First report of Botrytis cinerea causing grey mold on Corydalis yanhusuo in Panan, China.

Yanhusuo (Corydalis yanhusuo (Y. H. Chou & Chun C. Hsu) W. T. Wang ex Z.Y. Su & C.Y. Wu), a perennial herbaceous plant of the Papaveraceae family and genus Corydalis, is also known as Yuanhu and used as medicine by its tuberous roots. It is mainly planted in Zhejiang, Jiangsu and Anhui provinces of China, with the best quality produced in Panan County of Zhejiang province. Yanhusuo has the effects of promoting blood circulation, invigorating the flow of qi and relieving pain, and is widely used in Chinese traditional medicines. In surveys carried out in summer of 2020-2023, grey mold disease was found occurred on C. yanhusuo in Panan County. This disease begins at April, and lasts to July, with incidence of 20% to 70%. The diseased plants showed a large number of gray mold layers adhere to the leaves. When the disease infects from the leaf tips, it form V-shaped lesions; when the leaves are severely infected, the entire leaves die, shrink, curl, and have a large number of gray mold layers on the surface. To identify the causal agent of this leaf disease, diseased leaves were collected from Yanhusuo field at Panan County of Zhejiang province in China since 2020, and tissues at the junction of the healthy and diseased areas were cut off, disinfected with 75% ethanol for 30 seconds, rinsed with sterile water for 3 times for 1 minute each time, air-dried under sterile conditions, and then were inoculated in PDA medium and cultured in a 25℃ incubator. After 2-3 days, picked the edge hyphae of the fungi that grew on the PDA plate and cultured them on a new PDA plate. After 5 days, picked the single spore and inoculated on a new PDA plate for continuous cultivation until pure culture strains were obtained. Thirty strains were isolated from 30 samples that collected from 3 Yanhusuo fields in Panan County. One of the thirty purified strains was named "YH8" for further identification. When cultured on PDA medium, mycelia were initially whitish and turned gray with age. The hyphae accumulate into clusters, and no sclerotia are produced during the cultivation. The conidiophores are slender, septate. The base of the conidiophore is enlarged or slightly enlarged. The conidiophore often has branches and produces a large number of conidia, which are similar to grape clusters. The conidia are monosporous, ovoid, and colorless, 6.08 µm-12.76 µm×8.42 µm-19.34 µm, with an average size of 9.55 µm×14.50 µm. To further identify the species, YH8 genomic DNA was extracted, and the internal transcribed spacer (ITS), heat shock protein (HSP60), and glyceraldehyde-3-phosphate dehydrogenase (G3PDH) genes were amplified with the primers ITS1/4 (White et al. 1990), HSP60-F/HSP60-R, and G3PDH-F/G3PDH-R (Staats et al. 2005), respectively. A multilocus phylogenetic tree was constructed with the ITS, HSP60, and G3PDH reference sequences, and the sequences of PCR amplicons (Genbank number: PP388281, PP376066 and PP376067) were 100% (518 bp out of 518 bp), 99% (994 bp out of 995 bp) and 100% (880 bp out of 880 bp) identical to the Botrytis cinerea strain 5-3, respectively, and the grouping of strain YH8 was supported by 99% bootstrap value. To fulfill the Koch's postulates, spore suspension (approximately 103 CFU/mL) of YH8 was sprayed onto leaves of 3-week Yanhusuo seedlings, and sterile water was sprayed as negative control, 15 seedlings for each treatment, and the experiments was repeated for times. The seedlings were incubated in a growth chamber under 28℃ and 80% humidity. Seven days after inoculation, leaves of noninoculated controls were green and healthy, while the seedlings inoculated with spore suspension of YH8 showed lesions and molds, which were same with field symptoms. The causal pathogen was then reisolated from the lesions, and the gained pathogen showed same colony and spore morphology with YH8, which suggested the confirmation of Koch's postulates. Based on the morphological characteristics and molecular identification, the strain YH8 was identified as B. cinerea. To our knowledge, this is the first report of B. cinerea causing gray mold on the Corydalis yanhusuo in China. This report will provide guide to growers and local technicians for diagnostic and controlling grey mold disease of Yanhusuo.

