Search Results (239)

Ethylene response factors (ERFs) are transcription factors specific to plants that serve critical functions in various aspects of plant growth, development, and responses to environmental stressors. Despite the significance of these factors, the specific mechanisms by which mulberry ERFs interact with and respond to the pathogenic fungus Botrytis cinerea have not yet been fully elucidated. This study focuses on the isolation of a particular ERF transcription factor, known as MnERF/ABR1, which is localized in the cell nucleus and is derived from mulberry. Overexpression of MnERF/ABR1 in Arabidopsis or transient overexpression of MnERF/ABR1 in mulberry leaves can significantly enhance its resistance to B. cinerea. Our results suggest that empty vector control (CK) has higher levels of malondialdehyde (MDA), a marker of oxidative stress, compared to overexpression lines. In contrast, the catalase (CAT) activity of overexpression lines was higher than that of CK plants. Furthermore, staining with 3,3′-diaminobenzidine (DAB) and nitro blue tetrazolium (NBT) indicated that the resistance to B. cinerea was more pronounced in plants with overexpression than in those of the CK plants. These findings uncovered the molecular regulatory pathway involving MnERF/ABR1 in response to B. cinerea and established a basis for the development of disease-resistant mulberry varieties through genome editing. Full article

Potato (Solanum tuberosum L.) is a vital food security and cash crop in Eritrea, yet bacterial and fungal pathogens responsible for 15–30% yield losses remain molecularly uncharacterized across its production systems. Here, we present the first nationwide amplicon-based metagenomic survey of potato pathogen communities, sampling 81 farms across 14 sub-regions spanning four agroclimatic regions during July–August 2023. High-throughput amplicon sequencing targeting the bacterial 16S rRNA V3–V4 region and fungal ITS1–ITS2 loci revealed pronounced geographic heterogeneity in community composition and alpha diversity. Pseudomonas spp. were ubiquitous, with a peak relative abundance of 32.5% in Dekemhare. Dominant fungi included Alternaria spp. (14.3% in Berik), Fusarium spp. (highest diversity 53.8% in Adi Kuala), Botrytis cinerea (36.9% in Adi Keih), and Rhizoctonia solani (44.4% in Adi Tekeliezan). Bacterial Shannon diversity averaged 5.67; fungal, 4.70. Weighted UniFrac PCoA accounted for 56.5% of bacterial community variance along PC1, confirming distinct geographic clustering. Potential pathogen-associated taxa of Pseudomonas, Alternaria, Fusarium, and Colletotrichum were detected in every asymptomatic sample examined, demonstrating the inadequacy of visual-only disease surveillance. These findings establish the first molecular baseline for potato-associated taxon diversity in Eritrea, providing the empirical foundation for region-specific integrated disease management and evidence-based seed certification protocols. Full article

Botrytis cinerea, the causal agent of gray mold disease, is a major phytopathogen responsible for substantial losses in horticultural crops. In this study, cultivation conditions for Bacillus velezensis R22 were optimized to maximize overall antifungal activity against B. cinerea. A Plackett–Burman design was used to identify medium components affecting antifungal activity in flask cultures, followed by response surface methodology based on a central composite design (CCD) to optimize sucrose concentration, temperature, and agitation speed in a stirred bioreactor. Maximum antifungal activity was obtained at 17.45 g/L initial sucrose, 31.8 °C, and 293 rpm. The biological relevance of the optimized culture was confirmed in a tomato infection model, in which gray mold severity was reduced by 85.3% relative to the untreated control and by 59.9% relative to the non-optimized R22 culture. The same CCD approach was subsequently applied to determine cultivation conditions that maximize the concentration of R22 viable cells. The optimal parameters for 24-h growth (35.46 g/L sucrose, 36.5 °C, and 454 rpm) differed markedly from those identified for maximal antifungal activity. When evaluated on uninfected tomato plants, cultures produced under conditions favoring higher cell density showed enhanced plant growth-promoting activity compared to the non-optimized culture. Mass spectrometric analysis of lipopeptide extracts revealed that the enhanced antifungal activity was accompanied by an increased abundance of long-chain homologs across all major lipopeptide families, particularly surfactins. Thus, our results indicate that maximizing overall antifungal activity may be of greater practical significance than optimization of the individual fungicidal agent. Full article

