Anthracnose caused by severely impacts global crop and forest productivity. Here, we identify A22, a novel small-molecule inhibitor that targets myosin I (CsMyoI), and this represents a previously untapped antifungal mechanism. Phenotypic screening, ATPase inhibition, and microscale thermophoresis (MST) confirmed that A22 binds to CsMyoI and disrupts its motor activity. Molecular dynamics simulations revealed stable interactions with the conserved phenamacril binding pocket, with a critical K379 substitution explaining species-specific differences in inhibitor sensitivity. Notably, A22 exhibited no cross-resistance with widely used fungicides, such as carbendazim or pyraclostrobin. In greenhouse trials, A22 outperformed the commercial fungicide prochloraz in controlling anthracnose on pepper, mango, and strawberry, even at low concentrations, and also promoted strawberry growth. These findings establish myosin I as a novel antifungal target and present A22 as a potent agent with a unique mode of action, offering a promising strategy for managing anthracnose and fungicide-resistant fungal strains.

Gray mold caused by Botrytis cinerea severely impacts global crop productivity. In this study, we identified B33 [(Z)-3-amino-N-(3-chloro-4-fluorobenzyl)-2-cyano-3-phenylacrylamide], a novel phenamacril derivative bearing a distinctive substitution pattern where the ester group is replaced by an amino group. As an unprecedented small-molecule inhibitor targeting B. cinerea myosin I (BcMyoI), it represents an unexplored antifungal mode of action for gray mold disease management. Combined assays including phenotypic screening, ATPase inhibition, and microscale thermophoresis (MST) confirmed that B33 binds BcMyoI, and molecular dynamics simulations revealed stable interactions with the conserved phenamacril-binding pocket. In the greenhouse experiment, the protective effect of B33 on strawberry leaves was comparable to that of the commercial fungicide boscalid. These findings establish BcMyoI as a novel antifungal target and present B33 as a potent agent with a unique mode of action, thus providing a promising alternative strategy to control gray mold and diverse phytopathogens.

Fusarium head blight (FHB) is a devastating wheat disease causing yield losses and mycotoxin contamination. Phenamacril, a myosin I-targeting fungicide, has a limited antifungal spectrum. Using structure-based drug design focusing on M375 in Fusarium graminearum myosin I (FgMyoI), we synthesized 25 phenamacril derivatives. The II-2 emerged as the most potent inhibitor (EC50 = 0.0077 mg/L against F. graminearum), with broad-spectrum activity against various phytopathogens. Biochemical and computational analyses confirmed that II-2 tightly binds FgMyoI and inhibits its ATPase activity. In vivo tests demonstrated excellent protective and curative efficacy against FHB in wheat and corn, while toxicity assays revealed low risk to nontarget organisms. These results identify II-2 as a promising myosin inhibitor and a novel candidate for FHB management. This work also provides a foundation for the rational design of myosin-targeting fungicides against diverse plant fungal diseases.

Salicylic acid treatment significantly enhances the production of bioactive Euphorbia diterpenoids in Euphorbia lathyris suspension cultures by activating defense and diterpenoid biosynthetic pathways, including upregulating the key acyltransferase gene ElBAHD1. The medicinal plant Euphorbia lathyris produces pharmacologically active diterpenoids, but the content of which is limited in Euphorbia plants. An efficient suspension culture system of E. lathyris was established to enhance the production of bioactive diterpenoids. Salicylic acid (SA) treatment (550 μM, 10 days) significantly enhanced the production of lathyrane-type diterpenoids jolkinol A′ (1), jolkinol A (2), jolkinol B (5), and a new 16-hydroxylated jolkinol A derivative (7). In particular, the volumetric yield of jolkinol A (2) was achieved to 47.00 mg L−1, a 5.4-fold increase over unelicited controls. Intriguingly, SA also induced the production of polycyclic diterpenoids, including ent-atisane and ent-kaurane. Transcriptomic analysis revealed SA activated defense and diterpenoid biosynthetic pathways, upregulating a series of genes including the acyltransferase gene ElBAHD1. Overexpression of ElBAHD1 promoted the accumulation of jolkinol A′ (1), jolkinol A (2), and jolkinol B (5) in transgenic hairy roots and influenced the expression of diterpenoid biosynthetic genes. Notably, jolkinol A (2) exhibited potent antifungal activity against Phytophthora capsici, comparable to that of the commercial fungicide metalaxyl (84.76

