Genetic control of abiotic stress-related specialized metabolites in sunflower.

BACKGROUND: Abiotic stresses in plants include all the environmental conditions that significantly reduce yields, like drought and heat. One of the most significant effects they exert at the cellular level is the accumulation of reactive oxygen species, which cause extensive damage. Plants possess two mechanisms to counter these molecules, i.e. detoxifying enzymes and non-enzymatic antioxidants, which include many classes of specialized metabolites. Sunflower, the fourth global oilseed, is considered moderately drought resistant. Abiotic stress tolerance in this crop has been studied using many approaches, but the control of specialized metabolites in this context remains poorly understood. Here, we performed the first genome-wide association study using abiotic stress-related specialized metabolites as molecular phenotypes in sunflower. After analyzing leaf specialized metabolites of 450 hybrids using liquid chromatography-mass spectrometry, we selected a subset of these compounds based on their association with previously known abiotic stress-related quantitative trait loci. Eventually, we characterized these molecules and their associated genes. RESULTS: We putatively annotated 30 compounds which co-localized with abiotic stress-related quantitative trait loci and which were associated to seven most likely candidate genes. A large proportion of these compounds were potential antioxidants, which was in agreement with the role of specialized metabolites in abiotic stresses. The seven associated most likely candidate genes, instead, mainly belonged to cytochromes P450 and glycosyltransferases, two large superfamilies which catalyze greatly diverse reactions and create a wide variety of chemical modifications. This was consistent with the high plasticity of specialized metabolism in plants. CONCLUSIONS: This is the first characterization of the genetic control of abiotic stress-related specialized metabolites in sunflower. By providing hints concerning the importance of antioxidant molecules in this biological context, and by highlighting some of the potential molecular mechanisms underlying their biosynthesis, it could pave the way for novel applications in breeding. Although further analyses will be required to better understand this topic, studying how antioxidants contribute to the tolerance to abiotic stresses in sunflower appears as a promising area of research.

The dynamics of pattern matching in camouflaging cuttlefish.

Many cephalopods escape detection using camouflage1. This behaviour relies on a visual assessment of the surroundings, on an interpretation of visual-texture statistics2-4 and on matching these statistics using millions of skin chromatophores that are controlled by motoneurons located in the brain5-7. Analysis of cuttlefish images proposed that camouflage patterns are low dimensional and categorizable into three pattern classes, built from a small repertoire of components8-11. Behavioural experiments also indicated that, although camouflage requires vision, its execution does not require feedback5,12,13, suggesting that motion within skin-pattern space is stereotyped and lacks the possibility of correction. Here, using quantitative methods14, we studied camouflage in the cuttlefish Sepia officinalis as behavioural motion towards background matching in skin-pattern space. An analysis of hundreds of thousands of images over natural and artificial backgrounds revealed that the space of skin patterns is high-dimensional and that pattern matching is not stereotyped-each search meanders through skin-pattern space, decelerating and accelerating repeatedly before stabilizing. Chromatophores could be grouped into pattern components on the basis of their covariation during camouflaging. These components varied in shapes and sizes, and overlay one another. However, their identities varied even across transitions between identical skin-pattern pairs, indicating flexibility of implementation and absence of stereotypy. Components could also be differentiated by their sensitivity to spatial frequency. Finally, we compared camouflage to blanching, a skin-lightening reaction to threatening stimuli. Pattern motion during blanching was direct and fast, consistent with open-loop motion in low-dimensional pattern space, in contrast to that observed during camouflage.

Differential carbohydrate-active enzymes and secondary metabolite production by the grapevine trunk pathogen Neofusicoccum parvum Bt-67 grown on host and non-host biomass.

