Antifungal Activity of Phloroglucinol Derivatives against Botrytis cinerea and Monilinia fructicola.

Naturally derived compounds show promise as treatments for microbial infections. Polyphenols, abundantly found in various plants, fruits, and vegetables, are noted for their physiological benefits including antimicrobial effects. This study introduced a new set of acylated phloroglucinol derivatives, synthesized and tested for their antifungal activity in vitro against seven different pathogenic fungi. The standout compound, 3-methyl-1-(2,4,6-trihydroxyphenyl) butan-1-one (2b), exhibited remarkable fungicidal strength, with EC50 values of 1.39 µg/mL against Botrytis cinerea and 1.18 µg/mL against Monilinia fructicola, outperforming previously screened phenolic compounds. When tested in vivo, 2b demonstrated effective antifungal properties, with cure rates of 76.26% for brown rot and 83.35% for gray mold at a concentration of 200 µg/mL, rivaling the commercial fungicide Pyrimethanil in its efficacy against B. cinerea. Preliminary research suggests that 2b's antifungal mechanism may involve the disruption of spore germination, damage to the fungal cell membrane, and leakage of cellular contents. These results indicate that compound 2b has excellent fungicidal properties against B. cinerea and holds potential as a treatment for gray mold.

Methionine represses gray mold of tomato by keeping nitric oxide at an appropriate level via ethylene synthesis and signal transduction.

Methionine (Met) can inhibit plant diseases caused by phytopathogens. However, the effect of Met on gray mold resulted from Botrytis cinerea in tomato is still unclear. This study showed 5 mM Met alleviated disease development of gray mold, enhanced chitinase (CHI) and ß-1, 3-glucanase (GNS) activities and the expression of SlCHI, SlGNS, SlPR1 and SlNPR1 in tomatoes, rather than inhibited the growth of B. cinerea directly. Moreover, ethylene biosynthesis and signal transduction before pathogen inoculating were induced by 5 mM Met. Interestingly, Met reduced the nitrosylation levels of ACS4 and ACO6, enhanced the activities of nitric oxide synthase, nitrite reductase (NR) and S-nitrosoglutathione reductase (GSNOR) and the expression of SlNR and SlGSNOR. Tomatoes treated with aminoethoxyvinylglycine and carboxy-PTIO exhibited lower resistance to B. cinerea. These results indicate 5 mM Met promoted ethylene biosynthesis and signal transduction to facilitate NO synthesis and metabolism, enhancing the resistance of tomatoes to B. cinerea.

Novel pesticides design: co-encapsulation of citral and cyproconazole for the control of Botrytis cinerea using biocompatible nano-carriers.

BACKGROUND: Growing concerns about sustainability have driven the search for eco-friendly pest management solutions. Combining natural and synthetic compounds within controlled release systems is a promising strategy. This study investigated the co-encapsulation of the natural compound citral (Cit) and the synthetic antifungal cyproconazole (CPZ) using two water-based nanocarriers: solid lipid nanoparticles (SLNs) and chitosan nanoparticles (CSNPs). RESULTS: Both CSNPs and SLNs loaded with Cit + CPZ displayed superior antifungal activity against Botrytis cinerea compared to free compounds. Notably, CSNPs with a 2:1 Cit:CPZ ratio exhibited the highest efficacy, achieving a minimum inhibitory concentration (MIC100) of < 1.56 µg mL-1, lower than the 12.5 µg mL-1 of non-encapsulated compounds. This formulation significantly reduced the required synthetic CPZ while maintaining efficacy, highlighting its potential for environmentally friendly pest control. CONCLUSION: The successful co-encapsulation of Cit + CPZ within CSNPs, particularly at a 2:1 ratio, demonstrates a promising approach for developing effective and sustainable antifungal formulations against B. cinerea. © 2024 Society of Chemical Industry.

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.

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.

The binding pattern of ferric iron and iron-binding protein in Botrytis cinerea.

Iron-binding protein (Ibp) has protective effect on pathogen exposed to H2O2 in defense response of plants. Ibp in Botrytis cinerea (BcIbp) is related to its virulence. Bcibp mutation lead to virulence deficiencies in B. cinerea. BcIbp is involved in the Fe3+ homeostasis regulation. Recognition the binding site and binding pattern of ferric iron and iron-binding protein in B. cinerea are vital to understand its function. In this study, molecular dynamics (MD) simulations, gaussian accelerated molecular dynamics (GaMD) simulations, dynamic cross correlation analysis and quantum chemical energy calculation were used to explore binding pattern of ferric iron. MD results showed that the C-terminal region had little effect on the stability of residues in the Fe3+-binding pocket. Energy calculations suggested the most likely coordination pattern for ferric iron in iron-binding protein. These results will help to understand the binding of ferric iron to iron-binding protein and provide new ideas for regulating the virulence of B. cinerea.