Magnaporthe oryzae is a devastating fungal pathogen causing blast disease in rice and other crops, threatening global grain production and food security. Phenamacril (PHA) effectively inhibits Fusarium graminearum by targeting F. graminearum myosin I (FgMyoI). However, PHA shows limited activity against M. oryzae, despite the high sequence similarity between FgMyoI and M. oryzae myosin I (MoMyoI). Using our published PHA-FgMyoI complex structure as a template, we identified K378 in MoMyoI as a key determinant of insensitivity to PHA. Substitution of K378 with methionine, the corresponding residue in FgMyoI, markedly enhanced PHA-mediated inhibition of MoMyoI ATPase activity and improved PHA efficacy against M. oryzae. Guided by structure-based design, we synthesized PHA derivatives targeting MoMyoI. Among them, NJY-10 showed improved MoMyoI binding, ATPase inhibition, and protective efficacy against rice blast disease.

Euphorbia lathyris, a medicinal and bioenergy plant, suffers significant post-harvest losses due to fruit decay, which reduces biomass yield. Scoparone, a coumarin-type phytoalexin, is widely recognized as an indicator of increased resistance to post-harvest decay in plants. Scoparone acts as both a phytoalexin and a therapeutic compound for liver disorders. However, the O-methyltransferase from the E. lathyris (ElCOMT) gene family, key enzymes responsible for catalyzing the final methylation step in scoparone formation, remain poorly characterized. In this study, 34 ElCOMT genes were identified and classified into two groups: Type I and Type II. Chromosomal localization analysis revealed a non-uniform distribution of ElCOMTs across the E. lathyris genome. Tissue-specific expression profiling and metabolomic analysis in E. lathyris indicated that ElCOMT33 and ElCOMT34 are potentially involved in scoparone biosynthesis. In vitro enzyme activity assays demonstrated that ElCOMT33 functions as a multifunctional enzyme capable of catalyzing two successive O-methylation reactions, converting esculetin to scoparone via scopoletin as an intermediate. ElCOMT34 exhibits a more limited substrate specificity, mediating only the O-methylation of esculetin to yield isoscopoletin. Furthermore, ElCOMT33 demonstrated broad catalytic activity, effectively methylating coumarins, phenylpropanes, and flavonoids. Transgenic assays confirmed that overexpression of ElCOMT33 and ElCOMT34 increased scopoletin and isoscopoletin biosynthesis in E. lathyris hairy roots (in vivo), respectively. This study systematically characterized the OMT gene family in E. lathyris and revealed multiple O-methylation steps involved in the biosynthesis of the phytoalexin scoparone. The findings provided a solid theoretical framework for further research on increasing endogenous phytoalexin biosynthesis for breeding disease-resistant plants and reducing post-harvest loss.

Anthracnose caused by Colletotrichum siamense severely impacts global crop and forest productivity. Here, we identify A22, a novel small-molecule inhibitor that targets C. siamense myosin I (CsMyoI), and this represents a previously untapped antifungal mechanism. Phenotypic screening, ATPase inhibition, and microscale thermophoresis (MST) confirmed that A22 binds to CsMyoI and disrupts its motor activity. Molecular dynamics simulations revealed stable interactions with the conserved phenamacril binding pocket, with a critical K379 substitution explaining species-specific differences in inhibitor sensitivity. Notably, A22 exhibited no cross-resistance with widely used fungicides, such as carbendazim or pyraclostrobin. In greenhouse trials, A22 outperformed the commercial fungicide prochloraz in controlling anthracnose on pepper, mango, and strawberry, even at low concentrations, and also promoted strawberry growth. These findings establish myosin I as a novel antifungal target and present A22 as a potent agent with a unique mode of action, offering a promising strategy for managing anthracnose and fungicide-resistant fungal strains.