Characterization of wall-associated kinase/wall-associated kinase-like (WAK/WAKL) family in rose (Rosa chinensis) reveals the role of RcWAK4 in Botrytis resistance.

BACKGROUND: Wall-associated kinase (WAK)/WAK-like (WAKL) is one of the subfamily of receptor like kinases (RLK). Although previous studies reported that WAK/WAKL played an important role in plant cell elongation, response to biotic and abiotic stresses, there are no systematic studies on RcWAK/RcWAKL in rose. RESULTS: In this study, we identified a total of 68 RcWAK/RcWAKL gene family members within rose (Rosa chinensis) genome. The RcWAKs contained the extracellular galacturonan-binding domain and calcium-binding epidermal growth factor (EGF)-like domain, as well as an intracellular kinase domains. The RcWAKLs are missing either calcium-binding EGF-like domain or the galacturonan-binding domain in their extracellular region. The phylogenetic analysis showed the RcWAK/RcWAKL gene family has been divided into five groups, and these RcWAK/RcWAKL genes were unevenly distributed on the 7 chromosomes of rose. 12 of RcWAK/RcWAKL genes were significantly up-regulated by Botrytis cinerea-inoculated rose petals, where RcWAK4 was the most strongly expressed. Virus induced gene silencing of RcWAK4 increased the rose petal sensitivity to B. cinerea. The results indicated RcWAK4 is involved in the resistance of rose petal against B. cinerea. CONCLUSION: Our study provides useful information to further investigate the function of the RcWAK/RcWAKL gene family and breeding research for resistance to B. cinerea in rose.

Reduction of Botrytis cinerea Colonization of and Sporulation on Bunch Trash.

Botrytis bunch rot (BBR) of grapevine, caused by Botrytis cinerea, is commonly managed by fungicide (FUN) sprays at flowering (A), at prebunch closure (B), at veraison (C), and before harvest. Applications at A, B, and C are recommended to reduce B. cinerea colonization of bunch trash and the production of conidia during berry ripening. The effects of these applications were previously evaluated as reductions in BBR severity at harvest rather than as reductions in bunch trash colonization and sporulation by B. cinerea. This study investigated the effects of FUNs (a commercial mixture of fludioxonil and cyprodonil), biological control agents (BCAs; Aureobasium pullulans and Trichoderma atroviride), and botanicals (BOTs; a commercial mixture of eugenol, geraniol, and thymol) applied at different timings (A, B, C, or ABC) compared with a nontreated control (NT) on B. cinerea bunch trash colonization and sporulation in vineyards. The ability of B. cinerea to colonize the bunch trash (as indicated by B. cinerea DNA content) and sporulate (as indicated by the number of conidia produced under optimal laboratory conditions) was highly variable, and this variability was higher between years (2015 to 2018) than among the three vineyards and three sampling times (i.e., 1 week after applications at A, B, and C). B. cinerea sporulation on bunch trash was significantly lower in plots treated with FUN than in NT in only 3 of 18 cases (3 vineyards × 2 years × 3 sampling times). FUN applications, however, significantly reduced B. cinerea colonization of bunch trash compared with NT; for colonization, BCA efficacy was similar to that of FUN, but BOT efficacy was variable. For all products, colonization reduction was the same with application at A versus ABC, meaning that the effect of an early season application lasted from flowering to 1 week after veraison. These results indicate that the early season control of B. cinerea is important to reduce the saprophytic colonization of bunch trash, especially when the risk of BBR is high.

Genome-wide characterization of the rose (Rosa chinensis) WRKY family and role of RcWRKY41 in gray mold resistance.