The mechanistic insights of essential oil of Mentha piperita to control Botrytis cinerea and the prospection of lipid nanoparticles to its application.

Botrytis cinerea is the phytopathogenic fungus responsible for the gray mold disease that affects crops worldwide. Essential oils (EOs) have emerged as a sustainable tool to reduce the adverse impact of synthetic fungicides. Nevertheless, the scarce information about the physiological mechanism action and the limitations to applying EOs has restricted its use. This study focused on elucidating the physiological action mechanisms and prospection of lipid nanoparticles to apply EO of Mentha piperita. The results showed that the EO of M. piperita at 500, 700, and 900 µL L-1 inhibited the mycelial growth at 100 %. The inhibition of spore germination of B. cinerea reached 31.43 % at 900 µL L-1. The EO of M. piperita decreased the dry weight and increased pH, electrical conductivity, and cellular material absorbing OD260 nm of cultures of B. cinerea. The fluorescence technique revealed that EO reduced hyphae width, mitochondrial activity, and viability, and increased ROS production. The formulation of EO of M. piperita loaded- solid lipid nanoparticles (SLN) at 500, 700, and 900 µL L-1 had particle size ∼ 200 nm, polydispersity index < 0.2, and stability. Also, the thermogravimetric analysis indicated that the EO of M. piperita-loaded SLN has great thermal stability at 50 °C. EO of M. piperita-loaded SLN reduced the mycelial growth of B. cinerea by 70 %, while SLN formulation (without EO) reached 42 % inhibition. These results supported that EO of M. piperita-loaded SLN is a sustainable tool for reducing the disease produced by B. cinerea.

Study of phenoxy radical couplings using the enzymatic secretome of Botrytis cinerea.

Phenoxy radical coupling reactions are widely used in nature for the synthesis of complex molecules such as lignin. Their use in the laboratory has great potential for the production of high value compounds from the polyphenol family. While the enzymes responsible for the generation of the radicals are well known, the behavior of the latter is still enigmatic and difficult to control in a reaction flask. Previous work in our laboratory using the enzymatic secretome of B. cinerea containing laccases has shown that incubation of stilbenes leads to dimers, while incubation of phenylpropanoids leads to dimers as well as larger coupling products. Building on these previous studies, this paper investigates the role of different structural features in phenoxy radical couplings. We first demonstrate that the presence of an exocyclic conjugated double bond plays a role in the generation of efficient reactions. In addition, we show that the formation of phenylpropanoid trimers and tetramers can proceed via a decarboxylation reaction that regenerates this reactive moiety. Lastly, this study investigates the reactivity of other phenolic compounds: stilbene dimers, a dihydro-stilbene, a 4-O-methyl-stilbene and a simple phenol with the enzymatic secretome of B. cinerea. The observed efficient dimerization reactions consistently correlate with the presence of a para-phenol conjugated to an exocyclic double bond. The absence of this structural feature leads to variable results, with some compounds showing low conversion or no reaction at all. This research has allowed the development of a controlled method for the synthesis of specific dimers and tetramers of phenylpropanoid derivatives and novel stilbene derivatives, as well as an understanding of features that can promote efficient radical coupling reactions.

Characterization of two novel fusariviruses co-infecting a single isolate of phytopathogenic fungus Botrytis cinerea.

