激活蛋白PeaT1诱导烟草对TMV的系统抗性

枯斑三生烟草(Samsun-NN)经激活蛋白PeaT1诱导后接种烟草花叶病毒(Tobacco mosaic virus,TMV),对TMV产生了明显的系统获得抗性,枯斑抑制率达54.15%,枯斑大小也受到一定程度的限制。研究结果表明,PeaT1处理烟草植株下位三片叶不同时间后,其上部叶片中PPO、POD和PAL 3种抗病防御酶活性均比对照提高,第4 d酶活性达到最高值。实时荧光定量PCR检测结果显示,经PeaT1诱导4 d后烟草叶片中抗病相关基因PR1a、PR1b、NPR1在转录表达水平上较未诱导对照都有不同程度的上调。由以上结果我们推测PeaT1诱导烟草产生了系统获得抗性,本研究为进一步阐明PeaT1诱导植物抗病的信号传导途径奠定了基础。     

内生细菌EBS05对烟草诱导抗性的信号转导途径研究

 樟树内生枯草芽胞杆菌EBS05是一株对多种植物病原菌具有较强拮抗活性,并能诱导烟草系统抗性的生防菌株。本文以缺失Surfactin A合成相关基因的突变菌株EBS05^T为材料,研究了内生细菌EBS05对烟草诱导系统抗性的激发子及其信号转导途径。结果表明,菌株EBS05产生的Surfactin A是诱导烟草对TMV系统抗性的有效激发子;Surfactin A诱导处理后,SA信号转导途径下游的关键调节基因NPR1首先被激活,并持续超量表达,进而触发PR1b和PR1a基因持续超量表达,表明Surfactin A诱导烟草对TMV的系统抗性是通过激活SA信号转导途径实现的。同时,Surfactin A诱导处理后24~72 h,JA/ET信号转导途径调节基因PDF1.2被激活,且超量表达,表明在Surfactin A诱导烟草对TMV系统抗性的信号转导过程中,可能存在SA信号途径和JA/ET信号途径的交叉协同作用。     

枯草芽孢杆菌PTS-394诱导番茄对灰霉病的系统抗性

本文研究了枯草芽孢杆菌PTS-394对番茄的防御相关酶活性、抗病信号转导通路的标志基因表达的诱导情况和诱导抗病性对灰霉病的防治效果。结果显示,菌株PTS-394灌根番茄后,在24~72 h内番茄顶端叶片中PAL、PPO、POX、LOX的活性都有不同程度的持续增加,且72 h时达到最高峰值,随后在96 h下降,与对照相比差异显著;此外,番茄抗病信号通路节点基因NPR1和水杨酸(SA)信号通路激发的防卫基因PR-1a,在24~72 h得到了显著持续高表达。以上结果表明,利用菌株PTS-394灌根番茄后,能够诱导植株产生系统抗病性。菌株PTS-394灌根番茄后48 h,离体叶片挑战接种番茄灰霉菌,结果显示,菌株PTS-394处理的番茄叶片病斑面积仅为对照处理的50%,防控效果达47.1%;温室盆栽试验显示,菌株PTS-394处理后对番茄灰霉的防治效果为58.2%。综上所述,枯草芽孢杆菌PTS-394灌根番茄后,可以触发番茄植株系统性的抗病性,增强植株免疫能力。     

Involvement of jasmonic acid signalling in bacterial wilt disease resistance induced by biocontrol agent Pythium oligandrum in tomato