Botrytis cinerea is a major phytopathogenic fungus responsible for substantial economic losses in horticultural crops, underscoring the need for sustainable alternatives to synthetic fungicides. This study investigated the influence of physical, chemical and biological culture parameters on the antifungal activity of culture filtrates produced by Trichoderma harzianum BOL-12QD. Culture conditions were sequentially optimised by evaluating light-filter exposure, carbon and nitrogen source composition, potato ecotype selection, co-cultivation with Botrytis cinerea, and volatile-mediated interactions. Antifungal activity was assessed using mycelial growth inhibition assays against Botrytis cinerea. Among the individual factors, violet-filter illumination, a medium containing 5 g L⁻¹ glucose and 250 g L⁻¹ potato extract, the Leke Pek’e potato ecotype, ammonium nitrate as nitrogen source, and co-cultivation with Botrytis cinerea at 10⁴ conidia mL⁻¹ produced the highest inhibitory effects. Sequential integration of these optimised conditions resulted in enhanced antifungal activity, reaching up to 62% inhibition. Volatile organic compounds produced by Trichoderma harzianum BOL-12QD exhibited only minimal antifungal activity under the conditions tested, suggesting that volatile-mediated antagonism plays a limited role in this system. In contrast, culture-dependent modulation of extracellular metabolite profiles was evidenced by comparative ¹H NMR fingerprinting, which revealed condition-specific spectral differences, with the optimised treatment displaying a distinct metabolic signature relative to all other conditions. Cytotoxicity assays in murine peritoneal macrophages showed no significant reduction in cell viability at concentrations up to 200 μg mL⁻¹. In vivo exposure to the optimised culture filtrate (250 mg kg⁻¹ d⁻¹ for 10 days) induced transient treatment-related clinical observations without mortality, indicating a need for further detailed toxicological characterisation. Overall, these findings demonstrate that the antifungal activity of Trichoderma harzianum BOL-12QD is strongly modulated by interacting environmental, nutritional and biological culture parameters. The results support the potential of optimised culture filtrates as a source of bioactive metabolites for biocontrol applications, while highlighting the importance of integrated biochemical and toxicological evaluation. Full article

Necrosis- and ethylene-inducing peptide 1 (Nep1)-like proteins (NLPs) are conserved microbial proteins that contain immunogenic patterns capable of activating plant pattern-triggered immunity (PTI). NLP patterns from Sclerotinia sclerotiorum (SsNLPs), a destructive necrotrophic fungal pathogen with a broad host range, have been identified, and their roles in PTI have been revealed. Nevertheless, the molecular mechanisms by which SsNLPs stimulate plant immunity remain largely unknown. In this study, we phylogenetically characterized SsNLPs and demonstrated the involvement of the phytocytokine receptor-like kinases PEPRs in SsNLP1-triggered immunity. SsNLPs contained the NPP1 domain and GHRHDWE motif and were phylogenetically closely associated with Botrytis cinerea NLPs. SsNLP1 treatment strongly induced the expression of PEPR genes. Further genetic analyses using Arabidopsis wild-type, pepr1 pepr2 double mutant, and PEPR1 overexpression lines showed that SsNLP1 elicited diverse immune responses, including reactive oxygen species (ROS) accumulation and defense gene activation, and induced plant resistance to S. sclerotiorum. Notably, the induced plant resistance and immune responses were strengthened in PEPR1 overexpression lines and significantly reduced in the pepr1 pepr2 mutant, indicating a positive role of PEPR signaling in SsNLP1-triggered immunity. Overall, our results revealed that phytocytokine PEPR1 signaling amplifies PAMP SsNLP1-triggered immunity, thereby enhancing resistance against S. sclerotiorum. Our findings provide an example of the coordination between PAMP- and phytocytokine-triggered immunity for robust resistance to a necrotrophic pathogen. Full article

