Molecular bases of interactions in grapevine yellow diseases

The overall goal of the project is to significantly improve our understanding of the molecular interaction between grapevine and phytoplasmas, the causal agents of grapevine yellows diseases (GYs), through the integration of research approaches of the Slovenian and Austrian teams and with support of leading teams in plant physiology and in research of phytoplasmas. We will perform a comprehensive molecular study together with predictive modeling in which we will use natural and model experimental systems and innovative data analysis for identification of the key components of bacterial factors and plant responses to infection. Europe is the worlds main producer and exporter of grapevine planting material and wine. This important economic sector is facing epidemic threats of the GYs bois noir and flavescence doréecaused by taxonomically different types of phytoplasmas, BNp and FDp. To overcome research problems associated with the quarantine status of FDp, we use as a model BNp, which causes the same symptoms on the grapevine as FDp, presumably by very similar mechanisms. In the project we will characterize the current wave of BN disease in Austria and Slovenia by evaluating the infection rate and strain composition in insect vectors, alternative weed hosts and grapevine. Characterization of the strain composition will be evaluated by marker genes and both FDp and BNp strains will be transferred to a model plant for phytoplasma research Catharanthus roseus. To gain knowledge on the overall strain- and phytoplasma effector variability of Austrian and Slovenian BNp we will use next generation sequencing, which will give the basis to study evolution and mechanisms involved in disease development. Expression studies using high throughput RNA sequencing will be used to filter important factors (including effector proteins and regulatory RNAs) responsible for disease development and for differential expression of disease. Also the plant response will be evaluated and the potential contribution of these different factors will be evaluated in the model C. roseus by expressing of potential factors involved in disease development and plant responses. All results will be exploited for constructing an improved model of the phytoplasma/host system. This will look at the phytoplasma-plant interaction both from the plant as well as from the bacterial site which will allow a more comprehensive understanding of the disease. The study will use next generation sequencing and bioinformatics to filter phytoplasma specific information from the larger data set and will confirm the involvement of specific factors for disease development and plant tolerance. Coordinated efforts from Slovenian and Austrian sites will contribute to phytoplasma disease management important in both countries, especially in the wine and fruit sectors.

The stolbur phytoplasma is the cause of "Bois noir" of grapevine and is transmitted through phloem-sucking insects of the order Hemiptera from reservoir plants, namely bindweed and nettle, to grapevine. The disease causes economic damage in grapevine production in Europe. Phytoplasmas cannot be cultured in a conventional manner in the laboratory and therefore the characterization of the pathogen is difficult. Based on marker genes we characterized stolbur genotypes and observed waves of infection originating either from nettle or bindweed as reservoir hosts. The complete genome of six stolbur phytoplasma were obtained by metagenomic methods. Long read sequencing was used to complete the genomes of the two ecotypes of stolbur phytoplasma. The different types of strains show distinct symptoms in experimental plants and contain corresponding different effector genes likely responsible for the phenotype manifestation. For the different ecotypes also comparative expression analyzes were performed, which underline the important factors of the effectors in the interaction between phytoplasma and the plant host. To characterize the effects of phytoplasma effectors and of plant sugar metabolism transient and stable transformation of tobacco and periwinkle was performed. The phenotypes and the altered enzymatic activities of the plants were characterized. In summary, the results can lead to a better understanding of the mechanisms responsible for the symptom manifestations and disease development in phytoplasma - infected plants.