Sophie Trouvelot

A β-1,3 Glucan Sulfate Induces Resistance in Grapevine against Plasmopara viticola Through Priming of Defense Responses, Including HR-like Cell Death

Sulfated laminarin (PS3) has been shown previously to be an elicitor of plant defense reactions in tobacco and Arabidopsis and to induce protection against tobacco mosaic virus. Here, we have demonstrated the efficiency of PS3 in protecting a susceptible grapevine cultivar (Vitis vinifera cv. Marselan) against downy mildew (Plasmopara viticola) under glasshouse conditions. This induced resistance was associated with potentiated H2O2 production at the infection sites, upregulation of defense-related genes, callose and phenol depositions, and hypersensitive response-like cell death. Interestingly, similar responses were observed following P. viticola inoculation in a tolerant grapevine hybrid cultivar (Solaris). A pharmacological approach led us to conclude that both callose synthesis and jasmonic acid pathway contribute to PS3-induced resistance.

Carbohydrates in plant immunity and plant protection: roles and potential application as foliar sprays

Increasing interest is devoted to carbohydrates for their roles in plant immunity. Some of them are elicitors of plant defenses whereas other ones act as signaling molecules in a manner similar to phytohormones. This review first describes the main classes of carbohydrates associated to plant immunity, their role and mode of action. More precisely, the state of the art about perception of “PAMP, MAMP, and DAMP (Pathogen-, Microbe-, Damage-Associated Molecular Patterns) type” oligosaccharides is presented and examples of induced defense events are provided. A particular attention is paid to the structure/activity relationships of these compounds. The role of sugars as signaling molecules, especially in plant microbe interactions, is also presented. Secondly, the potentialities and limits of foliar sprays of carbohydrates to stimulate plant immunity for crop protection against diseases are discussed, with focus on the roles of the leaf cuticle and phyllosphere microflora.

Colonization of Flax Roots and Early Physiological Responses of Flax Cells Inoculated with Pathogenic and Nonpathogenic Strains of Fusarium oxysporum

ABSTRACT Fusarium oxysporum includes nonpathogenic strains and pathogenic strains that can induce necrosis or tracheomycosis in plants. The objective of this study was to compare the abilities of a pathogenic strain (Foln3) and a nonpathogenic strain (Fo47) to colonize flax roots and to induce early physiological responses in flax cell culture suspensions. Both strains colonized the outer cortex of the root; however, plant defense reactions, i.e., the presence of wall appositions, osmiophilic material, and collapsed cells, were less frequent and less intense in a root colonized by Foln3 than by Fo47. Early physiological responses were measured in flax cell suspensions confronted with germinated microconidia of both strains. Both pathogenic (Foln3) and nonpathogenic strains (Fo47) triggered transient H2O2 production in the first few minutes of the interaction, but the nonpathogenic strain also induced a second burst 3 h postinoculation. Ca2+ influx was more intense in cells inoculated with Fo47 than in cells inoculated with Foln3. Similarly, alkalinization of the extracellular medium was higher with Fo47 than with Foln3. Inoculation of the fungi into flax cell suspensions induced cell death 10 to 20 h postinoculation, with a higher percentage of dead cells observed with Fo47 than with Foln3 beginning at 14 h. This is the first report showing that early physiological responses of flax cells can be used to distinguish pathogenic and nonpathogenic strains of the soil-borne fungus F. oxysporum.

β-Aminobutyric acid primes an NADPH oxidase-dependent reactive oxygen species production during grapevine-triggered immunity.

The molecular mechanisms underlying the process of priming are poorly understood. In the present study, we investigated the early signaling events triggered by beta-aminobutyric acid (BABA), a well-known priming-mediated plant resistance inducer. Our results indicate that, in contrast to oligogalacturonides (OG), BABA does not elicit typical defense-related early signaling events nor defense-gene expression in grapevine. However, in OG-elicited cells pretreated with BABA, production of reactive oxygen species (ROS) and expression of the respiratory-burst oxidase homolog RbohD gene were primed. In response to the causal agent of downy mildew Plasmopara viticola, a stronger ROS production was specifically observed in BABA-treated leaves. This process was correlated with an increased resistance. The NADPH oxidase inhibitor diphenylene iodonium (DPI) abolished this primed ROS production and reduced the BABA-induced resistance (BABA-IR). These results suggest that priming of an NADPH oxidase-dependent ROS production contributes to BABA-IR in the Vitis-Plasmopara pathosystem.

Are grapevine stomata involved in the elicitor-induced protection against downy mildew?

Stomata, natural pores bordered by guard cells, regulate transpiration and gas exchanges between plant leaves and the atmosphere. These natural openings also constitute a way of penetration for microorganisms. In plants, the perception of potentially pathogenic microorganisms or elicitors of defense reactions induces a cascade of events, including H(2)O(2) production, that allows the activation of defense genes, leading to defense reactions. Similar signaling events occur in guard cells in response to the perception of abscisic acid (ABA), leading to stomatal closure. Moreover, few elicitors were reported to induce stomatal closure in Arabidopsis and Vicia faba leaves. Because responses to ABA and elicitors share common signaling events, it led us to question whether stomatal movements and H(2)O(2) production in guard cells could play a key role in elicitor-induced protection against pathogens that use stomata for infection. This study was performed using the grapevine-Plasmopara viticola pathosystem. Using epidermal peels, we showed that, as for ABA, the elicitor-induced stomatal closure is mediated by reactive oxygen species (ROS) production in guard cells. In plants, we observed that the protection against downy mildew induced by some elicitors is probably not due only to effects on stomatal movements or to a guard-cell-specific activation of ROS production.