BACKGROUND: The WRKYs are a major family of plant transcription factors that play roles in the responses to biotic and abiotic stresses; however, a comprehensive study of the WRKY family in roses (Rosa sp.) has not previously been performed. RESULTS: In the present study, we performed a genome-wide analysis of the WRKY genes in the rose (Rosa chinensis), including their phylogenetic relationships, gene structure, chromosomal locations, and collinearity. Using a phylogenetic analysis, we divided the 56 RcWRKY genes into three subgroups. The RcWRKYs were unevenly distributed across all seven rose chromosomes, and a study of their collinearity suggested that genome duplication may have played a major role in RcWRKY gene duplication. A Ka/Ks analysis indicated that they mainly underwent purifying selection. Botrytis cinerea infection induced the expression of 19 RcWRKYs, most of which had undergone gene duplication during evolution. These RcWRKYs may regulate rose resistance against B. cinerea. Based on our phylogenetic and expression analyses, RcWRKY41 was identified as a candidate regulatory gene in the response to B. cinerea infection, which was confirmed using virus-induced gene silencing. CONCLUSIONS: This study provides useful information to facilitate the further study of the function of the rose WRKY gene family.

Impact of foliar fungicides on target and non-target soil microbial communities in cucumber crops.

The application of foliar fungicides to horticultural crops has raised public concerns worldwide. In fact, it has been demonstrated that such fungicides have an impact on non-target microorganisms in the rhizosphere. Fluopyram, triadimenol and penthiopyrad are three broad-spectrum fungicides recommended to control foliar diseases. In our experiment, these fungicides were applied to a cucumber crop to mainly control downy mildew caused by Pseudoperonospora cubensis and grey mold caused by Botrytis cinerea. At the same time, we found that these treatments also controlled other fungal pathogens affecting cucumber crops, particularly penthiopyrad, which was more effective. Once the fungicide application period was over, the effect decreased, although fungicide traces remained in the soil. Furthermore, microbial soil community analysis indicated that both fungicide treatments affect fungal communities to a greater extent than bacterial communities.

A Conservative Mutant Version of the Mrr1 Transcription Factor Correlates with Reduced Sensitivity to Fludioxonil in Botrytis cinerea.

Fludioxonil is a highly effective phenylpyrrole fungicide for controlling Botrytis cinerea. Although the field efficacy of fludioxonil remains high, Botrytis cinerea isolates with reduced sensitivity have been reported globally. The molecular target of fludioxonil still remains unknown; however, a mechanism of reduced sensitivity to fludioxonil underlies the overexpression of the ATP binding cassette (ABC) transporter AtrB in a dependent pathway of the Mrr1 transcription factor. Fludioxonil is a key player in controlling B. cinerea infection in table grapes in Chile. However, some isolates with a reduced sensitivity to fludioxonil were detected. This study observed endogenous atrB overexpression in Chilean isolates with reduced sensitivity to fludioxonil (n = 22) compared to the sensitive isolates (n = 10). All isolates increased the expression of atrB in a growth medium supplemented with fludioxonil (0.05 µg/mL). However, sensitive isolates showed lower atrB expression than those with reduced fludioxonil sensitivity. Remarkably, a mutant version of the transcription factor Mrr1 carrying 21 amino acid modifications was identified in all isolates with reduced sensitivity to fludioxonil. These changes alter the protein's transcription factor domain and the C-terminal portion of the protein but not the Zn (2)-C6 fungal-type DNA-binding domain. These results suggest a direct relationship between the conserved and divergent mutant version of mrr1 and sensitivity to fludioxonil. This study provides a new target for developing molecular diagnostic strategies to monitor B. cinerea's sensitivity to fludioxonil in the field.

Pseudomonas synxantha volatile organic compounds: efficacy against Cadophora luteo-olivacea and Botrytis cinerea of kiwifruit.

Volatile organic compounds (VOCs) are responsible for the antagonistic activity exerted by different biological control agents (BCAs). In this study, VOCs produced by Pseudomonas synxantha strain 117-2b were tested against two kiwifruit fungal postharvest pathogens: Cadophora luteo-olivacea and Botrytis cinerea, through in vitro and in vivo assays. In vitro results demonstrated that P. synxantha 117-2b VOCs inhibit mycelial growth of C. luteo-olivacea and B. cinerea by 56% and 42.8% after 14 and 5 days of exposition, respectively. In vivo assay demonstrated significant inhibitory effects. VOCs used as a biofumigant treatment reduced skin-pitting symptoms disease severity by 28.5% and gray mold incidence by 66.6%, with respect to the untreated control. BCA volatiles were analyzed by solid-phase microextraction coupled with gas chromatography-mass spectrometry (SPME-GC/MS), and among the detected compounds, 1-butanol, 3-methyl and 1-nonene resulted as the most produced. Their efficacy as pure synthetic compounds was assayed against mycelial growth of fungal pathogens by different concentrations (0.34, 0.56, and 1.12 µL mL-1 headspace). The effect of the application of VOCs as a biofumigant was also investigated as the expression level of seven defense-related genes of kiwifruit at different exposition times. The results indicated an enhancement of the expression of almost all the genes starting from 3 h of treatment. These results described P. synxantha VOCs characteristics and their potential as a promising method to adopt for protecting kiwifruit from postharvest diseases caused by C. luteo-olivacea and B. cinerea.