O-methyltransferases (OMTs) play a crucial role in the methylation of a range of secondary metabolites, such as phenylpropanoids, flavonoids, and lignins. However, the genetic mechanism underlying flavonoid methylation in the medicinal plant Musella lasiocarpa, a rich source of methylated flavonoids, remains to be elucidated. Here, we identified and characterized a new caffeic acid O-methyltransferase (COMT) enzyme, designated Ml05G3914, along with three other OMTs (Ml01G0494, Ml04G2958, and Ml08G0855), through genome and transcriptome association analysis. The potential role of MlOMT was studied by testing the catalytic activity of recombinant protein with numerous possible substrates in vitro. Our findings demonstrate that Ml01G0494, Ml04G2958, and Ml05G3914 are capable of catalyzing the monomethylation of baicalein, quercetin, luteolin, and caffeic acid as well as dimethylation of myricetin. Additionally, M1080855 can catalyze monomethylation of luteolin and caffeic acid and progressively facilitate the production of both mono-and di-methylated products of myricetin. This study provides insights into methylated flavonoid biosynthesis in M. lasiocarpa and might potentially contribute to the large-scale production of methylated flavonoids via biosynthesis approaches in the future.

Macrocyclic diterpenoid esters from Euphorbiaceae plants hold significant medicinal value owing to their structural diversity and for attributing structural uniqueness and biological efficacy. However, the responsible enzymes for the acylation of macrocyclic diterpenoids remain unknown. We identified two macrocyclic diterpenoid O-acyltransferases, ElBAHD16 and ElBAHD35, from the diterpene biosynthetic gene cluster of Euphorbia lathyris. ElBAHD16 and ElBAHD35 were characterized both in vitro (using Escherichia coli) and in vivo (using Nicotiana benthamiana and E. lathyris) and exhibited mono-acylation activities toward the hydroxy groups of their substrates, 7-hydroxylathyrol and lathyrol. ElBAHD16 showed not only regioselectivity toward the 7-OH group of 7-hydroxylathyrol but also donor promiscuity, thereby producing three different mono-acylation products. Conversely, ElBAHD35 demonstrated specific recognition for the 5-OH group of 7-hydroxylathyrol and lathyrol, thereby mediating mono-acetylation reactions with acetyl-CoA, showing donor specificity. Site-directed mutagenesis revealed that residues H154 and T363 in ElBAHD16 are critical for its catalytic activity. Notably, the Q35 residue enhanced the efficiency of ElBAHD16, while the M296, N292, and F394 residues were crucial for its donor promiscuity. These findings elucidate the last step in the biosynthesis of macrocyclic diterpenoid esters and highlight the contribution of acyltransferases to the structural diversity of diterpenoids.

Bacterial leaf blight caused by Xanthomonas oryzae pv. oryzae poses a significant threat to rice production worldwide. The identification of resistant varieties is crucial for the development of rice varieties that can withstand this disease. Currently, the identification of resistant varieties primarily relies on resistance genes. However, due to vertical resistance of resistance genes, varieties identified based on this criterion still encounter significant challenges. In this study, we collected eight major cultivated rice varieties in Jiangsu Province, China, along with two experimental varieties, ZH11 and NP. Employing Liquid Chromatograph-Mass Spectrometer (LC-MS), we quantified the accumulation levels of 5,10-diketo-casbene (DKC), a casbene-type diterpene phytoalexin, in these ten rice varieties following treatment with methyl jasmonate (MeJA). These results demonstrated that despite the exogenous application of MeJA, which is known to trigger the biosynthesis of DKC in both shoots and roots, the accumulation levels of this crucial diterpene phytoalexin varied significantly among the ten cultivars. In addition to assessing DKC levels, we also evaluated the resistance of the ten varieties to bacterial leaf blight. Our analysis revealed a positive correlation between DKC content and resistance to this devastating pathogen. To further investigate the molecular basis of this resistance, we selected NG9108 and MN1332 as representative varieties with the higher and weaker resistance to bacterial leaf blight, respectively, and then examined their transcriptional responses to MeJA treatment. By comparing the transcriptional profiles of these two contrasting varieties, we found that genes involved in the synthesis of other diterpene phytoalexins, such as CPS2 and CPS4, were markedly up-regulated in the NG9108 variety as compared to MN1332. The contrasting resistance to bacterial leaf blight between NG9108 and MN1332 can be attributed, at least in part, to their disparate capabilities in diterpene phytoalexin synthesis. In conclusion, our research not only highlighted the importance of phytoalexin synthesis in rice resistance but also offers a practical framework for utilizing phytoalexin content as a criterion in the screening process for rice resistance breeding in the face of escalating disease challenges.