Neofusicoccum parvum is one of the most aggressive Botryosphaeriaceae species associated with grapevine trunk diseases. This species may secrete enzymes capable of overcoming the plant barriers, leading to wood colonization. In addition to their roles in pathogenicity, there is an interest in taking advantage of N. parvum carbohydrate-active enzymes (CAZymes), related to plant cell wall degradation, for lignocellulose biorefining. Furthermore, N. parvum produces toxic secondary metabolites that may contribute to its virulence. In order to increase knowledge on the mechanisms underlying pathogenicity and virulence, as well as the exploration of its metabolism and CAZymes for lignocellulose biorefining, we evaluated the N. parvum strain Bt-67 capacity in producing lignocellulolytic enzymes and secondary metabolites when grown in vitro with two lignocellulosic biomasses: grapevine canes (GP) and wheat straw (WS). For this purpose, a multiphasic study combining enzymology, transcriptomic, and metabolomic analyses was performed. Enzyme assays showed higher xylanase, xylosidase, arabinofuranosidase, and glucosidase activities when the fungus was grown with WS. Fourier transform infrared (FTIR) spectroscopy confirmed the lignocellulosic biomass degradation caused by the secreted enzymes. Transcriptomics indicated that the N. parvum Bt-67 gene expression profiles in the presence of both biomasses were similar. In total, 134 genes coding CAZymes were up-regulated, where 94 of them were expressed in both biomass growth conditions. Lytic polysaccharide monooxygenases (LPMOs), glucosidases, and endoglucanases were the most represented CAZymes and correlated with the enzymatic activities obtained. The secondary metabolite production, analyzed by high-performance liquid chromatography-ultraviolet/visible spectophotometry-mass spectrometry (HPLC-UV/Vis-MS), was variable depending on the carbon source. The diversity of differentially produced metabolites was higher when N. parvum Bt-67 was grown with GP. Overall, these results provide insight into the influence of lignocellulosic biomass on virulence factor expressions. Moreover, this study opens the possibility of optimizing the enzyme production from N. parvum with potential use for lignocellulose biorefining.

Botryosphaeriaceae gene machinery: Correlation between diversity and virulence.

The Botryosphaeriaceae family comprises numerous fungal pathogens capable of causing economically meaningful diseases in a wide range of crops. Many of its members can live as endophytes and turn into aggressive pathogens following the onset of environmental stress events. Their ability to cause disease may rely on the production of a broad set of effectors, such as cell wall-degrading enzymes, secondary metabolites, and peptidases. Here, we conducted comparative analyses of 41 genomes representing six Botryosphaeriaceae genera to provide insights into the genetic features linked to pathogenicity and virulence. We show that these Botryosphaeriaceae genomes possess a large diversity of carbohydrate-active enzymes (CAZymes; 128 families) and peptidases (45 families). Botryosphaeria, Neofusicoccum, and Lasiodiplodia presented the highest number of genes encoding CAZymes involved in the degradation of the plant cell wall components. The genus Botryosphaeria also exhibited the highest abundance of secreted CAZymes and peptidases. Generally, the secondary metabolites gene cluster profile was consistent in the Botryosphaeriaceae family, except for Diplodia and Neoscytalidium. At the strain level, Neofusicoccum parvum NpBt67 stood out among all the Botryosphaeriaceae genomes, presenting a higher number of secretome constituents. In contrast, the Diplodia strains showed the lowest richness of the pathogenicity- and virulence-related genes, which may correlate with their low virulence reported in previous studies. Overall, these results contribute to a better understanding of the mechanisms underlying pathogenicity and virulence in remarkable Botryosphaeriaceae species. Our results also support that Botryosphaeriaceae species could be used as an interesting biotechnological tool for lignocellulose fractionation and bioeconomy.

The Combination of Both Heat and Water Stresses May Worsen Botryosphaeria Dieback Symptoms in Grapevine.

(1) Background: Grapevine trunk diseases (GTDs) have become a global threat to vineyards worldwide. These diseases share three main common features. First, they are caused by multiple pathogenic micro-organisms. Second, these pathogens often maintain a long latent phase, which makes any research in pathology and symptomatology challenging. Third, a consensus is raising to pinpoint combined abiotic stresses as a key factor contributing to disease symptom expression. (2) Methods: We analyzed the impact of combined abiotic stresses in grapevine cuttings artificially infected by two fungi involved in Botryosphaeria dieback (one of the major GTDs), Neofusicoccum parvum and Diplodia seriata. Fungal-infected and control plants were subjected to single or combined abiotic stresses (heat stress, drought stress or both). Disease intensity was monitored thanks to the measurement of necrosis area size. (3) Results and conclusions: Overall, our results suggest that combined stresses might have a stronger impact on disease intensity upon infection by the less virulent pathogen Diplodia seriata. This conclusion is discussed through the impact on plant physiology using metabolomic and transcriptomic analyses of leaves sampled for the different conditions.

