Poplar (Populus spp.) canker severely threatens poplar shelterbelt stability in Wuwei, Gansu. Field experiments were conducted from 2024 to 2025 to screen 16 single fungicides and 10 compound formulations via spray and smearing, using lesion inhibition and callus formation rates as core criteria, and establish a precise hierarchical control scheme. Results showed preventive agents (300-fold Bordeaux mixture, 45% lime sulfur mixture) achieved >75% control efficacy. 43% tebuconazole and 1.8% xinjunamine acetate had the strongest bacteriostatic effects, while compound systems combining fungicides, penetrants and immune inducers showed >88% efficacy. A four-grade precise control strategy was finally established, providing technical support for eco-friendly poplar canker control in arid Northwest China.

Fungicides currently used to control poplar Cytospora canker (PCC), mainly caused by Cytospora chrysosperma, exhibit variable field efficacy, require high application frequency, pose resistance risks, and impose considerable environmental burdens. In this study, we developed a six-arm star polymer (SSP)-based nanocarrier system to deliver benomyl and carbendazim, aiming to enhance the control efficacy against C. chrysosperma. The SSP formed stable complexes with benomyl and carbendazim through hydrogen bonding and Van der Waals interactions, reducing their particle sizes from 864.36 nm to 14.76 nm and 2725.44 nm to 33.16 nm, respectively. Nanoagents demonstrated markedly enhanced antifungal efficacy over standalone fungicides. Specifically, SSP-benomyl at 3.8 mg/L (IC50 of benomyl) reduced the colony diameter of C. chrysosperma from 80 mm to 23 mm, while SSP-carbendazim at 2.6 mg/L (IC50 of carbendazim) reduced it from 80 mm to 29 mm. Besides high inhibition of fungal growth and control efficiency of PCC, these SSP-based nanoagents exhibited potential disease prevention. Our results provide a nanopesticide delivery system of SSP to manage PCC.

The fungal family Botryosphaeriaceae, which includes genera such as Diplodia, Dothiorella, and Phaeobotryon, comprises species commonly associated with woody plants such as endophytes, pathogens, and saprophytes. The Xizang Autonomous Region of China, known for its rich forest resources, harbors significant fungal diversity. However, limited research has been conducted on plant-disease-associated fungi in this region. In this study, we employed morphological characteristics and molecular phylogenetic analyses of the internal transcribed spacer region of rDNA (ITS), the ribosomal large subunit (LSU), the translation elongation factor 1-alpha (tef1) gene, and the partial beta-tubulin (tub2) gene to identify fungal species. As a result, two new species, Diplodia salicicola sp. nov. and Phaeobotryon xizangense sp. nov., are proposed and described herein. Additionally, Di. corticola, Di. mutila, Do. acericola, Do. magnoliae, Do. vidmadera, Do. yunnana comb. nov., and Do. zanthoxyli are reported for the first time in Xizang. Our findings contribute to advancing the knowledge of fungal biodiversity in Xizang’s high-altitude ecosystems.

Haloxylon ammodendron is a highly salt-tolerant plant vital for desertification control in northwest China. Despite its ecological importance, the molecular mechanisms underlying its exceptional salt tolerance remain largely unexplored. This study aimed to elucidate the temporal dynamics of its transcriptomic responses to varying salinity levels. Temporal analysis revealed distinct gene expression patterns across low, medium, and high salt concentrations, with unique regulatory trends over time. Differential expression analysis identified 2,630 DEGs at 7 days, 4,533 DEGs at 21 days, and 2,581 DEGs at 30 days, highlighting 21 days as a critical period for salt response. WGCNA on 19,399 genes at day 21 revealed three modules (ME4-yellow, ME6-red, ME9-magenta) significantly associated with salt stress. These modules were enriched in genes involved in photosynthesis, amino acid metabolism, carbohydrate metabolism, and stress response pathways. Hub gene analysis identified ATPD and five sub-key genes as central regulators of the salt response network. This study provides the first comprehensive temporal transcriptomic analysis of H. ammodendron under varying salinity concentrations, revealing novel molecular insights into its salt adaptation mechanisms. The identified hub genes and pathways offer valuable targets for understanding extreme salt tolerance and enhancing desert reclamation efforts in arid regions.