Botrytis cinerea is a major phytopathogen responsible for significant postharvest losses in plant-derived foods. The increasing resistance to synthetic fungicides has driven the search for sustainable alternatives, including enzyme-based biofungicides. In this study, the proteolytic fraction P1G10 from Vasconcellea pubescens latex was encapsulated in an alginate–chitosan (ALG-CS) matrix to improve its stability and antifungal performance. The encapsulated formulation (ALG-CS-P1G10) retained ~95% enzymatic activity after 8 h under stress conditions (37 °C, 1350 lux), compared with 67% for the free enzyme. In vitro assays demonstrated a dose-dependent inhibition of B. cinerea growth, with an IC₅₀ value of ~11 mg/mL determined using a logistic model. At this concentration, the formulation reduced fungal adhesion by more than 80% and increased sensitivity to cell wall-disrupting agents (Congo Red and Calcofluor White), pointing to alterations in cell wall integrity. Importantly, the encapsulated system provided a more stable and sustained antifungal effect, consistent with a controlled-release mechanism. These results demonstrate that coupling enzyme stabilization with controlled release can improve the functional performance of protease-based antifungal systems, offering a promising strategy for the development of biofungicides in postharvest applications. Full article

Plant hormones and their synthetic analogueues offer sustainable alternatives for crop protection, yet the direct antifungal activity of strigolactone (SL) and its analogues against necrotrophic pathogens remain largely unexplored. Here, we screened eight phytohormones and related analogues for treatments of Botrytis cinerea and identified the SL analogue rac-GR24 (racemic GR24) as a concentration-dependent growth inhibitor active at low micromolar concentrations. Given the stereochemical complexity of SLs and their analogues, we evaluated multiple enantiopure isomers and found that ent-5DS and GR24^ent-5DS, which differ in configuration from natural SLs, exhibited the strongest inhibitory activity. This stereospecific response was further validated using another filamentous fungus, Sclerotinia sclerotiorum, which displayed an identical susceptibility profile. Combinatorial treatments with enantiopure isomers and double-concentration rac-GR24 revealed that the antifungal effect of the racemate is primarily attributable to the GR24^ent-5DS enantiomer, whereas the opposite enantiomer GR24^5DS is almost inactive. Collectively, our findings uncover a stereospecific response in fungal pathogens, demonstrating that B. cinerea and S. sclerotiorum respond to exogenous SL analogues in a chirally selective manner. This work establishes a stereochemically defined framework for developing enantioselective fungicidal agents with potential applications in sustainable agriculture. Full article

Papain-like cysteine proteases (PLCPs) are central immune hubs frequently targeted by pathogen effectors. Sugarcane smut, caused by Sporisorium scitamineum, threatens global sugarcane yield, yet effector manipulation of host PLCPs remains unclear. Genome-wide analysis of Saccharum spontaneum AP85-441 identified 61 PLCP-encoding genes, which were classified into nine conserved subfamilies. Among these, ScRD21A, a member of the RD21 subfamily, was prioritized for functional characterization. Two Pit2 homologs, SsPit2A and SsPit2B, were identified from S. scitamineum. Yeast two-hybrid, BiFC and pull-down assays demonstrated that both effectors interact with ScRD21A, and that this interaction depends on a conserved LXRR motif within their PID14-like region. In total protein extracts from Nicotiana benthamiana, co-expression of SsPit2A or SsPit2B reduced ScRD21A-associated cysteine protease activity. Transient expression of ScRD21A enhanced flg22-induced ROS production, attenuated Pst DC3000-induced hypersensitive response-associated necrosis, and increased resistance to Botrytis cinerea. Together, these results support a conserved PLCP-targeting strategy in smut fungi and identify the ScRD21A–SsPit2A/B module as a tractable framework for studying effector–protease interactions relevant to sugarcane smut. Full article

Grapevine (Vitis vinifera L.) is an economically important fruit crop cultivated worldwide. However, its production and fruit quality are severely constrained by powdery mildew (Erysiphe necator) and Botrytis bunch rot (Botrytis cinerea) diseases. Increasing concerns regarding chemical fungicide resistance and environmental sustainability highlight the urgent need to develop alternative and more sustainable disease management strategies. This study assessed the field efficacy of Pseudomonas protegens-based formulations (TANIRI^® WP at 1 g·L⁻¹ and MaxGrowth at 1 mL·L⁻¹) within an integrated disease management program in cv. Chardonnay. Defense-related gene expression analysis revealed that biological treatments predominantly up-regulated pr1, pr2, and pr10 in both leaves and berries. In contrast, the chemical inducer acibenzolar-S-methyl (ASM) triggered earlier but less consistent induction of pr1 and pr2, alongside transient activation of pal and lox9. Repeated field applications of P. protegens formulations moderately reduced the severity of Botrytis bunch rot (20.89%) and powdery mildew (6.14%), though control levels remained below conventional sulfur/Bacillus subtilis-based treatments (30.04% and 13.56%, respectively). Overall, these findings suggest that biological inducers could complement conventional management practices for grapevine health. In particular, P. protegens may act mainly by systemically inducing host defense responses and partially suppressing pathogen development under field conditions. Full article