The microbiota of the grapevine holobiont: A key component of plant health.

BACKGROUND: Grapevine is a woody, perennial plant of high economic importance worldwide. Like other plants, it lives in close association with large numbers of microorganisms. Bacteria, fungi and viruses are structured in communities, and each individual can be beneficial, neutral or harmful to the plant. In this sense, microorganisms can interact with each other and regulate plant functions (including immunity) and even provide new ones. Thus, the grapevine associated with its microbial communities constitutes a supra-organism, also called a holobiont, whose functioning is linked to established plant-microorganism interactions. AIM OF REVIEW: The overall health of the plant may be conditioned by the diversity and structure of microbial communities. Consequently, an optimal microbial composition will consist of a microbial balance allowing the plant to be healthy. Conversely, an imbalance of microbial populations could lead to (or be generated by) a decline of the plant. The microbiome is an active component of the host also responsive to biotic and abiotic changes; in that respect, a better understanding of the most important drivers of the composition of plant microbiomes is needed. KEY SCIENTIFIC CONCEPTS OF REVIEW: This article presents the current state of the art about the grapevine microbiota and its composition according to the plant compartments and the influencing factors. We also focus on situations of imbalance, in particular during plant disease or decline. Finally, we discuss the possible interest of microbial engineering in an agrosystem such as viticulture.

Diversity of Neofusicoccum parvum for the Production of the Phytotoxic Metabolites (-)-Terremutin and (R)-Mellein.

Two Neofusicoccumparvum isolates and a UV mutant were characterized for their phytotoxin production in vitro, their pathogenicity on grapevine, and their genome sequenced. The isolate Np-Bt67 produced high level of (-)-terremutin, but almost no (R)-mellein, and it was the most aggressive on grapevine, triggering apoplexy. Similar symptoms were not induced by purified (-)-terremutin. The isolate Bourgogne S-116 (Np-B) produced 3-fold less (-)-terremutin and high amounts of (R)-mellein, but it was less aggressive on grapevine than Np-Bt67. The UV9 mutant obtained from Np-B (NpB-UV9) no longer produced (-)-terremutin but overproduced (R)-mellein by 2.5-fold, and it was as pathogenic as its parent. NpB-UV9 differed from its parent by simple mutations in two genes (transcription factor UCR-NP2_6692, regulatory protein UCR-NP2_9007), not located neither near (R)-mellein, nor (-)-terremutin biosynthetic genes, but likely involved in the control of (-)-terremutin biosynthesis. Grapevine immunity was disturbed upon challenge with these pathogens or purified phytotoxins, leading to an upregulation of SA-dependent defenses, while (-)-terremutin interfered with host JA/ET-dependent defenses. Our results suggest that neither (-)-terremutin nor (R)-mellein alone is essential for the pathogenicity of N. parvum on grapevine, since isolate/mutant non-producing these toxins in vitro is pathogenic. However, these phytotoxins could play a quantitative role in the infection process.

Assessment of a New Copper-Based Formulation to Control Esca Disease in Field and Study of Its Impact on the Vine Microbiome, Vine Physiology and Enological Parameters of the Juice.