Proteins harboring the PDZ domain are of utmost significance in the infection course of pathogenic bacteria, as well as in the response to external environmental stresses. In this study, we demonstrated that the Lonsdalea populi genome encodes a set of five proteins with the PDZ domain. Through a systematic inactivation of the genes responsible for encoding PDZ proteins, we showed that all these genes are closely related to the virulence of L. populi. Notably, deletion of prc results in suppression of the growth of L. populi and enhanced susceptibility to a diverse array of environmental stressors, such as hydrogen peroxide (H2O2), chloramphenicol, spectinomycin, and metal ions such as Mn2+, Fe2+, and Fe3+. Prc contains four domains: TSPn, PDZ, PEP, and DUF. By separately knocking out the four domains within Prc, we have demonstrated that the TSPn, PDZ, and PEP domains within Prc are all essential components in the pathogenic process of L. populi. Moreover, Prc interacts with the membrane protein YccA. This interaction regulates the biofilm formation capacity and motility of L. populi, thus exerting an impact on its virulence. Together, these findings suggest that Prc is an important regulator of virulence and stress response in L. populi.

Poplar Cytospora canker, caused by Cytospora chrysosperma, is one of the most destructive and widespread poplar diseases worldwide, especially in northern China. However, our current understanding of its pathogenic mechanisms remains limited. Here, we show that trehalose biosynthetic genes, such as trehalose-6-phosphate synthase 1 (CcTps1), trehalose-6-phosphate phosphatase (CcTps2), and the regulatory subunit (CcTps3), play important roles in the development and virulence of C. chrysosperma. The targeted deletion mutants showed reduced trehalose synthesis and were defective in hyphal growth and conidiation. Deletion of any of the three genes attenuated virulence in poplar twigs, and stronger poplar defense responses were triggered after inoculated by the mutants. Additionally, the mutants exhibited increased sensitivity to H2O2 and cell wall stressors. Taken together, the findings suggest that trehalose biosynthetic genes contribute to fungal development, stress responses, and full virulence in C. chrysosperma.

In recent years, Castanopsis carlesii, a keystone species in southern China’s forest ecosystems with high ecological and economic importance, has faced growing challenges from severe nut rot diseases. Gnomoniopsis (Gnomoniaceae, Diaporthales, Sordariomycetes, Ascomycota) represents a significant fungal genus that causes leaf spots, branch cankers, and fruit rot diseases. In this study, rotten nuts of C. carlesii were collected from Fujian Province, and fungal isolates were obtained using the tissue isolation method. Morphological characterization and molecular phylogenetic analysis, based on the combined sequences of the internal transcribed spacer region of rDNA (ITS), the translation elongation factor 1-alpha (tef1) gene, and the partial beta-tubulin (tub2) gene were used to identify these isolates. As a result, new isolates from diseased nuts of C. carlesii formed a distinct clade with Gnomoniopsis, and morphologically differentiated from the other species; hence, G. flava sp. nov. is proposed herein. Furthermore, pathogenicity tests involving three isolates of G. flava were conducted on healthy nuts of C. carlesii, confirming its role as the causal agent of this new plant disease. This study not only advances our understanding of species diversity within Gnomoniopsis but also lays the groundwork for developing control strategies for C. carlesii nut rot disease.