Gray mold caused by Botrytis cinerea poses a severe threat to tomato production. In this study, physiological, biochemical, transcriptomic, and proteomic analyses were integrated to characterize the dynamic responses of tomato ‘Ailsa Craig’ to B. cinerea infection. During B. cinerea infection, peroxidase (POD) activity showed a progressive increase, while catalase (CAT) activity was significantly upregulated at 24 hpi and remained stable through 48 hpi. Malondialdehyde (MDA) and hydrogen peroxide (H₂O₂) contents showed a delayed response, increasing significantly only at 48 hpi, whereas SOD activity exhibited a biphasic pattern. Transcriptome and proteome profiling identified 5824 differentially expressed genes and 124 differentially expressed proteins. Functional enrichment analysis highlighted defense-related pathways, including plant–pathogen interaction, flavonoid biosynthesis, and inositol phosphate metabolism. Notably, the chlorophyll a/b-binding protein SlLhcb13 exhibited post-transcriptional upregulation despite transcriptional suppression. Functional validation demonstrated that overexpression of SlLhcb13 enhanced resistance, whereas silencing increased susceptibility. These findings identify SlLhcb13 as a positive regulator linking photosynthesis to immunity and provide new insights into the defense mechanisms of tomato. Full article

Proteomic research in the genus Vitis has progressed from early biochemical studies of soluble proteins to high-resolution, quantitative analyses encompassing all major organs and derived products. This review provides a comprehensive synthesis of advances in grapevine and wine proteomics. In leaves, studies have revealed extensive remodeling of photosynthetic, antioxidant, and defense pathways under biotic (e.g., Plasmopara viticola, Erysiphe necator, Xylella fastidiosa, Candidatus Phytoplasma vitis) and abiotic stresses (drought, salinity, heat, light). Bud proteomics elucidated hormonal regulation and mechanisms of dormancy release, while root studies identified nitrate-dependent metabolic shifts and adaptive protein networks. Cell culture models enabled controlled investigation of elicitor responses, stilbene biosynthesis, and temperature-induced proteome changes. In berries, proteomics clarified developmental transitions from fruit set to ripening, emphasizing proteins related to secondary metabolism, vacuolar transport, and stress tolerance. Comparative analyses across cultivars and environments identified biomarkers linked to aroma, color, and texture. The wine proteome revealed selective persistence of grape-derived proteins (e.g., thaumatin-like proteins, chitinases) and yeast peptides influencing stability and sensory properties, while Botrytis cinerea infection significantly alters this balance by degrading PR proteins and introducing fungal enzymes. Altogether, the Vitis proteome emerges as a dynamic, multifunctional system crucial for understanding plant adaptation, enological quality, and biomarker discovery. Full article

Fungal pathogens pose a major threat to global agriculture and human health, necessitating alternative antifungal strategies with high efficacy and low resistance potential. Antifungal proteins (AFPs) from filamentous fungi are promising candidates due to their stability, selectivity, and diverse mechanisms of action. Here, we characterize Beauveria brongniartii antifungal protein (BbroAFP), a novel cysteine-rich protein from the entomopathogenic fungus B. brongniartii, and investigate its antifungal activity against Botrytis cinerea. Recombinant BbroAFP was expressed in Pichia pastoris, purified, and verified by liquid chromatography–tandem mass spectroscopy (LC–MS/MS) and in silico modeling. BbroAFP showed potent antifungal activity with minimum inhibitory concentrations (MICs) as low as 1 µM against several phytopathogenic fungi, while exhibiting no significant antibacterial activity. Activity was maintained across a wide range of pH and temperature conditions. Confocal microscopy revealed rapid surface binding followed by cytosolic internalization without major cell wall disruption. BbroAFP induced a rapid, transient burst of reactive oxygen species (ROS), accompanied by nuclear DNA fragmentation. Gene expression analysis revealed a transient increase in aif1, whereas mca1 expression decreased at later time points and mca2 remained largely unchanged, suggesting a metacaspase-independent response. Detached tomato leaf assays showed effective protection against B. cinerea without detectable phytotoxicity. Cytotoxicity assays confirmed a favorable safety profile, supporting further evaluation of BbroAFP for plant protection. Full article