Ginsenosides, the prominence active ingredients in ginseng, show various pharmacological activities. However, due to the difficulty of isomer separation, the research on the effect of the configuration of C-20 of ginsenoside compounds is restricted. Therefore, a systematic study is needed on the chiral preparation and separation mechanism of ginsenosides. An efficient and rapid HPLC analytical method with fifteen chiral stationary phases was developed for chiral separation of ginsenoside compounds in normal and reverse phase mode. Three and one ginsenoside compounds could be achieved completely separated (R-s > 1.5) by EnantioPak (R) Y1 and EnantioPak (R) R-C under the optimum chromatographic condition, respectively. Molecular simulation was performed to elucidate the chiral separation mechanism and chromatographic behavior of different analytes. The chiral recognition of ginsenoside compounds on EnantioPak (R) Y1 or EnantioPak (R) R-C was a spontaneous and entropy-enthalpy co-driven process, which governed by hydrogen bonding, van der Waals interaction, dipole-dipole interaction, electrostatic interaction and host-guest size-fit concept. Comprehensive analysis of experimental and calculated results, the relationship between structures of analytes, structures of chiral stationary phases and chiral separation parameters was discussed. This study aims to establish a rapid and efficient method for screening suitable chiral stationary phases; and also provide some guidance for the chiral separation of ginsenoside compounds, so then promote the pharmacological activity research of ginsenoside isomers.

BACKGROUND:Rice blast, a destructive fungal disease caused by Magnaporthe oryzae, severely impacts rice production worldwide. The increasing resistance of M. oryzae to multiple commercial fungicides has heightened the need for novel control agents. In previous studies, we identified two amaryllidaceous alkaloids, lycorine (LYC) and narciclasine (NAR), as potent antifungal agents against M. oryzae. However, their antifungal mechanisms remain poorly understood. RESULTS:In this study, RNA sequencing was used to examine the transcriptome of M. oryzae, followed by validation via quantitative real-time polymerase chain reaction analysis. The results suggested that both LYC and NAR greatly inhibited lipid metabolism and melanin metabolism in M. oryzae. In addition, LYC suppressed carbohydrate metabolism, whereas NAR interfered with mitosis. Liquid chromatography-mass spectrometry analysis further confirmed the suppression of lipid metabolism by both compounds, particularly the reduction in phosphatidylcholine content. Physiological and biochemical assays demonstrated that LYC and NAR significantly reduced melanin production and altered membrane permeability. LYC also evidently lowered carbohydrate and pyruvic acid levels, whereas NAR reduced both RNA and protein contents. CONCLUSION:In conclusion, LYC and NAR can both inhibit lipid metabolism and melanin synthesis of M. oryzae, with LYC also impacting carbohydrate metabolism and NAR disrupting mitosis. These findings provide a scientific basis for the development and utilization of LYC and NAR as novel control agents against rice blast. © 2025 Society of Chemical Industry.

The fruits of Lycium ruthenicum Murr. (Solanaceae) are employed in ethnomedicine and used as a functional food. Their antioxidant, anti-aging, and hypolipidemic activities have been investigated in modern research. This study indicated that the ethanolic extract of the fruits of L. ruthenicum Murr. (LRM) improved oxidative and heat stress tolerance, reduced the accumulation of lipofuscin, and retarded the aging process in Caenorhabditis elegans (Rhabditidae). Furthermore, the pharyngeal pumping rate and body length decreased under LRM treatment. Moreover, metabolomic analysis and the DPClusO algorithm revealed that LRM regulated a series of lifespan-related pathways centered on glycine, serine, and threonine metabolism. These results suggest that LRM prolongs the lifespan of Caenorhabditis elegans via dietary restriction. Moreover, feruloyl putrescine, a kind of polyamine, was found in differential metabolites, which may be the metabolite of caffeoyl-spermidine in LRM. These findings from this exploratory study offer a new insight into the roles of L. ruthenicum in anti-aging activity as a functional food.