Poplar canker, caused by the fungus Cytospora chrysosperma, results in tremendous losses in poplar plantations in China. Although NADPH oxidases (NOXs) play important roles in the development and pathogenicity of several pathogenic fungi, their roles in C. chrysosperma remain unclear. In this study, we characterized three NOX genes (CcNox1, CcNox2, and CcNoxR) in C. chrysosperma. All three genes were highly upregulated during poplar branch infection, and deletion of any of them severely reduced virulence on poplar branches. Furthermore, deletion of either CcNox1 or CcNoxR resulted in a significant increase in endogenous reactive oxygen species production in hyphae, enhanced influx of Ca2+, the disruption of redox homeostasis and compromised mitochondrial integrity. Moreover, biosynthesis and secretion of a known virulence factor oxalic acid was obviously defective and exogenous oxalic acid supplementation rescued the virulence of the mutants. Taken together, our findings reveal that NOXs play important roles in redox homeostasis, mitochondrial integrity and pathogenicity in C. chrysosperma.

Poplar bacterial canker, caused by Lonsdalea populi, seriously threatens the health of poplar plantations. The two-component system is one of the most important signal transduction pathways in bacteria, playing a crucial role in numerical traits including growth, stress tolerance, and pathogenicity; however, their functions in L. populi remain poorly understood. Here, we identified a two-component system (RstB/RstA) in L. populi. The deletion of either rstB or rstA led to a decrease in L. populi virulence, biofilm formation, and tolerance to H2O2 and iron stresses. Based on transcriptomic data of the rstB or rstA mutants, an electrophoretic mobility shift assay confirmed that RstA directly binds to the promoter of lpfetA. Given that lpfetA, lpfetB, and lpfetC are co-transcribed as an operon, these results suggested that RstA likely regulates the lpfetABC operon. The overexpression of either lpfetA, lpfetB, or lpfetC almost abolished bacterial virulence, while resulted in decreased biofilm formation and tolerance to H2O2. These findings demonstrated that RstA modulates L. populi virulence, at least in part, by regulating the lpfet operon. Overall, our work indicated that RstB/RstA plays an important role in bacterial virulence and stress tolerance.

Bacteria employ two-component systems (TCSs) to rapidly sense and respond to their surroundings often and during plant infection. Poplar canker caused by Lonsdalea populi is an emerging woody bacterial disease that leads to high mortality and poplar plantation losses in China. Nonetheless, the information about the underlying mechanism of pathogenesis remains scarce. Therefore, in this study, we reported the role of a TCS pair CpxA/CpxR in regulating virulence and stress responses in L. populi. The CpxA/R system is essential during infection, flagellum formation, and oxidative stress response. Specifically, the Cpx system affected flagellum formation by controlling the expression of flagellum-related genes. CpxR, which was activated by phosphorylation in the presence of CpxA, participated in the transcriptional regulation of a chaperone sctU and the type III secretion system (T3SS)-related genes, thereby influencing T3SS functions during L. populi infection. Phosphorylated CpxR directly manipulated the transcription of a membrane protein-coding gene yccA and the deletion of yccA resulted in reduced virulence and increased sensitivity to H2O2. Furthermore, we mutated the conserved phosphorylation site of CpxR and found that CpxRD51A could no longer bind to the yccA promoter but could still bind to the sctU promoter. Together, our findings elucidate the roles of the Cpx system in regulating virulence and reactive oxygen species resistance and provide further evidence that the TCS is crucial during infection and stress response.

Different poplar varieties vary in their tolerance to certain pathogens. However, knowledge about molecular regulation and critical responses of resistant poplars during pathogen infection remains scarce. To investigate adaptive responses to canker disease caused by the bacterium Lonsdalea populi, we screened three poplar varieties with contrasting tolerance, including Populus deltoides. 'Zhonglin 2025' (2025), Populus × Euramericana. '74/76' (107) and Populus tomentosa cv 'henan' (P. tomentosa). Transcriptomic analysis revealed significant changes in the expression levels of defence-related genes in different poplar varieties in response to infection, which reshaped the PTI and ETI processes. Intriguingly, photosynthesis-related genes were found to be highly expressed in the resistant variety, whereas the opposite was observed in the susceptible variety. Susceptible poplars maintained the activation of defence-related genes during early period of onset, which restricted the expression of photosynthesis-related and auxin signal-related genes. Furthermore, combined with metabolomic analysis, differences in the content of antibacterial substances and key differentially expressed genes in phenylpropane and flavonoid biosynthesis pathways were identified. Delayed induction of catechin in the susceptible variety and it's in vitro antibacterial activity were considered to be one of the important reasons for the differences in resistance to L. populi compared with the resistant variety, which is of practical interest for tree breeding. Moreover, the trade-off between growth and defence observed among the three poplar varieties during infection provides new insights into the multilevel regulatory circuits in tree-pathogen interactions.

