www.nature.com/scientificreports OPEN Plasmopara viticola infection afects mineral elements allocation and distribution in Vitis vinifera leaves Stefano Cesco1, Anna Tolotti1, Stefano Nadalini2, Stefano Rizzi2, Fabio Valentinuzzi1, Tanja Mimmo1,3, Carlo Porfdo4, Ignazio Allegretta4, Oscar Giovannini5, Michele Perazzolli5,6, Guido Cipriani2, Roberto Terzano4, Ilaria Pertot5,6 & Youry Pii1* Plasmopara viticola is one of the most important pathogens infecting Vitis vinifera plants. The interactions among P. viticola and both susceptible and resistant grapevine plants have been extensively characterised, at transcriptomic, proteomic and metabolomic levels. However, the involvement of plants ionome in the response against the pathogen has been completely neglected so far. Therefore, this study was aimed at investigating the possible role of leaf ionomic modulation during compatible and incompatible interactions between P. viticola and grapevine plants. In susceptible cultivars, a dramatic redistribution of mineral elements has been observed, thus uncovering a possible role for mineral nutrients in the response against pathogens. On the contrary, the resistant cultivars did not present substantial rearrangement of mineral elements at leaf level, except for manganese (Mn) and iron (Fe). This might demonstrate that, resistant cultivars, albeit expressing the resistance gene, still exploit a pathogen response mechanism based on the local increase in the concentration of microelements, which are involved in the synthesis of secondary metabolites and reactive oxygen species. Moreover, these data also highlight the link between the mineral nutrition and plants’ response to pathogens, further stressing that appropriate fertilization strategies can be fundamental for the expression of response mechanisms against pathogens. Grapevine is one of the most important crops worldwide, for production of fresh fruits, raisins, juices and wine1 and, within the genus Vitis, V. vinifera L. is the most relevant species for the wine industry. In 2018, the total surface cultivated with V. vinifera and the overall wine production was approximatively 7.5 million Ha and 30,000 million litres, respectively (https ://www.oiv.int/). However, grapevine can be afected by a wide number diseases caused by various pathogenic organisms, which can cause production losses both in terms of quan- tity and quality with considerable negative impacts on wine industry 2. Plasmopara viticola is one of the most important phytopathogens of V. vinifera, together with Botrytis cinerea and Erysiphe necator3. Downey mildew symptoms are characterised by initial yellow and oily spots on leaves and other green parts of the plant that evolve in necrosis and death of infected tissues, including bunches, possibly resulting in a total crop loss4. In order to control the disease, several fungicide treatments, including the copper (Cu)-based ones, are commonly applied in vineyards3,5,6. However, there are growing concerns about the possible negative impact of synthetic chemical fungicides and Cu on human health and the environment7–9. In particular, Cu can accumulate in the surface horizon of the soil and have adverse efects on soil biota10–12, as well as, on plants7,13–18. Terefore, with the aim of reducing the impact of synthetic chemical and Cu-based fungicides, the implementation of more sustainable practices aimed at controlling phytopathogen in viticulture are highly encouraged7. 1Faculty of Science and Technology, Free University of Bozen-Bolzano, Piazza Università 5, 39100 Bolzano, Italy. 2Department of Agricultural, Food, Environmental and Animal Sciences, University of Udine, 33100 Udine, Italy. 3Competence Centre for Plant Health, Free University of Bozen-Bolzano, 39100 Bolzano, Italy. 4Department of Soil, Plant and Food Sciences, University of Bari “Aldo Moro”, 70126 Bari, Italy. 5Department of Sustainable Agro-Ecosystems and Bioresources, Research and Innovation Centre, Fondazione Edmund Mach, 38010 San Michele all’Adige, Italy. 6Center Agriculture Food Environment (C3A), University of Trento, 38010 San Michele all’Adige, Italy. *email: [email protected] Scientifc Reports | (2020) 10:18759 | https://doi.org/10.1038/s41598-020-75990-x 1 Vol.:(0123456789) www.nature.com/scientificreports/ Despite the high importance of downy mildew on grapevine production, the molecular bases of P. viticola pathogenesis are still largely unknown2. Nonetheless, several studies have highlighted that, during the early stages of infection (i.e. hyphal structures and haustoria development), P. viticola expresses genes involved in the ATP synthesis and in the active acquisition of mineral elements from the host19,20. On the other hand, grapevine plant responses to P. viticola have been