapevine

Flavescence dorée (FD) is one of the most destructive grapevine yellows (GY) disease that caused huge losses in grape and wine industries. It is causes by phytoplasmas, which are bacteria living in the phloem and transmitted in vineyard by the leafhopper Scaphoideus titanus Ball. Grapevine varieties completely resistant to FD have not been uncovered yet, however inter and intraspecific differences in susceptibility have already been observed, such as poor presence of the disease in vineyard, no severe symptoms and recovery ability of the plant. The different behavior among the grapevine varieties suggests the presence of genetic traits in grapevine germplasm related to high or low susceptibility to FD. The aim of this work is to uncover the low susceptibility mechanisms, and also identify the genetic traits responsible for different phenotypes. The first work presented here, newly accepted, concerns the study of some mechanisms occurring during FD infection in ‘Tocai friulano’ plants, a scarcely susceptible variety. The surveys, performed in a highly FD-infected vineyard, showed a different response against the pathogen in symptomatic ‘Tocai friulano’ plants compared to ‘Pinot gris’ symptomatic ones, a variety highly susceptible. Compartmentalization of FDp in symptomatic portions of ‘Tocai friulano’, its absence in asymptomatic vegetative parts and the absence of the pathogen in permanent tissues (i.e. trunk) were observed. The ability of this cultivar to block and avoid the spread of the symptoms and the pathogen into the whole canopy suggested the possibility of recovery starting events; thus, metabolic analyses and gene expression studies were performed in the phloem tissues to investigate the possible defense mechanisms involved. The second PhD work concerns the study of a population obtained by a cross between ‘Tocai friulano’ and ‘Chardonnay’ (a highly susceptible cultivar) at CREA Viticulture and Enology in Conegliano (Treviso, Italy). Genotyping by sequencing (GBS) analysis was performed on the parentals and on 184 individuals of the progeny, and the resulting single nucleotide polymorphism (SNP) dataset was used both to construct a linkage map and to identify polymorphisms putatively involved in gene 2 regulation. In this work GBS was coupled with a in silico study to identify SNPs modifying transcription factor binding motifs (TFBM) in the proximal regulatory region of genes. Target genes were selected among constitutive differentially expressed genes (DEGs) between ‘Tocai friulano’ and ‘Chardonnay’, previously identified. Parallelly, the observation and the analysis of different FD symptoms in more than 100 infected plants belonging to the progeny led to the identification of valuable phenotypic traits that could be used during phenotype evaluation aimed to QTL identifications. Finally, the existence of differences in FD susceptibility among three ‘Chardonnay’ clones observed in vineyard suggested a further study focused on the transcriptomic and genomic differences among them. The genomes of the three clones were fully sequenced by PacBio HiFi on Sequel II, de novo assembled, and then compared each other to identify structural variants and SNPs. In addition, a transcriptomic study during the early stage of FD infection on two of the three clones showing different phenotype was performed. A dataset of the structural variants distinguishing the less susceptible clone from the other two was defined and it will be used to further investigation on the genes involved in FD infection and susceptibility. During the PhD course this third work was designed and partially performed, however further analyses are required to achieve more complete results.