Seasonal Dynamics and Damage of Halyomorpha halys in Italian Vineyards.

The brown marmorated stink bug, Halyomorpha halys (Stål) (Hemiptera: Pentatomidae), is an invasive polyphagous pest often observed in vineyards. In Europe, a gap needs to be filled in the knowledge on H. halys seasonal dynamics and damage on grapes. With this study, we described the seasonal dynamics of H. halys and its distribution in multi-cultivar vineyards, and we evaluated the damage on grape clusters induced by different pest densities. In vineyards, the seasonal occurrence of H. halys varied across time and grape cultivars, and the pest was more abundant on Cabernet Franc, Merlot and, to a lesser extent, Pinot gris. Moreover, higher densities of H. halys were found on red berry cultivars than on white ones, and on cultivars ripening late in the season. An edge effect was also detected in pest distribution within vineyards, with more stink bugs observed in the borders. In the study on pest infestation density, H. halys caused damage on berries, showing differences in susceptibility among different cultivars and with regard to the time of infestation (i.e., plant phenological stages). Halyomorpha halys infestation induced an increase in Botrytis cinerea and sour rot incidence, which probably represents the main issue related to the impact of brown marmorated stink bug on grapevine.

Curse or blessing: Growth- and laccase-modulating properties of polyphenols and their oxidized derivatives on Botrytis cinerea.

Infection of grapevines with the grey mold pathogen Botrytis cinerea results in severe problems for winemakers worldwide. Browning of wine is caused by the laccase-mediated oxidation of polyphenols. In the last decades, Botrytis management has become increasingly difficult due to the rising number of resistances and the genetic variety of Botrytis strains. During the search for sustainable fungicides, polyphenols showed great potential to inhibit fungal growth. The present study revealed two important aspects regarding the effects of grape-specific polyphenols and their polymerized oxidation products on Botrytis wild strains. On the one hand, laccase-mediated oxidized polyphenols, which resemble the products found in infected grapes, showed the same potential for inhibition of growth and laccase activity, but differed from their native forms. On the other hand, the impact of phenolic compounds on mycelial growth is not correlated to the effect on laccase activity. Instead, mycelial growth and relative specific laccase activity appear to be modulated independently. All phenolic compounds showed not only inhibitory but also inductive effects on fungal growth and/or laccase activity, an observation which is reported for the first time. The simultaneous inhibition of growth and laccase activity demonstrated may serve as a basis for the development of a natural botryticide. Yet, the results showed considerable differences between genetically distinguishable strains, impeding the use of a specific phenolic compound against the genetic variety of wild strains. The present findings might have important implications for future understanding of Botrytis cinerea infections and sustainable Botrytis management including the role of polyphenols.

Silene uniflora Extracts for Strawberry Postharvest Protection.

Halophytes are gaining considerable attention due to their applications in saline agriculture, phytoremediation, medicine, and secondary metabolite production. This study investigated the bioactive components present in Silene uniflora (sea campion) hydromethanolic extract, and their antimicrobial activity was evaluated both in vitro and ex situ against two strawberry phytopathogens, namely Botrytis cinerea (grey mold) and Colletotrichum nymphaeae (anthracnose fruit rot). The main identified phytochemicals were mome inositol, saturated fatty acid esters, and cyclotetracosane. In vitro tests demonstrated complete inhibition of the growth of B. cinerea and C. nymphaeae at extract concentrations of 1000 and 1500 µg·mL-1, respectively, with an activity comparable to that of fosetyl-Al and substantially higher than that of azoxystrobin. This activity was improved upon conjugation with chitosan oligomers (COS), yielding inhibition values of 750 and 1000 µg·mL-1. The COS-S. uniflora conjugate complexes were then tested as protective treatments for postharvest storage of strawberry fruit, resulting in high protection against artificially inoculated B. cinerea and C. nymphaeae at doses of 3750 and 5000 µg·mL-1, respectively. The reported results open the door to the valorization of this halophyte as a source of biorationals for strawberry protection.