thoroughly characterised at transcriptomic, proteomic and metabolomic levels21–26. In compatible interactions between the host and the pathogen, grapevine plants up-regulate genes related to the secondary metabolism, defence and response to external stimuli, whilst genes involved in the photosynthesis and the carbon metabolism are mostly down-regulated25. Transcriptional modulations were detected in micro-dissected stomata (i.e. the primary site of P. viticola infection) and surroundings cells, suggest- ing that grapevine plants can react to the pathogen by eliciting both a site-specifc response and a short distance signal(s) from the stomata to neighbouring cells27. In contrast, incompatible interactions are characterised by the up-regulation of stress and defence-related genes, as for instance the Pathogenesis Related (PR) proteins3,28. Several Quantitative Trait Loci (QTLs), named Resistance to P. viticola (Rpv) genes, have been identifed in diferent grapevine species and they have been associated with major phenotypic traits of resistance to downy mildew disease. In particular, Rpv1 and Rpv2 were found to be responsible for the resistance in Muscadinia rotun- difolia29, Rpv3 was associated with the resistance in ‘Villard blanc’30, whilst the resistance traits in V. amurensis was ascribed to Rpv8, Rpv10 and Rpv1231–33. Since the discovery of the resistance traits to P. viticola, breeding programs have been undertaken in order to introduce the features related to downy mildew resistance in sus- ceptible species34–38. Tus, the use of grapevine genotypes resistant to P. viticola infection might represent an important tool to improve the sustainability of viticulture39, in particular considering the current restriction on the use of Cu in the European Union40. Depending on the specifc QTL and the grapevine genotype, the resistance mechanisms elicited by the Rpv genes can involve diferent responses, as for instance the accumulation of callose and lignin41–43, the hypersensitive response33,44, synthesis of phytoalexins45,46, the accumulation of phenolics in the infected tissues47,48, the induction of either cell necrosis38,44,49 or peroxidase activity50,51. Recent studies have highlighted that also the ionome profle of plants might represent an important factor determining the success of the infection process by a pathogenic organism 52,53. Mineral elements, both macro- and microelements, play a fundamental role in plants, being essential for the life cycles completion. Tey are involved in a plethora of cell functions, including primary and secondary metabolisms, energy production, defence, signals transduction, genes regulation, hormones perception, reproduction, enzyme functioning, as well as in maintaining cell structure 54. In addition, an adequate intracellular concentration of essential metal ions was recognised as a necessary prerequisite for both pathogen virulence and plant defence responses55,56. In particular, considering the role played by metal ions in living organisms, it is conceivable that intracellular mineral concentrations of essential mineral elements can have a strong impact on a wide range of pathogens and on their ability to establish an interaction with host plants 57. A recent research carried out on olive trees infected with the bacterial pathogen Xylella fastidiosa demonstrated that the resistant variety “Leccino”, which usually shows low or no symptoms, was characterised by a diferent ionomic signature as compared to the symptomatic susceptible variety (e.g. “Ogliarola salentina”)53. In particular, increases in calcium (Ca) and manganese (Mn) leaf concentration were detected, suggesting that these mineral elements might be related to the enhanced resistance of the “Leccino” variety against X. fastidiosa53. Similarly, it was reported that the ionomic balance in Lactuca sativa L. infected with the bacterium Xanthomonas campestris pv. vitians correlated with the degree of resistance showed by diferent lettuce varieties52. Starting from this knowledge background, we investigated whether the resistance mechanisms showed by resistant grapevine cultivars, beside the specifc activities of Rpv genes, might be also associated to the modula- tion of the nutrient balance in the plants. Te ionomic signature of a plant tissue is the result of the regulation of transporter-based fuxes of mineral elements within the plant and among the diferent tissues, playing the leaf a relevant role 58. Te present study was undertaken on two Plasmopara-sensitive grapevine cultivars, Cabernet Sauvignon and Sauvignon Blanc, widely cultivated in wine-producing Countries (approx. 340 thousand and 120 thousand Ha, respectively) (https ://www.oiv.int/publi c/media s/5888/en-distr ibuti on-of-the-world s-grape vine-varie