[目的]欧美杨细菌性溃疡病是革兰氏阴性细菌Lonsdalea populi引起的杨树枝干病害,其危害严重,已造成欧美杨人工林的重要经济损失.双组分系统是细菌致病过程的关键调控途径之一.目前,欧美杨细菌性溃疡病菌的双组分系统如何调控致病过程仍缺乏系统研究.因此,本研究开展欧美杨细菌性溃疡病菌的双组分编码基因的缺失突变及突变体表型分析,为深入解析其致病机制提供遗传材料.[方法]本研究以欧美杨溃疡病菌菌株N-5-1为研究对象,利用双亲结合方法获得了28个双组分系统基因的缺失突变体,并通过表型测定方法分析了这些基因突变体的致病性、生长、游动性、生物膜形成和抗逆性等表型特征,研究不同双组分系统编码基因对该病菌致病过程的调控.[结果]构建了36个欧美杨溃疡病菌的双组分编码基因的敲除重组载体,获得了28个基因的缺失突变体.致病性测定表明18个双组分基因的敲除降低了病原菌的毒性,其中8个突变体毒性丧失.此外,还获得了调控游动性和生物膜形成能力的突变体以及在逆境胁迫反应(金属离子、盐离子、抗生素等胁迫)有缺陷的突变体.[结论]本研究获得了5个显著影响欧美杨细菌性溃疡病菌毒性及其他生物表型的双组分基因,为后续双组分信号调控致病机制研究提供了遗传材料.

An emerging poplar canker caused by the gram-negative bacterium, Lonsdalea populi, has led to high mortality of hybrid poplars Populus × euramericana in China and Europe. The molecular bases of pathogenicity and bark adaptation of L. populi have become a focus of recent research. This study revealed the whole genome sequence and identified putative virulence factors of L. populi. A high-quality L. populi genome sequence was assembled de novo, with a genome size of 3,859,707 bp, containing approximately 3434 genes and 107 RNAs (75 tRNA, 22 rRNA, and 10 ncRNA). The L. populi genome contained 380 virulence-associated genes, mainly encoding for adhesion, extracellular enzymes, secretory systems, and two-component transduction systems. The genome had 110 carbohydrate-active enzyme (CAZy)-coding genes and putative secreted proteins. The antibiotic-resistance database annotation listed that L. populi was resistant to penicillin, fluoroquinolone, and kasugamycin. Analysis of comparative genomics found that L. populi exhibited the highest homology with the L. britannica genome and L. populi encompassed 1905 specific genes, 1769 dispensable genes, and 1381 conserved genes, suggesting high evolutionary diversity and genomic plasticity. Moreover, the pan genome analysis revealed that the N-5-1 genome is an open genome. These findings provide important resources for understanding the molecular basis of the pathogenicity and biology of L. populi and the poplar-bacterium interaction.

The gram-negative bacterium Lonsdalea populi causes an emerging poplar (Populus × euramericana) canker resulting in severe losses to poplar production in China and Europe. Two-component signal transduction systems play important roles in the regulation of virulence and stress responses in phytopathogenic bacteria. We identified a two-component pair (Lqp2625-Lqp2624) in L. populi, highly homologous to DcuS-DcuR of Escherichia coli. Mutants lacking DcuS or DcuR displayed normal growth while their virulence on poplar twigs was impaired. An inability to produce flagella indicated that DcuS and DcuR are involved in biofilm formation and swimming motility. Moreover, the loss of DcuS or DcuR led to increased sensitivity to oxidative stress and chloramphenicol through downregulation of genes associated with catalases and the multidrug efflux pump, suggesting that the two-component pair contributes to cellular adaptation to oxidative and antibiotic stresses. We identified key domains and putative phosphorylation sites important for virulence and stress responses. Our findings reveal the functions of DcuS-DcuR in virulence and stress responses in L. populi and provide increasing evidence that two-component systems are crucial during the infection process and stress adaptation in bacteria.