Euphorbia lathyris L. is an important medicinal and fuel plant due to its abundant flavonoids, diterpenoids, and fatty acids. However, the lack of a reliable genetic transformation system in E. lathyris currently hinders the discovery and functional verification of genes related to its valuable secondary metabolites biosynthesis. In this study, we established and optimized tobacco rattle virus (TRV) based virus-induced gene silencing (VIGS) system in E. lathyris using magnesium chelatase I subunit (ElCHLI) as the reporter gene to expedite the gene verification process. The silencing efficiency reached the highest when the two-leaf stage seedlings were infiltrated at the concentration of Agrobacterium OD600 = 1.0 under the vacuum pressure of 6 kPa. E. lathyris ACT was validated as the reference gene for qRT-PCR experiments for TRV-infected plants. Based on the optimal conditions, the gene encoding casbene synthase (CBS) in E. lathyris was successfully silenced and the formation of ingenol in E. lathyris was significantly inhibited, indicating that CBS is a key enzyme for ingenol biosynthesis. These results demonstrate that TRV-mediated VIGS system provides an efficient approach for the gene functional verification in E. lathyris. The virus-induced gene silencing system in Euphorbia lathyris L. were established and optimized for the first time and casbene synthase was confirmed as a key enzyme in ingenol biosynthesis.

Arylmethylene bisindole alkaloids are predominantly found in cruciferous plants and as metabolites in marine organisms, exhibiting a diverse range of biological activities. In recent years, these alkaloids have been increasingly explored and utilized for controlling plant pathogenic fungi. However, the synthesis and underlying mechanisms, particularly their effectiveness against plant pathogens such as Clarireedia homoeocarpa, still require further investigation. In this study, a series of arylmethylene bisindoles were synthesized with high yield using aromatic aldehydes and indoles catalyzed by NiClO4 & sdot;6H2O. These compounds showed spectral antibacterial activity against six major phytopathogenic fungi and A13 exhibiting the highest antifungal activity against C. homoeocarpa with an EC50 value of 0.74 mg/L. Furthermore, in vivo antifungal assays were performed to assess the efficacy of the active compounds against C. homoeocarpa. To elucidate the antifungal mechanism of aryl-methylene bisindoles, a comprehensive analysis of physiological and biochemical indices, along with transcriptome profiling, was carried out to elucidate. The results demonstrated that A13 treatment can efficiently disrupt the cell membranes of C. homoeocarpa. Additionally, we identified and screened the candidate genes implicated in cell membrane alterations under A13 treatments. KEGG pathway analysis revealed that a considerable number of down-regulated DEGs were significantly enriched in metabolic pathways, biosynthesis of secondary metabolic, and ribosome biogenesis in eukaryotes. This research provides valuable insights for the effective management of lawn coin spot disease.

WRKY transcription factors (TFs) play pivotal roles in regulating plant nutrient uptake, particularly phosphate (Pi) acquisition, and biosynthesis of secondary metabolites. Euphorbia lathyris, a significant medicinal plant with diverse pharmacological activities, lacks a systematic analysis of WRKY members and their functional roles. In this study, 58 ElWRKY genes were identified in the E. lathyris genome, classified into seven subgroups through comparative genomics analysis, and distributed on 10 chromosomes. Phylogenetic and expression pattern analyses identified ElWRKY48 as a candidate gene involved in Pi uptake regulation. The transgenic validation assay demonstrated that ElWRKY48 overexpression negatively regulated Pi uptake and led to phosphorus-deficient phenotypes in the hairy roots of E. lathyris. Furthermore, the transcriptome analysis revealed an opposite expression pattern between the Pi transporter gene ElPHT1 (Elat0034050.1) and ElWRKY48 in the transgenic lines overexpressing ElWRKY48. The negative regulation of ElPHT1 expression by ElWRKY48 was validated through qRT-PCR, Y1H, EMSA, gene knockout (CRISPR/Cas9), and LUC assays. Additionally, the overexpression of ElWRKY48 reduced diterpenoid ingenol biosynthesis by suppressing the expression of its biosynthesis-related genes. These findings provide valuable insights into the role of WRKY in Pi uptake and offer potential avenues for genetic improvement in the yield and quality of E. lathyris.