Dipicolinic acid enhances kiwifruit resistance to Botrytis cinerea by promoting phenolics accumulation.

BACKGROUND: Kiwifruit is highly susceptible to fungal pathogens, such as Botrytis cinerea, which reduce crop production and quality. In this study, dipicolinic acid (DPA), which is one of the main components of Bacillus spores, was evaluated as a new elicitor to enhance kiwifruit resistance to B. cinerea. RESULTS: DPA enhances antioxidant capacity and induces the accumulation of phenolics in B. cinerea-infected 'Xuxiang' kiwifruit. The contents of the main antifungal phenolics in kiwifruit, including caffeic acid, chlorogenic acid and isoferulic acid, increased after DPA treatment. DPA enhanced H2 O2 levels after 0 and 1 days, which promoted catalase (CAT) and superoxide dismutase (SOD) activities, reducing long-term H2 O2 levels. DPA promoted the up-regulation of several kiwifruit defense genes, including CERK1, MPK3, PR1-1, PR1-2, PR5-1 and PR5-2. Furthermore, DPA at 5 mM inhibited B. cinerea symptoms in kiwifruit (95.1% lesion length inhibition) more effectively than the commercial fungicides carbendazim, difenoconazole, prochloraz and thiram. CONCLUSIONS: The antioxidant properties of DPA and the main antifungal phenolics of kiwifruit were examined for the first time. This study uncovers new insights regarding the potential mechanisms used by Bacillus species to induce disease resistance. © 2023 Society of Chemical Industry.

Defense response of strawberry plants against Botrytis cinerea influenced by coriander extract and essential oil.

Essential oils and extracts are investigated in sustainable plant protection area lately. Alternative antifungal substances are especially relevant for major economic-relevance pathogens, like Botrytis cinerea (causal agent of strawberry grey mold), control. However, the reaction of plants to alternative protection with plant-origin products is currently unknown. Induced stress in plants causes changes in antioxidant and photosynthetic systems. The aim of the research was to determine the defense response of strawberry plants under application of coriander seed products. In the first step of the research, we determined coriander seed (Coriandrum sativum), black seed (Nigella sativa) and peppermint leaf (Menta × piperita) products' antifungal activity against B. cinerea in vitro. Secondly, we continued evaluation of antifungal activity under controlled environment on strawberry plants of the most effective coriander seed products. Additionally, we evaluated the antioxidant and photosynthetic parameters in strawberries, to examine the response of plants. Antifungal activity on strawberries was determined based on grey mold incidence and severity after application of coriander products. Impact on photosynthetic system was examined measuring photosynthetic rate, transpiration rate, stomatal conductance, and intercellular to ambient CO2 concentration. Strawberry leaves were collected at the end of the experiment to analyze the antioxidant response. The highest antifungal activity both in vitro and on strawberries had coriander seed essential oil, which decreased grey mold severity. Coriander extract increased the photosynthetic capacity and antioxidant response of strawberry plants, however had negative effect on suppression of grey mold. In most cases, the essential oil activated antioxidant response of strawberry plants lower than extract. Our study results provide no direct impact of increased photosynthetic capacity values and antifungal effect after treatment with natural oils. The highest concentrations of coriander essential oil and extract potentially demonstrated a phytotoxic effect.

Trichoderma-Induced Resistance to Botrytis cinerea in Solanum Species: A Meta-Analysis.