Lonsdalea quercina subsp.populi引起的欧美杨溃疡病是严重威胁欧美杨人工林生产的细菌性病害.双组分系统是细菌最重要的信号转导通路之一,在细菌的生长繁殖、环境适应、胁迫耐受性以及致病过程中发挥重要作用.为探索L.quercina中双组分系统编码基因的功能,本研究利用同源重组对双组分系统编码基因lqp0812和lqp0813进行缺失突变,并研究其生物学功能.表型分析显示,与野生型菌株N-5-1相比,△lqp0812和△lqp0813突变体在生长速率、金属离子胁迫、盐胁迫、渗透胁迫及致病性方面没有显著差异;但△lqp0812突变体在游动性上与野生型N-5-1相比显著减弱;△lqp0812和△lqp0813突变体生物被膜的形成能力,对过氧化氢、抗生素的耐受性都显著增强.qRT-PCR结果显示,在△lqp0812和△lqp0813突变体中,外排泵基因acrA、mdtB、aaeB的表达量与野生型相比显著升高.研究结果表明,lqp0812参与病原菌的游动性,lqp0812与lqp0813共同负调控菌株生物膜的形成和对氧化胁迫、抗生素胁迫的耐受性.

MarR family transcription regulators are ubiquitous among bacteria and archaea. They extensively control multiple cellular processes and elaborately regulate the expression of genes involved in virulence, stress response and antibiotics at translational level. In Xanthomonas campestris pv. campestris, insertional inactivation of MarR family transcription regulator HpaR (XC2827) resulted in significantly decrease in virulence and increase in the production of the extracellular proteases. Here, we reported that the genome of Xcc 8004 encodes nine MarR family transcription regulators. The MarR family transcription regulators, HpaR (XC2827) and XC0449, were heterologous expressed and purified. In vitro MST and Pull-down assay confirmed the physical interaction between HpaR and XC0449. Phenotypical assay determined that deletion of XC0449 resulted in substantial virulence attenuation. In vitro EMSA, in vivo qRT-PCR and GUS activity assay identified that HpaR and XC0449 coordinately act as the transcriptional activator to regulate the expression of the virulence-associated gene XC0705, and eventually control the bacterial virulence and the production of extracellular proteases.

The Gram-negative bacterium Lonsdalea populi causes a lethal disease known as bark canker on Populus × euramericana in China and Europe. Typical symptoms of bark canker include an abundant white-colored fluid, which oozes from the infected tissues. The availability of the genomic sequence of the bacterium provided the necessary resource to launch genome-scale investigations into the mechanisms fundamental to pathogenesis. Functional analyses of a diverse group of genes encoding virulence factors and components of signaling pathways indicate that successful bark infection depends on specific responses by the pathogen to various stresses, including oxidative stress. Although physiology of resistance is well studied, the molecular processes underlying the defense responses and the genetic basis of resistance to L. populi and in other poplar species remain largely unknown. Control of the disease has relied on chemical measures. Due to the genetic amenability of Lonsdalea and poplar, this pathosystem will become an important model system to unravel molecular mechanisms of bacterial pathogenicity on woody plants. Increased understanding of pathogenesis and signaling in the interaction will facilitate the management of this kind of poplar canker.