Phenylphenalenones (PhPNs), phytoalexins in wild bananas (Musaceae), are known to act against various pathogens. However, the abundance of PhPNs in many Musaceae plants of economic importance is low. Knowledge of the biosynthesis of PhPNs and the application of biosynthetic approaches to improve their yield is vital for fighting banana diseases. However, the processes of PhPN biosynthesis, especially those involved in methylation modification, remain unclear. Musella lasiocarpa is a herbaceous plant belonging to Musaceae, and due to the abundant PhPNs, their biosynthesis in M. lasiocarpa has been the subject of much attention. In this study, we assembled a telomere-to-telomere gapless genome of M. lasiocarpa as the reference, and further integrated transcriptomic and metabolomic data to mine the candidate genes involved in PhPN biosynthesis. To elucidate the diversity of PhPNs in M. lasiocarpa, three screened O-methyltransferases (Ml01G0494, Ml04G2958, and Ml08G0855) by phylogenetic and expressional clues were subjected to in vitro enzymatic assays. The results show that the three were all novel O-methyltransferases involved in the biosynthesis of PhPN phytoalexins, among which Ml08G0855 was proved to function as a multifunctional enzyme targeting multiple hydroxyl groups in PhPN structure. Moreover, we tested the antifungal activity of PhPNs against Fusarium oxysporum and found that the methylated modification of PhPNs enhanced their antifungal activity. These findings provide valuable genetic resources in banana breeding and lay a foundation for improving disease resistance through molecular breeding.

Euphorbia lathyris L. is a biennial herb in the Euphorbiaceae that has been used as a medicinal plant. It is distributed or cultivated worldwide, and the seeds of E. lathyris are the main source of ingenol, which is the precursor of Picato, the first medicine approved by USFDA for the treatment of solar keratosis (Abramovits et al. 2013). However, the production of E. lathyris can be severely hampered by the occurrence of plant diseases. Between 2020-2022 (specifically in October-November of each year), anthracnose-like symptoms were observed on E. lathyris in fields (E 118°49'50″, N 32°3'33″) in Nanjing, Jiangsu Province, China. The incidence of E. lathyris with disease symptoms was between 25%-30% (n = 100). The lesions on the leaves were evident initially as dark brown spots, which expanded into larger necrotic spots, finally resulting in leaves withering and dropping off. In severe cases, stem wilting was also observed. To determine the causal agent, we collected diseased leaf samples (n = 20) from different E. lathyris plants in the field (~ 1800 m2). After cleaning, the junctions of the diseased and healthy parts were excised and sterilized in 75% ethanol for 20-25 seconds, and rinsed with sterile water. After that, they were transferred onto potato sucrose agar (PSA) plates and placed at 25℃ for 3-4 days, until fungal growth was evident. The fungus was purified by recovering single conidia and growing them on PSA (Hu et al. 2015). A consistent fungal colony, based on morphological characteristics, was recovered from 17 samples. The colony color was initially white, green in the middle, and gradually changed into gray green as the colony matured. Conidia were transparent and cylindrical (22-28 µm × 7-9 µm, n = 50). Five loci informative (ITS, TUB, ACT, GAPDH, and CHS-1) (Weir et al. 2012) for Colletotrichum spp. identification were sequenced from two isolates ELC-1 and ELC-2 obtained from different plant individuals. Compared with a reference isolate (Colletotrichum gloeosporioides ZH3), the GAPDH, CHS-1, and TUB2 sequences of ELC-1 and ELC-2 showed 95% (263 bp out of 275 bp), 98% (295 bp out of 299 bp), and 99% (711 bp out of 712 bp and 717 bp out of 719 bp) similarity, respectively. The ITS sequence identities were 100% (577 bp out of 577 bp) and 99% (594 bp out of 597 bp), while the ACT sequence identities were 100% (281 bp out of 281 bp) and 98% (279 bp out of 284 bp). All sequences have been deposited in Genbank database (OR865865-OR865866 and OR873625-OR873632). After performing phylogenetic analysis with Mega 11, the pathogen was confirmed as C. gloeosporioides. To fulfil Koch's postulates, we sprayed six-week-old healthy plants with a conidia suspension of C. gloeosporioides (106 spores/mL) or sterile water (serve as control). The inoculated plants were placed at 25℃, 100% relative humidity, and 12-h photoperiod (Zhang et al. 2021). Six plants were inoculated for each treatment, and the experiment was repeated three times. After 6-8 days, the plants inoculated with C. gloeosporioides showed similar symptoms to those observed on diseased plants in the field, while the control plants remained healthy and free of disease. The pathogens were then re-isolated and identified as C. gloeosporioides. To our knowledge, this is the first report of C. gloeosporioides causing anthracnose on E. lathyris. Anthracnose may cause significant yield losses in E. lathyris production, and our results will provide experimental and theoretical basis for the management of the disease.

