Dissecting the susceptibility/resistance mechanism of Vitis vinifera for the future control of downy mildew

23CO.NA.VI. 2020 – 8° Convegno Nazionale di Viticoltura, Udine, Italy, July 5-7, 2021openInternationalBothThe Eurasian grapevine (Vitis vinifera), a species cultivated worldwide for high-quality wine production, is extremely susceptible to the agent of downy mildew, Plasmopara viticola. Nevertheless, germplasm from Georgia (Southern Caucasus, the first grapevine domestication centre), characterized by a high genetic variability, showed resistance traits to P. viticola. The cultivar Mgaloblishvili exhibited the most promising phenotype in terms of resistance against P. viticola. Its defence response results in: i) low disease intensity; ii) low sporulation; iii) damaged mycelium; iv) production of antimicrobial compounds such as volatile organic compounds (VOCs), whose effectiveness on the pathogen was evaluated by leafdisc assays. At the transcriptomic level, its resistance mechanism is determined by the differential expression of both resistance and susceptible genes. The resistance genes are related to: i) pathogen recognition through PAMP, DAMP and effector receptors; ii) ethylene signalling pathway; iii) synthesis of antimicrobial compounds (VOCs) and fungal wall degrading enzymes; iv) development of structural barriers (cell wall reinforcement). The first putative susceptible gene was the transcription factor VviLBDIf7 gene, whose validation was carried out by dsRNA (double-stranded RNA) assay. In this work, these unique results on plant-pathogen interaction are reviewed with the aim of developing new strategies to control the disease.openRicciardi, Valentina; Marcianò, Demetrio; Sargolzaei, Maryam; Marrone Fassolo, Elena; Fracassetti, Daniela; Brilli, Matteo; Moser, Mirko; Vahid, Shariati J.; Tavakole, Elahe; Maddalena, Giuliana; Passera, Alessandro; Casati, Paola; Pindo, Massimo; Cestaro, Alessandro; Costa, Alex; Bonza, Maria Cristina; Maghradze, David; Tirelli, Antonio; Failla, Osvaldo; Bianco, Piero Attilio; Quaglino, Fabio; Toffolatti, Silvia Laura; De Lorenzis, GabriellaRicciardi, V.; Marcianò, D.; Sargolzaei, M.; Marrone Fassolo, E.; Fracassetti, D.; Brilli, M.; Moser, M.; Vahid, S.J.; Tavakole, E.; Maddalena, G.; Passera, A.; Casati, P.; Pindo, M.; Cestaro, A.; Costa, A.; Bonza, M.C.; Maghradze, D.; Tirelli, A.; Failla, O.; Bianco, P.A.; Quaglino, F.; Toffolatti, S.L.; De Lorenzis, G

Methods for Fungicide Efficacy Screenings: Multiwell Testing Procedures for the Oomycetes <i>Phytophthora infestans</i> and <i>Pythium ultimum</i>

Oomycetes-borne diseases represent a serious problem for agriculture sustainability due to the high use of chemical products employed for their control. In recent years, increasing concerns on side effects associated with fungicide utilization have led to the reduction of the permissible modes of action, with the remaining ones continuously threatened by the increase of resistant strains in the pathogen populations. In this context, it is mandatory to develop new generation fungicides characterized by high specificity towards the target species and low environmental impact to guarantee the sustainability, productivity, and quality of food production. Fungicide discovery is a lengthy and costly process, and despite these urgent needs, poor description and formalization of high-throughput methodologies for screening the efficacy of active compounds are commonly reported for these kinds of organisms. In this study, a comprehensive picture of two high-throughput practices for efficient fungicide screening against plant-pathogenic oomycetes has been provided. Different protocols using multiwell plates were validated on approved crop protection products using Phytophthora infestans and Pythium ultimum as the model species. In addition, detailed statistical inputs useful for the analysis of data related to the efficacy of screenings are included

Pathogen Adaptation to American (Rpv3-1) and Eurasian (Rpv29) Grapevine Loci Conferring Resistance to Downy Mildew