Copper-based preparations have been used for more than 100 years in viticulture to control downy mildew caused by Plasmopara viticola. LC2017, and a new low-copper-based formulation, has been developed to control grapevine trunk diseases (GTDs). Previous greenhouse studies showed the potential of LC2017 to control GTDs by both fungistatic and plant defense elicitor effects. Here, we further characterize the effects of LC2017 in the field determining its impact on: (i) incidence of Esca, (ii) the vine microbiome, (iii) the vine physiology and (iv) enological parameters of juices. We observed a progressive decrease of cumulate Esca incidence in treated vines over the years with annual fluctuation related to the known erratic emergence of GTD symptoms. Neither harmful effects of LC2017 on the vine microbiota, nor on vine physiology were observed (at both transcriptomic and metabolomic levels). Similarly, no impact of LC2017 was observed on the enological properties of berries except for sugar content in juice from esca-diseased vines. The most important result concerns the transcriptomic profiles: that of diseased and LC2017 treated vines differs from that of disease untreated ones, showing a treatment effect. Moreover, the transcriptomic profile of diseased and LC2017-treated vines is similar to that of untreated asymptomatic vines, suggesting control of the disease.

In planta Activity of the Novel Copper Product HA + Cu(II) Based on a Biocompatible Drug Delivery System on Vine Physiology and Trials for the Control of Botryosphaeria Dieback.

The growing concerns on human and environment health are forcing the plant protection industry toward the formulation of more eco-sustainable plant protection products (PPP), both efficient and innovative in their approach to disease control. A large number of these innovative formulations now rely on a combination of pathogens antagonistic properties and stimulation of natural plant defense to pathogens. The formulation HA + Cu(II), in which copper is delivered to the plants by the drug-delivery molecule hydroxyapatite (HA), was found efficient against the grapevine pathogens Plasmopara viticola and Phaeoacremonium minimum and able to induce the host-plant defense system. We investigated the HA + Cu(II) impacts on grapevine physiology, both in uninfected and when infected by the Botryosphaeria dieback agents Diplodia seriata and Neofusicoccum parvum. This study of plant physiology and disease impact were addressed to evaluate both the HA + Cu(II) potential as a plant defense elicitor and its possible and future use as PPP in vineyard. Our results showed that HA + Cu(II) induced several key-defense genes without negatively affecting plant growth and photosynthetic activity. In addition, fungistatic effect on the two Botryosphaeriaceae at the in planta tested concentrations is reported. Altogether, our results obtained under controlled conditions fully support the potential of HA + Cu(II) as a promising PPP toward grapevine trunk diseases in vineyard.

Metabolic Profile Discriminates and Predicts Arabidopsis Susceptibility to Virus under Field Conditions.

As obligatory parasites, plant viruses alter host cellular metabolism. There is a lack of information on the variability of virus-induced metabolic responses among genetically diverse plants in a natural context with daily changing conditions. To decipher the metabolic landscape of plant-virus interactions in a natural setting, twenty-six and ten accessions of Arabidopsis thaliana were inoculated with Turnip mosaic virus (TuMV), in two field experiments over 2 years. The accessions were measured for viral accumulation, above-ground biomass, targeted and untargeted metabolic profiles. The phenotypes of the accessions ranged from susceptibility to resistance. Susceptible and resistant accessions were shown to have different metabolic routes after inoculation. Susceptible genotypes accumulate primary and secondary metabolites upon infection, at the cost of hindered growth. Twenty-one metabolic signatures significantly accumulated in resistant accessions whereas they maintained their growth as mock-inoculated plants without biomass penalty. Metabolic content was demonstrated to discriminate and be highly predictive of the susceptibility of inoculated Arabidopsis. This study is the first to describe the metabolic landscape of plant-virus interactions in a natural setting and its predictive link to susceptibility. It provides new insights on plant-virus interactions. In this undomesticated species and in ecologically realistic conditions, growth and resistance are in a permanent conversation.

Woody Plant Declines. What's Wrong with the Microbiome?

Woody plant (WP) declines have multifactorial determinants as well as a biological and economic reality. The vascular system of WPs involved in the transport of carbon, nitrogen, and water from sources to sinks has a seasonal activity, which places it at a central position for mediating plant-environment interactions from nutrient cycling to community assembly and for regulating a variety of processes. To limit effects and to fight against declines, we propose: (i) to consider the WP and its associated microbiota as an holobiont and as a set of functions; (ii) to consider simultaneously, without looking at what comes first, the physiological or pathogenic disorders; and (iii) to define pragmatic strategies, including preventive and curative agronomical practices based on microbiota engineering.