When the biocontrol agent Pythium oligandrum (PO) colonizes the rhizosphere, it suppresses bacterial wilt disease in tomato (Solanum lycopersicum cv. Micro‐Tom) caused by Ralstonia solanacearum, and a homogenate of its mycelia exhibits elicitor activity, inducing an ethylene (ET)‐dependent defence response in Micro‐Tom. Since salicylic acid (SA) and jasmonic acid (JA) play an important role in plant defence responses to pathogens, the involvement of SA‐ and JA‐dependent signal transduction pathways in resistance to R. solanacearum was investigated in tomato roots treated with a mycelial homogenate of PO. Bacterial wilt disease was also suppressed in tomato cv. Moneymaker treated with the PO homogenate. However, the SA‐inducible PR‐1(P6) gene was not up‐regulated in either Micro‐Tom or Moneymaker. SA did not accumulate in homogenate‐treated roots in comparison with distilled water‐treated controls, even 24 h after inoculation. Induced resistance against R. solanacearum was not compromised in SA‐non‐accumulating NahG transgenic plants treated with the PO homogenate. On the other hand, the expression of the JA‐responsive gene for the basic PR‐6 protein was induced in both tomato cultivars treated with the PO homogenate. Furthermore, quantitative disease assays showed that the induced resistance against R. solanacearum was compromized in PO homogenate‐treated jai1‐1 mutant plants defective in JA signalling. These results indicated that the JA‐dependent signalling pathway is required for PO‐induced resistance against R. solanacearum in tomato.     

真核表达产物Y3诱导烟草对烟草花叶病毒的抗性研究

从毛头鬼伞Coprinus comatus中提取的碱性糖蛋白Y3可以降低烟草花叶病毒（TMV）的侵染。克隆获得Y3蛋白cDNA后与真核表达载体pPIC-9k连接,重组载体pPIC-9k-Y3成功电转化入毕赤酵母Pichia pastoris后,转化子在28℃、250 r/min培养条件下,使用1.0%甲醇诱导表达6 d,成功实现了Y3蛋白的真核表达。300 μg/mL浓度的Y3蛋白对普通烟进行诱导处理后摩擦接种TMV结果表明,诱导处理后烟草体内除过氧化物酶POD外,多酚氧化酶PPO、苯丙氨酸解氨酶PAL和β-1,3葡聚糖酶活性都有不同程度提高。蛋白处理24 h后植株中PPO活性达到最大,为对照组的2倍。利用实时荧光定量PCR （qRT-PCR）研究抗性相关基因表达情况,Y3诱导后烟草植株碱性病程相关基因1（PR1-b）、病程相关基因非表达子1（NPR1）、PAL在转录水平上均显著上调（P<0.05）。推测真核表达产物Y3蛋白可通过提高水杨酸信号途径相关防御酶活性以及抗病基因转录来诱导烟草对TMV产生系统抗性,为Y3蛋白作为生物农药开发提供理论基础。     

植物抗病激活蛋白harpinXooc防治水稻病害的研究

植物抗病激活蛋白harpinXooc由水稻条斑病细菌hrp基因簇中hpa1基因编码。用构建于表达载体pET21a（＋）上的hpa1诱导表达产物harpinXooc处理烟草，可激发烟草产生过敏反应，以及激活与烟草抗病信号途径相关的基因PR-1a、hin1和hsr203J的表达；处理水稻后，NPR1、OsPR1a、OsPR1b和PAL被激活。表明harpinXooc蛋白与植物互作后，通过水杨酸信号传导途径激活病程相关蛋白等防卫反应基因的转录表达，从而使植物产生系统获得抗病性。harpinXooc经加工后制成含量达1％的可溶性微颗粒制剂在水稻上进行应用，试验结果显示，harpinXooc蛋白可诱导水稻产生抗病性，防治水稻稻瘟病效果与杀菌剂稻瘟必克（三环唑）相当，防治水稻纹枯病和稻曲病效果与井冈霉素效果相当。对水稻增产的效果主要表现在增加粒实重上，增产达6％以上。     

Inverse effects of Arabidopsis NPR1 gene on fusarium seedling blight and fusarium head blight in transgenic wheat

In this study, the Arabidopsis thaliana NPR1 (non‐expressor of PR genes) gene was integrated into an elite wheat cultivar, and the response of the transgenic wheat expressing NPR1 to inoculation with Fusarium asiaticum was analysed. With seedling inoculation, the transgenic lines showed significantly increased fusarium seedling blight (FSB) susceptibility, whereas floret inoculation resulted in enhanced fusarium head blight (FHB) resistance. Quantitative real‐time PCR revealed that expression of two defence genes, PR3 and PR5, was associated with susceptible reactions to FSB and FHB, whereas the PR1 gene was activated in resistance responses. This inverse modulation by the constitutively expressed NPR1 gene suggests that NPR1 has a bifunctional role in regulating defence responses in plants. Therefore, it is unsuitable for improving overall resistance to FSB and FHB in wheat.     