A wide diversity of mycoviruses has been reported from Botrytis species, some with the potential to suppress the pathogenic abilities of this fungus. Considering their importance, this study was devised to find potential hypovirulence-associated mycoviruses found in Botrytis cinerea strains isolated from Pakistani strawberry fields. Here we report the complete genome characterization of two fusariviruses co-infecting a single isolate of phytopathogenic fungus B. cinerea (Kst14a). The viral genomes were sequenced by deep sequencing using total RNA fractions of the Kst14a isolate. The identified viruses were tentatively named Botrytis cinerea fusarivirus 9 (BcFV9) and Botrytis cinerea fusarivirus 3a (BcFV3a). Both viruses had a single-segmented (ssRNA) genome having a size of 6424 and 8370 nucleotides encoding two discontinuous open reading frames (ORFs). ORF-1 of both mycoviruses encodes for a polyprotein having a conserved domain of RNA-dependent RNA polymerase (RdRP) and a helicase domain (Hel) which function in RNA replication, while ORF2 encodes a hypothetical protein with an unknown function, respectively. Phylogenetic analysis indicated that BcFV9 made a clade with the genus Alphafusarivirus and BcFV3a fall in the genus Betafusarivirus in the family Fusariviridae. To our knowledge, this is the first report of two fusariviruses identified in isolates of B. cinerea from Pakistan. Both mycoviruses successfully transfected to a compatible strain of B. cinerea (Mst11). A comparison of virus-free (VF) and virus-infected (VI) isogenic lines showed the presence of these viruses was causing hypovirulence in infected strains. Virus-infected strains also had a small lesion size while testing the pathogenicity via apple assay.

Photodynamic Inactivation in agriculture: combating fungal phytopathogens resistant to conventional treatment.

Botrytis cinerea is a severe threat in agriculture, as it can infect over 200 different crop species with gray mold affecting food yields and quality. The conventional treatment using fungicides lead to emerging resistance over the past decades. Here, we introduce Photodynamic Inactivation (PDI) as a strategy to combat B. cinerea infections, independent of fungicide resistance. PDI uses photoactive compounds, which upon illumination create reactive oxygen species toxic for killing target organisms. This study focuses on different formulations of sodium-magnesium-chlorophyllin (Chl, food additive E140) as photoactive compound in combination with EDTA disodium salt dihydrate (Na2EDTA) as cell-wall permeabilizer and a surfactant. In an in vitro experiment, three different photosensitizers (PS) with varying Chl and Na2EDTA concentrations were tested against five B. cinerea strains with different resistance mechanisms. We showed that all B. cinerea mycelial spheres of all tested strains were eradicated with concentrations as low as 224 µM Chl and 3.076 mM Na2EDTA (LED illumination with main wavelength of 395 nm, radiant exposure 106 J cm-2). To further test PDI as a Botrytis treatment strategy in agriculture a greenhouse trial was performed on B. cinerea infected bell pepper plants (Capsicum annum L). Two different rates (560 or 1120 g Ha-1) of PS formulation (0.204 M Chl and 1.279 M Na2EDTA) and a combination of PS formulation with 0.05% of the surfactant BRIJ L4 (560 g Ha-1) were applied weekly for 4 weeks by spray application. Foliar lesions, percentage of leaves affected, percentage of leaf area diseased and AUDPC were significantly reduced, while percentage of marketable plants were increased by all treatments compared to a water treated control, however, did not statistically differ from each other. No phytotoxicity was observed in any treatment. These results add to the proposition of employing PDI with the naturally sourced PS Chl in agricultural settings aimed at controlling B. cinerea disease. This approach seems to be effective regardless of the evolving resistance mechanisms observed in response to conventional antifungal treatments.

Deciphering molecular events behind Systemin-induced resistance to Botrytis cinerea in tomato plants.

Plant defence peptides are paramount endogenous danger signals secreted after a challenge, intensifying the plant immune response. The peptidic hormone Systemin (Sys) was shown to participate in resistance in several plant pathosystems, although the mechanisms behind Sys-induced resistance when exogenously applied remain elusive. We performed proteomic, metabolomic, and enzymatic studies to decipher the Sys-induced changes in tomato plants in either the absence or the presence of Botrytis cinerea infection. Sys treatments triggered direct proteomic rearrangement mostly involved in carbon metabolism and photosynthesis. However, the final induction of defence proteins required concurrent challenge, triggering priming of pathogen-targeted proteins. Conversely, at the metabolomic level, Sys-treated plants showed an alternative behaviour following a general priming profile. Of the primed metabolites, the flavonoids rutin and isorhamnetin and two alkaloids correlated with the proteins 4-coumarate-CoA-ligase and chalcone-flavanone-isomerase triggered by Sys treatment. In addition, proteomic and enzymatic analyses revealed that Sys conditioned the primary metabolism towards the production of available sugars that could be fuelling the priming of callose deposition in Sys-treated plants; furthermore, PR1 appeared as a key element in Sys-induced resistance. Collectively, the direct induction of proteins and priming of specific secondary metabolites in Sys-treated plants indicated that post-translational protein regulation is an additional component of priming against necrotrophic fungi.