Metabolomic characterization of sunflower leaf allows discriminating genotype groups or stress levels with a minimal set of metabolic markers.

INTRODUCTION: Plant and crop metabolomic analyses may be used to study metabolism across genetic and environmental diversity. Complementary analytical strategies are useful for investigating metabolic changes and searching for biomarkers of response or performance. METHODS AND OBJECTIVES: The experimental material consisted in eight sunflower lines with two line status, four restorers (R, used as males) and four maintainers (B, corresponding to females) routinely used for sunflower hybrid varietal production, respectively to complement or maintain the cytoplasmic male sterility PET1. These lines were either irrigated at full soil capacity (WW) or submitted to drought stress (DS). Our aim was to combine targeted and non-targeted metabolomics to characterize sunflower leaf composition in order to investigate the effect of line status genotypes and environmental conditions and to find the best and smallest set of biomarkers for line status and stress response using a custom-made process of variables selection. RESULTS: Five hundred and eighty-eight metabolic variables were measured by using complementary analytical methods such as 1H-NMR, MS-based profiles and targeted analyses of major metabolites. Based on statistical analyses, a limited number of markers were able to separate WW and DS samples in a more discriminant manner than previously published physiological data. Another metabolic marker set was able to discriminate line status. CONCLUSION: This study underlines the potential of metabolic markers for discriminating genotype groups and environmental conditions. Their potential use for prediction is discussed.

Untargeted Analysis of Semipolar Compounds by LC-MS and Targeted Analysis of Fatty Acids by GC-MS/GC-FID: From Plant Cultivation to Extract Preparation.

The way plants are grown and samples are harvested, prepared, and extracted has a profound impact on the output of a metabolomics experiment. In this chapter, we detail the experimental procedures from plant cultivation to extract preparation, in order to avoid difficulties that could result in contamination or undesired changes of the analytes. Two plant organs are mentioned as examples: tomato fruits (Solanum lycopersicum) and flax seeds (Linum usitatissimum). Extractions designed for the untargeted analysis of semipolar compounds by liquid chromatography-mass spectrometry (LC-MS) and targeted analysis of fatty acids by gas chromatography-mass spectrometry (GC-MS) or gas chromatography with flame ionization detector (GC-FID) are described.

Leaf Starch Turnover Occurs in Long Days and in Falling Light at the End of the Day.

We investigated whether starch degradation occurs at the same time as starch synthesis in Arabidopsis (Arabidopsis thaliana) leaves in the light. Starch accumulated in a linear fashion for about 12 h after dawn, then accumulation slowed and content plateaued. Following decreases in light intensity, the rate of accumulation of starch declined in proportion to the decline in photosynthesis if the decrease occurred <10 h after dawn, but accumulation ceased or loss of starch occurred if the same decrease in light intensity was imposed more than 10 h after dawn. These changes in starch accumulation patterns after prolonged periods in the light occurred at both high and low starch contents and were not related to time-dependent changes in either the rate of photosynthesis or the partitioning of assimilate between starch and Suc, as assessed from metabolite measurements and 14CO2 pulse experiments. Instead, measurements of incorporation of 13C from 13CO2 into starch and of levels of the starch degradation product maltose showed that substantial starch degradation occurred simultaneously with synthesis at time points >14 h after dawn and in response to decreases in light intensity that occurred >10 h after dawn. Starch measurements in circadian clock mutants suggested that the clock influences the timing of onset of degradation. We conclude that the propensity for leaf starch to be degraded increases with time after dawn. The importance of this phenomenon for efficient use of carbon for growth in long days and for prevention of starvation during twilight is discussed.

Fortune telling: metabolic markers of plant performance.