The sulfated laminarin triggers a stress transcriptome before priming the SA- and ROS-dependent defenses during grapevine's induced resistance against Plasmopara viticola.

Grapevine (Vitis vinifera) is susceptible to many pathogens which cause significant losses to viticulture worldwide. Chemical control is available, but agro-ecological concerns have raised interest in alternative methods, especially in triggering plant immunity by elicitor treatments. The β-glucan laminarin (Lam) and its sulfated derivative (PS3) have been previously demonstrated to induce resistance in grapevine against downy mildew (Plasmopara viticola). However, if Lam elicits classical grapevine defenses such as oxidative burst, pathogenesis-related (PR)-proteins and phytoalexin production, PS3 triggered grapevine resistance via a poorly understood priming phenomenon. The aim of this study was to identify the molecular mechanisms of the PS3-induced resistance. For this purpose we studied i) the signaling events and transcriptome reprogramming triggered by PS3 treatment on uninfected grapevine, ii) grapevine immune responses primed by PS3 during P. viticola infection. Our results showed that i) PS3 was unable to elicit reactive oxygen species (ROS) production, cytosolic Ca2+ concentration variations, mitogen-activated protein kinase (MAPK) activation but triggered a long lasting plasma membrane depolarization in grapevine cells, ii) PS3 and Lam shared a common stress-responsive transcriptome profile that partly overlapped the salicylate- (SA) and jasmonate-(JA)-dependent ones. After P. viticola inoculation, PS3 specifically primed the SA- and ROS-dependent defense pathways leading to grapevine induced resistance against this biotroph. Interestingly pharmacological approaches suggested that the plasma membrane depolarization and the downstream ROS production are key events of the PS3-induced resistance.

Changes in Carbohydrate Metabolism in Plasmopara viticola-Infected Grapevine Leaves

The oomycete Plasmopara viticola is responsible for downy mildew, a severe grapevine disease. In infected grapevine leaves, we have observed an abnormal starch accumulation at the end of the dark period, suggesting modifications in starch metabolism. Therefore, several complementary approaches, including transcriptomic analyses, measurements of enzyme activities, and sugar quantification, were performed in order to investigate and to understand the effects of P. viticola infection on leaf starch and-to a larger extent-carbohydrate metabolism. Our results indicate that starch accumulation is associated with an increase in ADP-glucose pyrophosphorylase (AGPase) activity and modifications in the starch degradation pathway, especially an increased α-amylase activity. Together with these alterations in starch metabolism, we have observed an accumulation of hexoses, an increase in invertase activity, and a reduction of photosynthesis, indicating a source-to-sink transition in infected leaf tissue. Additionally, we have measured an accumulation of the disaccharide trehalose correlated to an increased trehalase gene expression and enzyme activity. Altogether, these results highlight a dramatic alteration of carbohydrate metabolism correlated with later stages of P. viticola development in leaves.

Molecular Interactions of β-(1→3)-Glucans with Their Receptors

β-(1→3)-Glucans can be found as structural polysaccharides in cereals, in algae or as exo-polysaccharides secreted on the surfaces of mushrooms or fungi. Research has now established that β-(1→3)-glucans can trigger different immune responses and act as efficient immunostimulating agents. They constitute prevalent sources of carbons for microorganisms after subsequent recognition by digesting enzymes. Nevertheless, mechanisms associated with both roles are not yet clearly understood. This review focuses on the variety of elucidated molecular interactions that involve these natural or synthetic polysaccharides and their receptors, i.e., Dectin-1, CR3, glycolipids, langerin and carbohydrate-binding modules.

Recovery of Fusarium oxysporum Fo47 Mutants Affected in Their Biocontrol Activity After Transposition of the Fot1 Element

ABSTRACT To investigate the biocontrol mechanisms by which the antagonistic Fusarium oxysporum strain Fo47 is active against Fusarium wilt, a Fot1 transposon-mediated insertional mutagenesis approach was adopted to generate mutants affected in their antagonistic activity. Ninety strains in which an active Fot1 copy had transposed were identified with a phenotypic assay for excision and tested for their biocontrol activity against F. oxysporum f. sp. lini on flax in greenhouse experiments. Sixteen strains were affected in their capacity to protect flax plants, either positively (more antagonistic than Fo47) or negatively (less antagonistic). The molecular characterization of these mutants confirms the excision of Fot1 and its reinsertion in most of the cases. Moreover, we demonstrate that other transposable elements such as Fot2, impala, and Hop have no transposition activity in the mutant genomes. The phenotypic characterization of these mutants shows that they are affected neither in their in vitro growth habit nor in their competitiveness in soil compared with wild-type strain Fo47. These results show that mutants are not impaired in their saprophytic phase and suggest that the altered biocontrol phenotype should likely be expressed during the interaction with the host plant.

Image analysis methods for assessment of H2O2 production and Plasmopara viticola development in grapevine leaves: Application to the evaluation of resistance to downy mildew

The grapevine downy mildew (Plasmopara viticola) provokes severe damages and destroys the harvest in the absence of an effective protection. Numerous fungicide treatments are thus generally necessary. To promote a sustainable production, alternative strategies of protection including new antifungal molecules, resistant genotypes or elicitor-induced resistance are under trial. To evaluate the relevance of these strategies, resistance tests are required. In this context, three image analysis methods were developed to read the results of tests performed to assess P. viticola sporulation and mycelial development, and H(2)O(2) production in leaves. They have been validated using elicitors of plant defenses. These methods are reliable, innovative, rapid, and their modular concept allows their further adaptation to other host-pathogen systems.