Epigenetic regulation plays a central role in modulating plant immune responses and interactions with beneficial microbes. In this study, we investigated the contribution of RNA-directed DNA methylation (RdDM) components—DCL3; AGO9; DCL1; and the de novo DNA methyltransferases CMT3, DRM1, and DRM2—to the interaction between Arabidopsis thaliana, Trichoderma atroviride, and foliar pathogens. We show that DCL3 and AGO9 differentially regulate basal and inducible immunity, negatively affecting resistance to the necrotrophic fungus Botrytis cinerea, while promoting defense against the hemibiotrophic bacterium Pseudomonas syringae pv. tomato DC3000. Transcriptional analyses revealed that RdDM components modulate the balance between jasmonic acid/ethylene (JA/ET) and salicylic acid (SA) signaling pathways, influencing the amplitude and coordination of defense responses. In addition, DCL3 and DCL1 appear to be required for the full expression of T. atroviride-mediated systemic resistance, whereas AGO9 and DNA methyltransferases contribute to efficient root colonization. Notably, mutants in these pathways displayed enhanced basal resistance but impaired responsiveness to beneficial microbial signals, revealing a trade-off between constitutive defense activation and inducible systemic protection. Consistent with this, alterations in RdDM components were also associated with changes in plant growth dynamics under specific conditions, supporting a role for epigenetic regulation in coordinating growth–defense trade-offs. Together, our findings support a model in which epigenetic regulation controls defense responsiveness, enabling plants to balance immune activation, growth and compatibility toward beneficial microbes. Full article

Botrytis cinerea, the causal agent of grey mold in grapevine, remains one of the most economically important pathogens in viticulture and a key target for sustainable biocontrol strategies. This study evaluated the antagonistic potential of seven Trichoderma isolates (T1–T7), collected from the rhizosphere of grapevine in Morocco, using a combination of in vitro and in planta assays designed to capture multiple direct and indirect modes of action. The isolates exhibited variable levels of antagonism through competition, volatile organic compounds, extracellular metabolites, and elicitation responses. Preliminary in planta assays on detached grape berries further demonstrated that all selected isolates reduced lesion development, with preventive applications yielding the strongest protection. Overall, the study highlights the complementary and strain-specific mechanisms underlying Trichoderma & B. cinerea interactions and underscores the importance of isolate selection and application timing for the development of effective and environmentally friendly grey mold management strategies. These findings provide a mechanistic basis for the future evaluation of promising isolates under vineyard conditions. Full article

The existence of lignin in Physcomitrium patens has been controversial. However, cinnamyl alcohol dehydrogenase (CAD), the key enzyme in monolignol biosynthesis, has been identified with four gene members in P. patens. Despite the roles of PpCAD1 in moss architecture being proven in a previous study, the functions and molecular mechanisms of PpCAD4 remain largely unexplored in early terrestrial plants. This study aims to unravel this mystery via a comprehensive analysis of the transcriptome and metabolome of PpCAD4-overexpression (OE) lines compared with wild type (WT) under Botrytis cinerea treatment, firstly. A total of 475 and 1368 significantly differentially expressed genes in PpCAD4-OE lines compared to the wild type at 6 h and 12 h post-inoculation, which were predominantly enriched in pathways involving flavonoid, phenylpropanoid biosynthesis, and plant hormone signal transduction. Concurrently, metabolomic profiling revealed 160 and 114 differentially accumulated metabolites in PpCAD4-OE at the corresponding time points, with phenolic acids and flavonoids collectively constituting over 45% of these compounds. Furthermore, the MADS-box transcriptional factor PpMC6 negatively regulated PpCAD4 expression by yeast-one-hybrid and dual-luciferase assays. Finally, Catalase isozyme 2 (PpCAT2) and E3 ubiquitin-protein ligase (PpE3) were identified as interactive partners with PpCAD4, respectively, deducing that the increasing of reactive oxygen species might be promoted by PpCAT2 degradation through PpE3 after B. cinerea assault. Our results demonstrated that the essential roles and potential mechanisms of PpCAD4 are essential for defense against pathogens during the adaptation to land in moss. Full article