毁灭炭疽菌Colletotrichum destructivum诱抗蛋白对烟草抗病性的诱导

为明确毁灭炭疽菌Colletotrichum destructivum诱抗蛋白诱导烟草的抗病性及其作用,采用喷雾、摩擦接种方法及RT-PCR技术研究了诱抗蛋白的预防保护作用,以及烟草悬浮细胞经诱导后过氧化物酶(POD)、多酚氧化酶(PPO)、苯丙氨酸解氨酶(PAL)活性及脯氨酸(Pro)含量和病程相关基因表达的变化。结果表明,接种3、5和7 d后,该诱抗蛋白对烟草炭疽病的诱抗效果分别为58.00%、48.99%和49.65%,对烟草白粉病的诱抗效果分别为83.26%、80.76%和78.60%,并可以抑制烟草普通花叶病毒的复制及在寄主体内的扩增;经诱抗蛋白处理后,烟草悬浮细胞POD、PPO、PAL活性及Pro含量明显提高;诱抗蛋白能够诱导烟草病程相关蛋白基因PR-1a、PR-1b以及抗病信号传导途径关键基因NPR1的表达。表明毁灭炭疽菌诱抗蛋白可诱导烟草产生抗病性,可能与烟草悬浮细胞中POD、PAL、PPO的活性及Pro的含量提高以及相关病程基因表达有关。     

β-葡寡糖诱导植物抗病性的研究进展

β-葡寡糖作为一种植物激发子,可高效诱导植物产生抗病性,因此被普遍认为是一种病原物相关的分子模式.其作用的发挥主要是通过与细胞膜上的受体相互识别,引起受体构象改变产生跨膜信号,再经过一系列的胞内信号传导,调控防卫基因的表达,积累次生代谢产物,诱导植物抗性来实现.诱抗活性不仅受到寡糖聚合度和化学修饰基团的影响,而且植物对于结构上有差异的β-葡寡糖激发子的识别也是大相径庭.本文就β-葡寡糖诱导植物产生抗病性的研究进展进行了综述.     

Suppressive effect of abscisic acid on systemic acquired resistance in tobacco plants

Recent studies have indicated that the phytohormone abscisic acid (ABA), induced in response to a variety of environmental stresses, plays an important role in modulating diverse plant–pathogen interactions. In Arabidopsis thaliana, we previously clarified that ABA suppressed the induction of systemic acquired resistance (SAR), a plant defense system induced by pathogen infection through salicylic acid (SA) accumulation. We investigated the generality of this suppressive effect by ABA on SAR using tobacco plants. For SAR induction, we used 1,2-benzisothiazole-3(2H)-one 1,1-dioxide (BIT) and benzo(1,2,3)thiadiazole-7-carbothioic acid S-methyl ester (BTH) that activate upstream and downstream of SA in the SAR signaling pathway, respectively. Wild-type tobacco plants treated with BIT or BTH exhibited enhanced disease resistance against Tobacco mosaic virus (TMV) and tobacco wildfire bacterium, Pseudomonas syringae pv. tabaci (Pst), however, which was suppressed by pretreatment of plants with ABA. Pretreatment with ABA also suppressed the expression of SAR-marker genes by BIT and BTH, indicating that ABA suppressed the induction of SAR. ABA suppressed BTH-induced disease resistance and pathogenesis-related (PR) gene expression in NahG-transgenic plants that are unable to accumulate SA. The accumulation of SA in wild-type plants after BIT treatment was also suppressed by pretreatment with ABA. These data suggest that ABA suppresses both upstream and downstream of SA in the SAR signaling pathway in tobacco.     