Durable resistance is a key objective in genetic improvement for disease resistance in grapevines, which must survive for years in the field in the presence of adaptable pathogen populations. In this study, the adaptation of 72 Northern Italian isolates of Plasmopara viticola, the downy mildew agent, has been investigated into Bianca, possessing Rpv3-1, the most frequently exploited resistance locus for genetic improvement, and Mgaloblishvili, a Vitis vinifera variety possessing the newly discovered Rpv29 locus. Infection parameters (latency period, infection frequency, and disease severity) and oospore production and viability were evaluated and compared to those of Pinot noir, the susceptible reference. The expected levels of disease control were achieved by both resistant cultivars (&gt;90% on Bianca; &gt;25% on Mgaloblishvili), despite the high frequency of isolates able to grow on one (28%) or both (46%) accessions. The disease incidence and severity were limited by both resistant cultivars and the strains able to grow on resistant accessions showed signatures of fitness penalties (reduced virulence, infection frequency, and oospore density). Together, these results indicate an adequate pathogen control but suitable practices must be adopted in the field to prevent the diffusion of the partially adapted P. viticola strains to protect resistance genes from erosion

A comparative cell wall analysis of Trichoderma spp. confirms a conserved polysaccharide scaffold and suggests an important role for chitosan in mycoparasitism

ABSTRACT Fungal cell walls are dynamic extracellular matrices that enable efficient adaptation to changing environments. While the cell wall compositions of yeasts, human, and plant pathogenic fungi have been studied to some extent, the cell walls of mycoparasites remain poorly characterized. Trichoderma species comprise a diverse group of soil fungi with different survival strategies and lifestyles. The comparative study of cell wall carbohydrate-active enzymes in 13 Trichoderma spp. revealed that the types of enzymes involved in chitin and chitosan metabolism are phylogenetically distant between mycoparasitic and saprotrophic species. Here, we compare the carbohydrate composition and function of the cell wall of a saprotrophic strain Trichoderma reesei with that of the mycoparasitic, biological control agent Trichoderma atroviride. Monosaccharide and glycosidic linkage analyses as well as dual in situ interaction assays showed that the cell wall polysaccharide composition is conserved between both species, except for the amounts of chitin detected. The results suggest that the observed accumulation of chitosan during mycoparasitism may prevent host recognition. Remarkably, Trichoderma atroviride undergoes dynamic cell wall adaptations during both vegetative development and mycoparasitism, which appears to be confirmed by an evolutionarily expanded group of specialized enzymes. Overall, our analyses support the notion that habitat specialization is reflected in cell wall architecture and that plastic chitin remodeling may confer an advantage to mycoparasites, ultimately enabling the successful invasion and parasitism of plant pathogens. This information may potentially be exploited for the control of crop diseases using biological agents.IMPORTANCETrichoderma species are emerging model fungi for the development of biocontrol agents and are used in industrial biotechnology as efficient enzyme producers. Fungal cell walls are complex structures that differ in carbohydrate, protein, and enzyme composition across taxa. Here, we present a chemical characterization of the cell walls of two Trichoderma spp., namely the predominantly saprotrophic Trichoderma reesei and the mycoparasite Trichoderma atroviride. Chemical profiling revealed that Trichoderma spp. remodel their cell wall to adapt to particular lifestyles, with dynamic changes during vegetative development. Importantly, we found that chitosan accumulation during mycoparasitism of a fungal host emerged as a sophisticated strategy underpinning an effective attack. These insights shed light on the molecular mechanisms that allow mycoparasites to overcome host defenses and can be exploited to improve the application of T. atroviride in biological pest control. Moreover, our results provide valuable information for targeting the fungal cell wall for therapeutic purposes

Influence of Nitrogen on Grapevine Susceptibility to Downy Mildew

Downy mildew, caused by the obligate parasite Plasmopara viticola, is one of the most important threats to viticulture. The exploitation of resistant and susceptibility traits of grapevine is one of the most promising ways to increase the sustainability of disease management. Nitrogen (N) fertilization is known for influencing disease severity in the open field, but no information is available on its effect on plant-pathogen interaction. A previous RNAseq study showed that several genes of N metabolism are differentially regulated in grapevine upon P. viticola inoculation, and could be involved in susceptibility or resistance to the pathogen. The aim of this study was to evaluate if N fertilization influences: (i) the foliar leaf content and photosynthetic activity of the plant, (ii) P. viticola infectivity, and (iii) the expression of the candidate susceptibility/resistance genes. Results showed that N level positively correlated with P. viticola infectivity, confirming that particular attention should be taken in vineyard to the fertilization, but did not influence the expression of the candidate genes. Therefore, these genes are manipulated by the pathogen and can be exploited for developing new, environmentally friendly disease management tools, such as dsRNAs, to silence the susceptibility genes or breeding for resistance