Modes of action and potential as a peptide-based biofungicide of a plant defensin MtDef4.

Due to rapidly emerging resistance to single-site fungicides in fungal pathogens of plants, there is a burgeoning need for safe and multisite fungicides. Plant antifungal peptides with multisite modes of action (MoA) have potential as bioinspired fungicides. Medicago truncatula defensin MtDef4 was previously reported to exhibit potent antifungal activity against fungal pathogens. Its MoA involves plasma membrane disruption and binding to intracellular targets. However, specific biochemical processes inhibited by this defensin and causing cell death have not been determined. Here, we show that MtDef4 exhibited potent antifungal activity against Botrytis cinerea. It induced severe plasma membrane and organelle irregularities in the germlings of this pathogen. It bound to fungal ribosomes and inhibited protein translation in vitro. A MtDef4 variant lacking antifungal activity exhibited greatly reduced protein translation inhibitory activity. A cation-tolerant MtDef4 variant was generated that bound to ß-glucan of the fungal cell wall with higher affinity than MtDef4. It also conferred a greater reduction in the grey mould disease symptoms than MtDef4 when applied exogenously on Nicotiana benthamiana plants, tomato fruits and rose petals. Our findings revealed inhibition of protein synthesis as a likely target of MtDef4 and the potential of its cation-tolerant variant as a peptide-based fungicide.

Ethylene-MPK8-ERF.C1-PR module confers resistance against Botrytis cinerea in tomato fruit without compromising ripening.

The plant hormone ethylene plays a critical role in fruit defense against Botrytis cinerea attack, but the underlying mechanisms remain poorly understood. Here, we showed that ethylene response factor SlERF.C1 acts as a key regulator to trigger the ethylene-mediated defense against B. cinerea in tomato fruits without compromising ripening. Knockout of SlERF.C1 increased fruit susceptibility to B. cinerea with no effect on ripening process, while overexpression enhanced resistance. RNA-Seq, transactivation assays, EMSA and ChIP-qPCR results indicated that SlERF.C1 activated the transcription of PR genes by binding to their promoters. Moreover, SlERF.C1 interacted with the mitogen-activated protein kinase SlMPK8 which allowed SlMPK8 to phosphorylate SlERF.C1 at the Ser174 residue and increases its transcriptional activity. Knocking out of SlMPK8 increased fruit susceptibility to B. cinerea, whereas overexpression enhanced resistance without affecting ripening. Furthermore, genetic crosses between SlMPK8-KO and SlERF.C1-OE lines reduced the resistance to B. cinerea attack in SlERF.C1-OE fruits. In addition, B. cinerea infection induced ethylene production which in turn triggered SlMPK8 transcription and enhanced the phosphorylation of SlERF.C1. Overall, our findings reveal the regulatory mechanism of the 'Ethylene-MPK8-ERF.C1-PR' module in resistance against B. cinerea and provide new insight into the manipulation of gray mold disease in fruits.

A new point mutation (D1158N) in histidine kinase Bos1 confers high-level resistance to fludioxonil in field gray mold disease.

Gray mold, caused by the fungus Botrytis cinerea, is one of the most important plant diseases worldwide that is prone to developing resistance to fungicides. Currently, the phenylpyrrole fungicide fludioxonil exhibits excellent efficacy in the control of gray mold in China. In this study, we detected the fludioxonil resistance of gray mold disease in Shouguang City of Shandong Province, where we first found fludioxonil-resistant isolates of B. cinerea in 2014. A total of 87 single spore isolates of B. cinerea were obtained from cucumbers in greenhouse, and 3 of which could grow on PDA plates amended with 50 µg/mL fludioxonil that was defined as high-level resistance, with a resistance frequency of 3.4%. Furthermore, the 3 fludioxonil-resistant isolates also showed high-level resistance to the dicarboximide fungicides iprodione and procymidone. Sequencing comparison revealed that all the 3 fludioxonil-resistant isolates had a point mutation at codon 1158, GAC (Asp) â†’ AAC (Asn) in the histidine kinase Bos1, which was proved to be the reason for fludioxonil resistance. In addition, the fludioxonil-resistant isolates possessed an impaired biological fitness compared to the sensitive isolates based on the results of mycelial growth, conidiation, virulence, and osmotic stress tolerance determination. Taken together, our results indicate that the high-level resistance to fludioxonil caused by the Bos1 point mutation (D1158N) has emerged in the field gray mold disease, and the resistance risk is relatively high, and fludioxonil should be used sparingly.