BACKGROUND: In the last decade, metabolomics has emerged as a powerful diagnostic and predictive tool in many branches of science. Researchers in microbes, animal, food, medical and plant science have generated a large number of targeted or non-targeted metabolic profiles by using a vast array of analytical methods (GC-MS, LC-MS, 1H-NMR….). Comprehensive analysis of such profiles using adapted statistical methods and modeling has opened up the possibility of using single or combinations of metabolites as markers. Metabolic markers have been proposed as proxy, diagnostic or predictors of key traits in a range of model species and accurate predictions of disease outbreak frequency, developmental stages, food sensory evaluation and crop yield have been obtained. AIM OF REVIEW: (i) To provide a definition of plant performance and metabolic markers, (ii) to highlight recent key applications involving metabolic markers as tools for monitoring or predicting plant performance, and (iii) to propose a workable and cost-efficient pipeline to generate and use metabolic markers with a special focus on plant breeding. KEY MESSAGE: Using examples in other models and domains, the review proposes that metabolic markers are tending to complement and possibly replace traditional molecular markers in plant science as efficient estimators of performance.

Plant polysaccharides initiate underground crosstalk with bacilli by inducing synthesis of the immunogenic lipopeptide surfactin.

Some plant-associated bacteria such as Bacillus sp. can protect their host from pathogen ingress and this biocontrol activity correlates with their potential to form multiple antibiotics upon in vitro growth. However, our knowledge on antibiotic production by soil bacilli evolving on roots in natural conditions is still limited. In this work, antibiome imaging first revealed that the lipopeptide surfactin is the main bacterial ingredient produced in planta within the first hours of interaction with root tissues. We further demonstrated that surfactin synthesis is specifically stimulated upon perception of plant cell wall polymers such as xylan or arabinogalactan, leading to fast accumulation of micromolar amounts in the root environment. At such concentrations, the lipopeptide may not only favour the ecological fitness of the producing strain in term of root colonization, but also triggers systemic resistance in the host plant. This surfactin-induced immunity primes the plant to better resist further pathogen ingress, and involves only limited expression of defence-related molecular events and does not provoke seedling growth inhibition. By contrast with the strong response mounted upon perception of pathogens, this strongly attenuated defensive reaction induced by surfactin in plant tissues should help Bacillus to be tolerated as saprophytic partner by its host.

Cyclic lipopeptides from Bacillus subtilis activate distinct patterns of defence responses in grapevine.

Non-self-recognition of microorganisms partly relies on the perception of microbe-associated molecular patterns (MAMPs) and leads to the activation of an innate immune response. Bacillus subtilis produces three main families of cyclic lipopeptides (LPs), namely surfactins, iturins and fengycins. Although LPs are involved in induced systemic resistance (ISR) activation, little is known about defence responses induced by these molecules and their involvement in local resistance to fungi. Here, we showed that purified surfactin, mycosubtilin (iturin family) and plipastatin (fengycin family) are perceived by grapevine plant cells. Although surfactin and mycosubtilin stimulated grapevine innate immune responses, they differentially activated early signalling pathways and defence gene expression. By contrast, plipastatin perception by grapevine cells only resulted in early signalling activation. Gene expression analysis suggested that mycosubtilin activated salicylic acid (SA) and jasmonic acid (JA) signalling pathways, whereas surfactin mainly induced an SA-regulated response. Although mycosubtilin and plipastatin displayed direct antifungal activity, only surfactin and mycosubtilin treatments resulted in a local long-lasting enhanced tolerance to the necrotrophic fungus Botrytis cinerea in grapevine leaves. Moreover, challenge with specific strains overproducing surfactin and mycosubtilin led to a slightly enhanced stimulation of the defence response compared with the LP-non-producing strain of B. subtilis. Altogether, our results provide the first comprehensive view of the involvement of LPs from B. subtilis in grapevine plant defence and local resistance against the necrotrophic pathogen Bo. cinerea. Moreover, this work is the first to highlight the ability of mycosubtilin to trigger an immune response in plants.

Regulatory properties of ADP glucose pyrophosphorylase are required for adjustment of leaf starch synthesis in different photoperiods.