Systemic resistance induced in Arabidopsis thaliana by Trichoderma asperellum SKT-1, a microbial pesticide of seedborne diseases of rice

BACKGROUND: Trichoderma asperellum SKT-1 is a microbial pesticide of seedborne diseases of rice. To investigate the mechanisms of disease suppression in SKT-1, the ability to induce systemic resistance by SKT-1, or its cell-free culture filtrate (CF), was tested using Arabidopsis thaliana Col-0 plants. RESULTS: Both SKT-1 and its CF elicit an induced systemic resistance against the bacterial leaf speck pathogen Pseudomonas syringae pv. tomato DC3000 in Col-0 plants. Involvement of plant hormones in the induced resistance by SKT-1 and CF was assessed using Arabidopsis genotypes such as the jasmonic acid (JA)-resistant mutant jar1, the ethylene (ET)-resistant mutant etr1, the plant impaired in salicylic acid (SA) signalling transgenic NahG and the mutant npr1 impaired in NPR1 activity. In soil experiments using SKT-1, no significant disease suppression effect was observed in NahG transgenic plants or npr1 mutant plants. Expression levels of SA-inducible genes such as PR-1, PR-2 and PR-5 increased substantially in the leaves of Col-0 plants. Expression levels of JA/ET-induced genes such as PDF1.2a, PR-3, PR-4 and AtVsp1 were also induced, but the levels were not as high as for SA-inducible genes. In a hydroponic experiment using CF from SKT-1, all Arabidopsis genotypes showed an induced systemic resistance by CF and increased expression levels of JA/ET- and SA-inducible genes in leaves of CF-treated plants. CONCLUSION: The SA signalling pathway is important in inducing systemic resistance to colonisation by SKT-1, and both SA and JA/ET signalling pathways combine in the signalling of induced resistance by CF. These results indicate that the response of A. thaliana is different from that found in root treatments with barley grain inoculum and CF from SKT-1. Copyright © 2011 Society of Chemical Industry     

Salicylic Acid Suppresses Potato virus Y Isolate N:O-Induced Symptoms in Tobacco Plants

ABSTRACT The effects of salicylic acid (SA) and 1-aminocyclopropane-1-carboxylic acid (ACC) on the systemic development of symptoms induced by a severe isolate of Potato virus Y group N:O (PVY(N:O)) in tobacco were investigated. Upon inoculation, the systemic development of symptoms in tobacco plants could be divided into three stages: virus incubation stage, rapid symptom-progress stage, and partial recovery and symptom-shifting stage. Treatment of seedlings with SA delayed the virus-induced necrosis in stems by 1 to 2 days. SA, not ACC, also significantly suppressed the symptom severity in stems. However, neither SA nor ACC treatment affected the partial recovery phenotype exhibited in the latterly emerged upper parts of the plants. Further analysis indicated that the accumulation of PVY was retarded by SA at the early stage of infection, and the effects were more profound in stems than leaves. Peroxidase (POX) activity and pathogenesis-related (PR) genes PR-1a and PR-1b were enhanced by PVY infection. SA not only increased POX activity in stems and PR genes in stems and leaves of mock-inoculated plants, but also elevated the activity of POX in both leaves and stems and the expression of PR-1a in leaves of PVY-infected plants. Together, the results suggest that systemic acquired resistance plays a key role in suppressing PVY(N:O)-induced symptom development through SA-mediated and ethylene-independent pathways. The symptom suppression was correlated with reduced replication/ accumulation of virus at the early stage of infection. The results also suggest that neither SA nor ethylene plays a role in the recovery phenotype.     

Pseudomonas aeruginosa 7NSK2-induced Systemic Resistance in Tobacco Depends on in planta Salicylic Acid Accumulation but is not Associated with PR1a Expression