RAS signalling genes can be used as host-induced gene silencing targets to control fungal diseases caused by Sclerotinia sclerotiorum and Botrytis cinerea.

Sclerotinia sclerotiorum causes white mold (also called stem rot, Sclerotinia blight, etc.) in many economically important plants. It is a notorious soilborne fungal pathogen due to its wide host range and ability to survive in soil for long periods of time as sclerotia. Although host-induced gene silencing (HIGS) was recently demonstrated to be an effective method for controlling white mold, limited gene targets are available. Here, using a forward genetics approach, we identified a RAS-GTPase activating protein, SsGAP1, which plays essential roles in sclerotia formation, compound appressoria production and virulence. In parallel, as revealed by our knockout analysis, the SsGAP1 ortholog in Botrytis cinerea, BcGAP1, plays similar roles in fungal development and virulence. By knocking down SsRAS1 and SsRAS2, we also revealed that both SsRAS1 and SsRAS2 are required for vegetative growth, sclerotia development, compound appressoria production and virulence in S. sclerotiorum. Due to the major roles these RAS signalling components play in Sclerotiniaceae biology, they can be used as HIGS targets to control diseases caused by both S. sclerotiorum and B. cinerea. Indeed, when we introduced HIGS constructs targeting SsGAP1, SsRAS1 and SsRAS2 in Nicotiana benthamiana and Arabidopsis thaliana, we observed reduced virulence. Taken together, our forward genetics gene discovery pipeline in S. sclerotiorum is highly effective in identifying novel HIGS targets to control S. sclerotiorum and B. cinerea.

The apocarotenoid ß-ionone regulates the transcriptome of Arabidopsis thaliana and increases its resistance against Botrytis cinerea.

Carotenoids are isoprenoid pigments indispensable for photosynthesis. Moreover, they are the precursor of apocarotenoids, which include the phytohormones abscisic acid (ABA) and strigolactones (SLs) as well as retrograde signaling molecules and growth regulators, such as ß-cyclocitral and zaxinone. Here, we show that the application of the volatile apocarotenoid ß-ionone (ß-I) to Arabidopsis plants at micromolar concentrations caused a global reprogramming of gene expression, affecting thousands of transcripts involved in stress tolerance, growth, hormone metabolism, pathogen defense, and photosynthesis. This transcriptional reprogramming changes, along with induced changes in the level of the phytohormones ABA, jasmonic acid, and salicylic acid, led to enhanced Arabidopsis resistance to the widespread necrotrophic fungus Botrytis cinerea (B.c.) that causes the gray mold disease in many crop species and spoilage of harvested fruits. Pre-treatment of tobacco and tomato plants with ß-I followed by inoculation with B.c. confirmed the effect of ß-I in increasing the resistance to this pathogen in crop plants. Moreover, we observed reduced susceptibility to B.c. in fruits of transgenic tomato plants overexpressing LYCOPENE ß-CYCLASE, which contains elevated levels of endogenous ß-I, providing a further evidence for its effect on B.c. infestation. Our work unraveled ß-I as a further carotenoid-derived regulatory metabolite and indicates the possibility of establishing this natural volatile as an environmentally friendly bio-fungicide to control B.c.

Inhibition activity of tomato endophyte Bacillus velezensis FQ-G3 against postharvest Botrytis cinerea.

Grey mold, caused by Botrytis cinerea, is a widespread and harmful disease of tomato. Biocontrol agents derived from endophytic bacteria are known to hold great potential for inhibition of phytopathogen. We conducted this study to explore the tomato endophytic strains with inhibition activity against B. cinerea. Endophytic strain Bacillus velezensis FQ-G3 exhibited excellent inhibition activity against B. cinerea. Inhibitory effects against B. cinerea were investigated both in vitro and in vivo. The in vitro assays displayed that FQ-G3 could significantly inhibit mycelia growth with inhibition rate of 85.93%, and delay conidia germination of B. cinerea. Tomato fruit inoculated with B. velezensis FQ-G3 revealed lower grey mold during treatment. The antifungal activity was attributed to activation of defense-related enzymes, as evidenced by the higher levels of peroxidase, polyphenol oxidase, and phenylalanine ammonia lyase in tomatoes after inoculation. In addition, scanning electron microscope was applied to elucidate the interaction between endophytes and pathogen, and bacterial colonization and antibiosis appeared to be the underlying mechanisms that FQ-G3 could suppress growth of B. cinerea. Collectively, our present results suggested that FQ-G3 may potentially be useful as a biocontrol agent in postharvest tomatoes.