Arabidopsis (Arabidopsis thaliana) leaves synthesize starch faster in short days than in long days, but the mechanism that adjusts the rate of starch synthesis to daylength is unknown. To understand this mechanism, we first investigated whether adjustment occurs in mutants lacking components of the circadian clock or clock output pathways. Most mutants adjusted starch synthesis to daylength, but adjustment was compromised in plants lacking the GIGANTEA or FLAVIN-BINDING, KELCH REPEAT, F BOX1 components of the photoperiod-signaling pathway involved in flowering. We then examined whether the properties of the starch synthesis enzyme adenosine 5'-diphosphate-glucose pyrophosphorylase (AGPase) are important for adjustment of starch synthesis to daylength. Modulation of AGPase activity is known to bring about short-term adjustments of photosynthate partitioning between starch and sucrose (Suc) synthesis. We found that adjustment of starch synthesis to daylength was compromised in plants expressing a deregulated bacterial AGPase in place of the endogenous AGPase and in plants containing mutant forms of the endogenous AGPase with altered allosteric regulatory properties. We suggest that the rate of starch synthesis is in part determined by growth rate at the end of the preceding night. If growth at night is low, as in short days, there is a delay before growth recovers during the next day, leading to accumulation of Suc and stimulation of starch synthesis via activation of AGPase. If growth at night is fast, photosynthate is used for growth at the start of the day, Suc does not accumulate, and starch synthesis is not up-regulated.

The grapevine flagellin receptor VvFLS2 differentially recognizes flagellin-derived epitopes from the endophytic growth-promoting bacterium Burkholderia phytofirmans and plant pathogenic bacteria.

• The role of flagellin perception in the context of plant beneficial bacteria still remains unclear. Here, we characterized the flagellin sensing system flg22-FLAGELLIN SENSING 2 (FLS2) in grapevine, and analyzed the flagellin perception in the interaction with the endophytic plant growth-promoting rhizobacterium (PGPR) Burkholderia phytofirmans. • The functionality of the grapevine FLS2 receptor, VvFLS2, was demonstrated by complementation assays in the Arabidopsis thaliana fls2 mutant, which restored flg22-induced H2O2 production and growth inhibition. Using synthetic flg22 peptides from different bacterial origins, we compared recognition specificities between VvFLS2 and AtFLS2. • In grapevine, flg22-triggered immune responses are conserved and led to partial resistance against Botrytis cinerea. Unlike flg22 peptides derived from Pseudomonas aeruginosa or Xanthomonas campestris, flg22 peptide derived from B. phytofirmans triggered only a small oxidative burst, weak and transient defense gene induction and no growth inhibition in grapevine. Although, in Arabidopsis, all the flg22 epitopes exhibited similar biological activities, the expression of VvFLS2 into the fls2 background conferred differential flg22 responses characteristic for grapevine. • These results demonstrate that VvFLS2 differentially recognizes flg22 from different bacteria, and suggest that flagellin from the beneficial PGPR B. phytofirmans has evolved to evade this grapevine immune recognition system.

Burkholderia phytofirmans PsJN acclimates grapevine to cold by modulating carbohydrate metabolism.

Low temperatures damage many temperate crops, including grapevine, which, when exposed to chilling, can be affected by symptoms ranging from reduced yield up to complete infertility. We have previously demonstrated that Burkholderia phytofirmans PsJN, a plant growth-promoting rhizobacteria (PGPR) that colonizes grapevine, is able to reduce chilling-induced damage. We hypothesized that the induced tolerance may be explained at least partly by the impact of bacteria on grapevine photosynthesis or carbohydrate metabolism during cold acclimation. To investigate this hypothesis, we monitored herein the fluctuations of photosynthesis parameters (net photosynthesis [P(n)], intercellular CO(2) concentration, stomatal conductances, ΦPSII, and total chlorophyll concentration), starch, soluble sugars (glucose, fructose, saccharose, mannose, raffinose, and maltose), and their precursors during 5 days of chilling exposure (4°C) on grapevine plantlets. Bacterization affects photosynthesis in a non-stomatal dependent pattern and reduced long-term impact of chilling on P(n). Furthermore, all studied carbohydrates known to be involved in cold stress tolerance accumulate in non-chilled bacterized plantlets, although some of them remained more concentrated in the latter after chilling exposure. Overall, our results suggest that modification of carbohydrate metabolism in bacterized grapevine plantlets may be one of the major effects by which this PGPR reduces chilling-induced damage.