由Lonsdalea quercina subsp.populi引起的欧美杨溃疡病于2006年在国内首次发现,不同于其它病原菌造成的杨树溃疡病,该病害对欧美杨速生林的生长造成毁灭性破坏,已造成了严重的经济损失,该病原菌的致病分子机制尚不清楚.双组分系统是细菌重要的信号传递通路,在细菌的生长繁殖、逆境胁迫应答、环境适应以及病原菌致病过程中发挥重要作用.开展双组份系统研究将有助于解析欧美杨细菌性溃疡病菌的致病机制.本研究鉴定了欧美杨细菌性溃疡病菌L.quercina双组份孤儿反应调节基因LqRR2,并对其生物学功能进行了研究.通过同源重组获得了LqRR2基因的缺失突变体△LqRR2.表型测定结果显示,与野生型菌株相比,突变体△LqRR2在半固体培养基上的游动能力显著减弱,对欧美杨‘107杨’枝干的毒性也显著降低.但是,突变体的生长速率、生物膜形成能力以及胞外多糖产量较野生型无显著差别.此外,荧光定量PCR分析结果显示,游动性相关基因flgB、flgC和flgE的表达量在突变体中明显降低.综上所述,双组份调节蛋白编码基因LqRR2是欧美杨细菌性溃疡病菌L.quercina维持病原菌游动性和全毒性所必需的.

Poplar, which is a dominant species in plant communities distributed in the northern hemisphere, is commonly used as a model plant in forestry studies. Poplar production can be inhibited by infections caused by bacteria, including Lonsdalea quercina subsp. populi, which is a gram-negative bacterium responsible for bark canker disease. However, the molecular basis of the pathogenesis remains uncharacterized. In this study, we annotated the two-component signal transduction systems (TCSs) encoded by the L. quercina subsp. populi N-5-1 genome and identified 18 putative histidine kinases and 24 response regulators. A large-scale mutational analysis revealed that 19 TCS genes regulated bacterial virulence against poplar trees. Additionally, the deletion of kdpE or overexpression of kdpD resulted in almost complete loss of bacterial virulence. We observed that kdpE and kdpD formed a bi-cistronic operon. KdpD exhibited autokinase activity and could bind to KdpE (K d = 5.73 ± 0.64 μM). Furthermore, KdpE is an OmpR family response regulator. A chromatin immunoprecipitation sequencing analysis revealed that KdpE binds to an imperfect palindromic sequence within the promoters of 44 genes, including stress response genes Lqp0434, Lqp3037, and Lqp3270. A comprehensive analysis of TCS functions may help to characterize the regulation of poplar bark canker disease.

[Objective]Poplar bacterial canker caused by Lonsdalea quercina subsp.populi is a disease, which is serious harm to poplar industry.In this study, the biological function of the LqHK1 gene in L.quercina subsp.populi was investigated to provide viable knowledge for further understanding the pathogenic mechanism of pathogen.[Method]The bioinformatics method was used to indentify the two component systems LqHK1.The gene deletion mutant strain △LqHK1 was constructed by homologous recombination.Verifying PCR and Southern blot were used to investigate the biological characteristics of the mutant strains and their complementary strains,such as growth rate, motility, biofilm formation, and pathogenicity.At the same time, qRT-PCR was used to test the expression levels of the motility related genes flgB, flgC, flgE, and quantitatively analyze DNA content of poplar tissue pathogen in the vaccination site after pathogen inoculation.[Result]The two-component system gene LqHK1 was identified and the deletion mutant △LqHK1 had been obtained by homologous recombination.Phenotypic analysis showed that pathogenicity test on annual poplar branches was significantly less virulent than wide-type, while the complemented mutant HB LqHK1 restored the virulence to the wild-type level.The mutant had significantly less colonization in host than the wild type strain.Mutant strain had reduced swimming ability compared with the wild-type.The motility related genes flgB, flgC, flgE expression significantly decreased in mutant;forming ability LqHK1 mutant biofilms significantly decreased, but the growth rate of LqHK1 mutant strains had no significant difference from wild type.[Conclusion]Studies indicate that the bacterial poplar canker Lonsdalea quercina subsp.populi two-component signal transduction system LqHK1 gene is closely related to pathogenpathogenicity.