With the idea of summarizing the outcomes of studies focusing on the resistance induced by Trichoderma spp. against Botrytis cinerea in tomato, the present paper shows, for the first time, results of a meta-analysis performed on studies published from 2010 to 2021 concerning the cross-talk occurring in the tomato-Trichoderma-B. cinerea system. Starting from an initial set of 40 papers, the analysis was performed on 15 works and included nine parameters, as a result of a stringent selection mainly based on the availability of more than one article including the same indicator. The resulting work not only emphasizes the beneficial effects of Trichoderma in the control of grey mold in tomato leaves (reduction in disease intensity, severity and incidence and modulation of resistance genes in the host), but carefully drives the readers to reply to two questions: (i) What are the overall effects of Trichoderma on B. cinerea infection in tomato? (ii) Do the main effects of Trichoderma differ based on the tomato species, Trichoderma species, amount, type and duration of treatment? At the same time, this meta-analysis highlights some weak points of the available literature and should be seen as an invitation to improve future works to better the conceptualization and measure.

High CO2 Reduces Spoilage Caused by Botrytis cinerea in Strawberry Without Impairing Fruit Quality.

High CO2 (> 20 kPa) conditions are beneficial for suppressing spoilage caused by Botrytis cinerea in strawberry fruit; however, these conditions are often accompanied by discoloration, off-flavors, and faster softening. Stepwise increments of CO2 concentrations have been proposed to alleviate injuries in fruits caused by high CO2. In this study, we investigated whether stepwise increments of CO2, up to 30 kPa and under a reduced O2 concentration, are beneficial for reducing fungal spoilage without inducing CO2 injury symptoms in strawberry fruit. Based on recommended settings (5-10 kPa O2 with 15-20 kPa CO2), we first selected optimal O2 and CO2 concentrations that best-reduced spoilage caused by B. cinerea in red ripe "Sonsation" strawberry fruit. We found that higher O2 (10 kPa) and CO2 (20 kPa) concentrations were most beneficial for prolonging strawberry fruit shelf life. Subsequently, we studied the performance of red ripe "Arabella" strawberry fruit stored at 5°C under different controlled atmosphere (CA) conditions (10 kPa O2 with either 0, 20, or 30 kPa CO2). The CO2 concentrations were achieved either within 8 h or in a stepwise manner within the first 4 days of storage. As a control, 21 kPa O2 and 0 kPa CO2 were used. Following storage for up to 11 days, the spoilage incidence was assessed at 12°C for 5 days. The application of high CO2 (20 and 30 kPa) combined with 10 kPa O2 greatly suppressed fruit spoilage during storage and subsequent shelf life. High CO2 suppressed respiration as well as maintained a higher pH and firmness in treated fruit. The level of total sugars did not change, but during storage, a substantial part of sucrose was converted into glucose and fructose, especially under high CO2 conditions. High CO2 did not affect ascorbic acid and anthocyanin levels. The stepwise increments of CO2 did not result in beneficial effects compared to the static application of high CO2. Our results show that "Arabella" strawberry fruit are highly tolerant to elevated CO2 and can be stored under 30 kPa CO2 to prolong the shelf life.

Discovery of a gene cluster for the biosynthesis of novel cyclic peptide compound, KK-1, in Curvularia clavata.

KK-1, a cyclic depsipeptide with 10 residues produced by a filamentous fungus Curvularia clavata BAUA-2787, is a promising pesticide active compound with high activity against many plant pathogens, especially Botrytis cinerea. As a first step toward the future mass production of KK-1 through synthetic biological approaches, we aimed to identify the genes responsible for the KK-1 biosynthesis. To achieve this, we conducted whole genome sequencing and transcriptome analysis of C. clavata BAUA-2787 to predict the KK-1 biosynthetic gene cluster. We then generated the overexpression and deletion mutants for each cluster gene using our originally developed transformation system for this fungus, and analyzed the KK-1 production and the cluster gene expression levels to confirm their involvement in KK-1 biosynthesis. As a result of these, a region of approximately 71 kb was found, containing 10 open reading frames, which were co-induced during KK-1 production, as a biosynthetic gene cluster. These include kk1B, which encodes nonribosomal peptide synthetase with a domain structure that is consistent with the structural features of KK-1, and kk1F, which encodes a transcription factor. The overexpression of kk1F increased the expression of the entire cluster genes and, consequently, improved KK-1 production, whereas its deletion decreased the expression of the entire cluster genes and almost eliminated KK-1 production, demonstrating that the protein encoded by kk1F regulates the expressions of the other nine cluster genes cooperatively as the pathway-specific transcription factor. Furthermore, the deletion of each cluster gene caused a reduction in KK-1 productivity, indicating that each gene is involved in KK-1 production. The genes kk1A, kk1D, kk1H, and kk1I, which showed a significant decrease in KK-1 productivity due to deletion, were presumed to be directly involved in KK-1 structure formation, including the biosynthesis of the constituent residues. kk1C, kk1E, kk1G, and kk1J, which maintained a certain level of KK-1 productivity despite deletion, were possibly involved in promoting or assisting KK-1 production, such as extracellular transportation and the removal of aberrant units incorporated into the peptide chain.