NAC61 regulates late- and post-ripening osmotic, oxidative, and biotic stress responses in grapevine.

During late- and post-ripening stages, grape berry undergoes profound biochemical and physiological changes whose molecular control is poorly understood. Here, we report the role of NAC61, a grapevine NAC transcription factor, in regulating different processes involved in berry ripening progression. NAC61 is highly expressed during post-harvest berry dehydration and its expression pattern is closely related to sugar concentration. The ectopic expression of NAC61 in Nicotiana benthamiana leaves resulted in low stomatal conductance, high leaf temperature, tissue collapse and a higher relative water content. Transcriptome analysis of grapevine leaves transiently overexpressing NAC61 and DNA affinity purification and sequencing analyses allowed us to narrow down a list of NAC61-regulated genes. Direct regulation of the stilbene synthase regulator MYB14, the osmotic stress-related gene DHN1b, the Botrytis cinerea susceptibility gene WRKY52, and NAC61 itself was validated. We also demonstrate that NAC61 interacts with NAC60, a proposed master regulator of grapevine organ maturation, in the activation of MYB14 and NAC61 expression. Overall, our findings establish NAC61 as a key player in a regulatory network that governs stilbenoid metabolism and osmotic, oxidative, and biotic stress responses that are the hallmark of late- and post-ripening grape stages.

Arabidopsis thaliana phosphoinositide-specific phospholipase C 2 is required for Botrytis cinerea proliferation.

Phospholipase C (PLC) plays a key role in lipid signaling during plant development and stress responses. PLC activation is one of the earliest responses during pathogen perception. Arabidopsis thaliana contains seven PLC encoding genes (AtPLC1 to AtPLC7) and two pseudogenes (AtPLC8 and AtPLC9), being AtPLC2 the most abundant isoform with constitutive expression in all plant organs. PLC has been linked to plant defense signaling, in particular to the production of reactive oxygen species (ROS). Previously, we demonstrated that AtPLC2 is involved in ROS production via the NADPH oxidase isoforms RBOHD activation during stomata plant immunity. Here we studied the role of AtPLC2 on plant resistance against the necrotrophic fungus Botrytis cinerea, a broad host-range and serious agricultural pathogen. We show that the AtPLC2-silenced (amiR PLC2) or null mutant (plc2-1) plants developed smaller B. cinerea lesions. Moreover, plc2-1 showed less ROS production and an intensified SA-dependent signaling upon infection, indicating that B. cinerea uses AtPLC2-triggered responses for a successful proliferation. Therefore, AtPLC2 is a susceptibility (S) gene that facilitates B. cinerea infection and proliferation.

Alleviative effects of quercetin of Botrytis cinerea-induced toxicity in zebrafish (Danio rerio) larvae.

Quercetin is a kind of flavonoid substance extensively existing in the plant, which has antioxidant, anti-inflammatory, and anti-apoptosis effects. It was reported that the higher concentration of spores present in the environment could cause abnormal development in zebrafish larvae. Therefore, this study set out to investigate whether quercetin could reduce the zebrafish larvae damage caused by Botrytis cinerea exposure as well as to examine the molecular basis for this action. The findings demonstrated that 50 µM quercetin improved the developmental dysplasia of zebrafish larvae induced by 102 CFU/mL Botrytis cinerea spore suspension, reduced abnormal apoptosis, enhanced antioxidant system, relieved inflammation, reshaped intestinal morphology and recovered intestinal motility. At the molecular level, quercetin decreased the transcriptional abundance of pro-apoptotic factors (bax, p53, caspase3, and caspase9) and up-regulated the anti-apoptotic gene (bcl-2) expression to reduce apoptosis. Moreover, quercetin enhanced the activities of downstream antioxidant enzymes (SOD and CAT) to clear excess ROS and MDA due to Botrytis cinerea exposure by up-regulating the expression of antioxidant genes (nrf2, ho-1, sod, and cat) in the Keap1-Nrf2 pathway. Additionally, quercetin inhibited the elevation of TNF-α by regulating the gene expression of key targets (jak3, pi3k, pdk1, akt, and ikk2) and the content of major proteins NF-κB (P65) and IκB in the NF-κB pathway. In conclusion, this work enriched the contents of the biological research of Botrytis cinerea and provided a new direction for the drug development and targeted therapy of quercetin.