Root colonization by rhizobacteria can induce a systemic resistance in plants that is phenotypically similar to systemic acquired resistance induced by a localized pathogen infection. We used the tobacco–tobacco mosaic virus model to investigate whether the systemic resistance induced by the rhizobacterium Pseudomonas aeruginosa 7NSK2 is mediated by the systemic acquired resistance signal transduction pathway. Experiments with nahG-transformed tobacco revealed that Pseudomonas aeruginosa 7NSK2-induced resistance depended on in planta salicylic acid accumulation for its expression but not for its induction and is, in this respect, similar to systemic acquired resistance. However, Pseudomonas aeruginosa 7NSK2-induced resistance was, unlike systemic acquired resistance, not associated with PR1a expression at the time of challenge with tobacco mosaic virus. This suggests that Pseudomonas aeruginosa 7NSK2 treatment would only potentiate defense gene expression in systemic tissue, which would also explain why its level of resistance is lower than in case of systemic acquired resistance. Because we demonstrated that induced resistance by Pseudomonas aeruginosa 7NSK2 exclusively depends on the production of salicylic acid by this strain our conclusions might also account for other salicylic acid-producing and resistance-inducing rhizobacteria.     

心叶烟NPR1基因全长cDNA序列的克隆及表达分析

 NPR1(non-expressor of pathogenesis-related gene 1)基因在拟南芥系统获得抗性中起着关键作用,可调控拟南芥植株广谱抗性的发生。本文报道了从心叶烟中克隆NPR1同源基因(NgNPR1)及其表达特性的研究结果。NgNPR1 cDNA全长2253 bp,编码588个氨基酸。将NgNPR1基因组全长与cDNA序列进行比对发现,NgNPR1基因组DNA含有4个外显子和3个内含子。Southern杂交分析表明,在心叶烟基因组中NgNPR1为单拷贝基因。采用绿色荧光蛋白在洋葱表皮瞬时表达的试验,证明了NgNPR1蛋白在水杨酸诱导时会从细胞质转运到细胞核中。Northern杂交分析发现,NgNPR1基因可以被与植物抗病相关的信号分子如水杨酸、茉莉酸甲酯、过氧化氢和乙烯所诱导。进一步研究发现,植物病原物如赤星病菌、青枯病菌和烟草花叶病毒对心叶烟植株的侵染也会使NgNPR1表达量增加。这些结果表明,NgNPR1基因在心叶烟植株抵御病原物侵染过程中可能起着重要作用。     

植物系统性获得抗病性的产生机理和途径

坏死型病原物侵染或某些生化制剂诱导处理后,植株未受侵染或处理部位产生对随后病原物侵染的抗性,称为植物系统性获得抗性,ＳＡＲ具有抗性表现系统、持久、抗病对象广谱三大特点。坏死型病原物侵染或某些生化制剂处理后,植株受处理部位迅速产生系统性信号,经韧皮部传导到未侵染或处理部位,诱发ＳＡＲ基因表达。水杨酸是诱发ＳＡＲ的系统性信号之一。此外,上部非处理部位处于敏化状态,能更迅速有效地产生针对挑战接种病原物的     

Salicylic Acid Induces Resistance to Alternaria solani in Hydroponically Grown Tomato

ABSTRACT Alternaria solani is the causal agent of early blight disease in tomato and is responsible for significant economic losses sustained by tomato producers each year. Because salicylic acid (SA) is an important signal molecule that plays a critical role in plant defense against pathogen invasion, we investigated if the exogenous application of SA would activate systemic acquired resistance (SAR) against A. solani in tomato leaves. The addition of 200 muM SA to the root system significantly increased the endogenous SA content of leaves. Free SA levels increased 65-fold over basal levels to 5.85 mug g(-1) fresh weight (FW) after 48 h. This level of SA had no visible phytotoxic effects. Total SA content (free SA + SA-glucose conjugate) increased to 108 mug g(-1) FW after 48 h. Concomitant with elevated SA levels, expression of the tomato pathogenesis-related (PR)-1B gene was strongly induced within 24 h of the addition of 200 muM SA. PR-1B expression was still evident after 48 h; however, PR-1B induction was not observed in plants not receiving SA treatment. Challenge inoculation of SA-treated tomato plants using conidia of A. solani resulted in 83% fewer lesions per leaf and a 77% reduction in blighted leaf area as compared with control plants not receiving SA. Our data indicate that root feeding 200 muM SA to tomato plants can (i) significantly elevate foliar SA levels, (ii) induce PR-1B gene expression, and (iii) activate SAR that is effective against A. solani.