Fatty acid modulation and desaturase gene expression are differentially triggered in grapevine incompatible interaction with biotrophs and necrotrophs.

Grapevine (Vitis vinifera L.) is prone to fungal and oomycete diseases. Downy and powdery mildews and grey mold, are caused by Plasmopara viticola, Erisiphe necator and Botrytis cinerea, respectively. P. viticola and E. necator are obligatory biotrophs whereas B. cinerea is a necrotroph. In tolerant grapevine cultivars, plant-pathogen interaction induces defence responses, including metabolite and protein accumulation and hypersensitive reaction. Lipid and lipid-derived molecules may have a key role in the activation of defence mechanisms. Previous results suggest that V. vinifera cv Regent tolerance to P. viticola may be mediated in the first hours post inoculation by fatty acid (FA) associated signalling. In the present study we characterized FA modulation in V. vinifera cv Regent leaves upon inoculation with P. viticola, E. necator and B. cinerea and correlated FA modulation with the expression profiles of genes encoding the FA desaturases FAD6 and FAD8. In all the interactions, a progressive desaturation of stearic acid to α-linolenic acid, precursor of jasmonic acid, occurred, which was observed for a longer period against B. cinerea. Our results provide evidence of a distinct FA meditated signalling pattern in grapevine interaction with biotrophs and necrotrophs. While the interaction with the biotrophs may trigger a higher synthesis of polyunsaturated FA (PUFA) at early time-points with a tendency to return to basal levels, the interaction with B. cinerea may trigger a later and more durable induction of PUFA synthesis. In all interactions, membrane fluidity modulation occurred, which may be crucial to maintain cellular function during infection.

Genome sequence analysis of the beneficial Bacillus subtilis PTA-271 isolated from a Vitis vinifera (cv. Chardonnay) rhizospheric soil: assets for sustainable biocontrol.

BACKGROUND: Bacillus subtilis strains have been widely studied for their numerous benefits in agriculture, including viticulture. Providing several assets, B. subtilis spp. are described as promising plant-protectors against many pathogens and as influencers to adaptations in a changing environment. This study reports the draft genome sequence of the beneficial Bacillus subtilis PTA-271, isolated from the rhizospheric soil of healthy Vitis vinifera cv. Chardonnay at Champagne Region in France, attempting to draw outlines of its full biocontrol capacity. RESULTS: The PTA-271 genome has a size of 4,001,755 bp, with 43.78% of G + C content and 3945 protein coding genes. The draft genome of PTA-271 putatively highlights a functional swarming motility system hypothesizing a colonizing capacity and a strong interacting capacity, strong survival capacities and a set of genes encoding for bioactive substances. Predicted bioactive compounds are known to: stimulate plant growth or defenses such as hormones and elicitors, influence beneficial microbiota, and counteract pathogen aggressiveness such as effectors and many kinds of detoxifying enzymes. CONCLUSIONS: Plurality of the putatively encoded biomolecules by Bacillus subtilis PTA-271 genome suggests environmentally robust biocontrol potential of PTA-271, protecting plants against a broad spectrum of pathogens.

Timing of light quality affects susceptibility to Botrytis cinerea in strawberry leaves.