Pecan [Carya illinoinensis (Wangenh.) Koch] is a widely cultivated dried fruit and woody oil tree with high economic value. Continuous sole planting of pecan caused some land to lie idle and deterioration of soil conditions. Tree and medicinal crops intercropping represents an environmental-friendly and economically feasible solution to these issues. Thus, we aimed to explore the underlying mechanism by which intercropping improved soil condition by regulating the interactions of the soil microbiome and metabolome. In this study, pecans were intercropped with medicinal crops caper spurge and honeysuckle on a tree farm in China. A combined analysis of soil microbiomes and metabolomes was performed to discover the effects of intercropping on bulk and rhizosphere soils. The results showed that intercropping improved the edaphic properties of bulk soil and promoted the growth of pecan and caper spurge. Intercropping also significantly altered the structures of both bacterial and fungal communities in bulk soil, stabilised the enrichment of nitrogen-cyclic bacteria, for instance, Bacillus, and decreased the relative abundances of plant–pathogenic fungi, for instance, Fusarium. In addition, the result of metabolomic analysis showed that intercropping promoted the synthesis of functional compounds, such as trehalose and ethanolamine, which enhanced plant disease resistance in bulk soils. Moreover, the co-occurrence networks of microbiomes and metabolomes of bulk soils revealed that Bacillus was significantly correlated with Fusarium, Alternaria, and trehalose under intercropping patterns. Furthermore, analysis of microbiomes and metabolomes in the rhizosphere soil of caper spurge and honeysuckle revealed that Penicillium and Rhizomicrobium were significantly increased by intercropping and showed more dynamic connections with other genera and metabolites compared with single planting. Overall, intercropping pecans with caper spurge and honeysuckle can improve soil conditions and promote plant growth through microbiological and metabolomics integrated analyses. This study provides valuable information and theoretical basis for optimizing land resource utilisation and improving soil conditions in tree fields like pecan fields via intercropping, thereby reducing production losses and ensuring economic benefits.

BACKGROUND: Bacterial leaf blight caused by Xanthomonas oryzae pv. oryzae (Xoo) is one of the most serious diseases of rice, and there is a lack of bactericides for controlling this disease. We previously found parthenolide (PTL) is a potential lead for developing bactericides against Xoo, and subunit F of respiratory chain complex I (NuoF) is an important target protein of PTL. However, the binding modes of PTL with NuoF need further elucidation. RESULTS: In this study, we obtained the crystal structure of Xoo NuoEF (complex of subunit E and F of respiratory chain complex I) with a resolution of 2.36 angstrom, which is the first report on the protein structure of NuoEF in plant-pathogenic bacteria. The possible binding sites of PTL with NuoF (Cys105 and Cys187) were predicted with molecular docking and mutated into alanine using a base mismatch method. The mutated proteins were expressed in Escherichia coli and purified with affinity chromatography. The binding abilities of PTL with mutated proteins were investigated via pull-down assay and BIAcore analysis, which revealed that double mutation of Cys105 and Cys187 in NuoF severely affected the binding ability of PTL with NuoF. In addition, the binding modes were further simulated with combined quantum mechanical/molecular mechanical calculations, and the results indicated that PTL may have a stronger binding with Cys105 than Cys187. CONCLUSION: NuoEF protein structure of Xoo was resolved, and Cys105 and Cys187 in NuoF are important binding sites of PTL. This study further clarified the action mechanism of PTL against Xoo, and will promote the innovation of bactericides targeting Xoo complex I. (c) 2024 Society of Chemical Industry.