Light plays an essential role in the outcome of plant-pathogen interactions. In this study, the effect of pre- and post-inoculation light quality on Botrytis cinerea disease development in strawberry leaves was investigated. Circadian variations in leaf susceptibility to Botrytis cinerea were tested by inoculating at 5 h, 12 h, or 16 h after the start of photoperiod. The effects of 4 h and/or 12 h of red or blue light combined with white light in a 16 h photoperiod on leaf susceptibility and metabolites were studied. Light presence significantly decreased leaf susceptibility to Botrytis irrespective of light quality. Leaf susceptibility to B. cinerea showed significant circadian variations. Susceptibility was highest in leaves inoculated 12 h after the start of photoperiod followed by incubation in the dark. When disease development took place under the light regime, the shorter the light period after inoculation the stronger the disease symptoms. Twelve hours of red light radiation in a 16 h photoperiod increased, while 12 h blue light radiation decreased leaf resistance, resulting in alterations in the circadian variation of leaf susceptibility. These alterations correlated with changes in leaf metabolites, such as hydrogen peroxide, total phenolics and hexoses. Overall, we conclude that twelve hours of red light followed by white light in a 16 h photoperiod efficiently increased leaf resistance to B. cinerea.

The Xylanase Inhibitor TAXI-I Increases Plant Resistance to Botrytis cinerea by Inhibiting the BcXyn11a Xylanase Necrotizing Activity.

During host plant infection, pathogens produce a wide array of cell wall degrading enzymes (CWDEs) to break the plant cell wall. Among CWDEs, xylanases are key enzymes in the degradation of xylan, the main component of hemicellulose. Targeted deletion experiments support the direct involvement of the xylanase BcXyn11a in the pathogenesis of Botrytis cinerea. Since the Triticum aestivum xylanase inhibitor-I (TAXI-I) has been shown to inhibit BcXyn11a, we verified if TAXI-I could be exploited to counteract B. cinerea infections. With this aim, we first produced Nicotiana tabacum plants transiently expressing TAXI-I, observing increased resistance to B. cinerea. Subsequently, we transformed Arabidopsis thaliana to express TAXI-I constitutively, and we obtained three transgenic lines exhibiting a variable amount of TAXI-I. The line with the higher level of TAXI-I showed increased resistance to B. cinerea and the absence of necrotic lesions when infiltrated with BcXyn11a. Finally, in a droplet application experiment on wild-type Arabidopsis leaves, TAXI-I prevented the necrotizing activity of BcXyn11a. These results would confirm that the contribution of BcXyn11a to virulence is due to its necrotizing rather than enzymatic activity. In conclusion, our experiments highlight the ability of the TAXI-I xylanase inhibitor to counteract B. cinerea infection presumably by preventing the necrotizing activity of BcXyn11a.

VvSWEET7 Is a Mono- and Disaccharide Transporter Up-Regulated in Response to Botrytis cinerea Infection in Grape Berries.

The newly-identified SWEETs are high-capacity, low-affinity sugar transporters with important roles in numerous physiological mechanisms where sugar efflux is critical. SWEETs are desirable targets for manipulation by pathogens and their expression may be transcriptionally reprogrammed during infection. So far, few plant SWEET transporters have been functionally characterized, especially in grapevine. In this study, in the Botrytis-susceptible variety "Trincadeira," we thoroughly analyzed modifications in the gene expression profile of key SWEET genes in Botrytis cinerea-infected grape berries. VvSWEET7 and VvSWEET15 are likely to play an important role during fruit development and Botrytis infection as they are strongly expressed at the green and mature stage, respectively, and were clearly up-regulated in response to infection. Also, B. cinerea infection down-regulated VvSWEET17a expression at the green stage, VvSWEET10 and VvSWEET17d expression at the veraison stage, and VvSWEET11 expression at the mature stage. VvSWEET7 was functionally characterized by heterologous expression in Saccharomyces cerevisiae as a low-affinity, high-capacity glucose and sucrose transporter with a K m of 15.42 mM for glucose and a K m of 40.08 mM for sucrose. VvSWEET7-GFP and VvSWEET15-GFP fusion proteins were transiently expressed in Nicotiana benthamiana epidermal cells and confocal microscopy allowed to observe that both proteins clearly localize to the plasma membrane. In sum, VvSWEETs transporters are important players in sugar mobilization during grape berry development and their expression is transcriptionally reprogrammed in response to Botrytis infection.