番茄细菌性斑点病菌无毒基因研究进展

番茄细菌性斑点病是影响番茄产量和品质的重要病害,Pseudomonas syringaepv.tomato(Pst)为其病原菌,其与番茄的互作系统是研究植物抗感病机理的典型模式系统。Pst存在2种无毒基因:avrPto和avrPtoB,它们编码的蛋白质均能与番茄抗性基因Pto编码的Ser-Thr蛋白激酶互作,符合Flor"基因对基因"学说。AvrPto和AvrPtoB在表达Pto的抗性植物中,与Pto互作,表现无毒功能,引发植物防御反应;而在缺失Pto的感病植物中,它们具有毒性,促进细菌的生长。本文综述了番茄细菌性斑点病菌无毒基因avrPto及avrPtoB的结构特点及其功能,这有助于了解病原物与植物的互作机制,对认识植物的感病性、抗病性以及植物防御反应都具有重要意义。     

Compost and compost water extract-induced systemic acquired resistance in cucumber and Arabidopsis

ABSTRACT A biocontrol agent-fortified compost mix, suppressive to several diseases caused by soilborne plant pathogens, induced systemic acquired resistance (SAR) in cucumber against anthracnose caused by Colletotrichum orbiculare and in Arabidopsis against bacterial speck caused by Pseudomonas syringae pv. maculicola KD4326. A peat mix conducive to soilborne diseases did not induce SAR. The population size of P. syringae pv. maculicola KD4326 was significantly lower in leaves of Arabidopsis plants grown in the compost mix compared to those grown in the peat mix. Autoclaving destroyed the SAR-inducing effect of the compost mix, and inoculation of the autoclaved mix with nonautoclaved compost mix or Pantoea agglomerans 278A restored the effect, suggesting the SAR-inducing activity of the compost mix was biological in nature. Topical sprays with water extract prepared from the compost mix reduced symptoms of bacterial speck and the population size of pathogenic KD4326 in Arabidopsis grown in the peat mix but not in the compost mix. The peat mix water extract applied as a spray did not control bacterial speck on plants grown in either mix. Topical sprays with salicylic acid (SA) reduced the severity of bacterial speck on plants in the peat mix but did not further reduce the severity of symptoms on plants in the compost mix. The activity of the compost water extract was heat-stable and passed through a 0.2-mum membrane filter. beta-1,3-Glucanase activity was low in cucumber plants grown in either mix, but when infected with C. orbiculare, this activity was induced to significantly higher levels in plants grown in the compost mix than in plants grown in the peat mix. Similar results were obtained for beta-D-glucuronidase (GUS) activity driven by a PR2 (beta-1,3-glucanase) gene promoter in transgenic Arabidopsis plants grown in the compost or peat mix. GUS activity was induced with topical sprays of the compost water extract or SA in plants not inoculated with the pathogen, suggesting that compost-induced disease suppression more than likely involved the potentiation of resistance responses rather than their activation and that compost-induced SAR differed from SAR induced by pathogens, SA, or compost water extract.     

组蛋白修饰调控植物水杨酸信号转导的研究进展

真核生物中组蛋白翻译后共价修饰直接影响染色质空间结构变化,调控相关基因表达,在植物胁迫应答过程中起重要作用。水杨酸(salicylic acid,SA)作为植物中关键信号分子,诱导多种病毒、真菌及细菌病害抗性。本文简要介绍了植物细胞对病原菌的感知、转导信号产生与防御系统激活机制,着重阐述了组蛋白甲基化、乙酰化、SUMO化修饰、组蛋白变体H2A.Z等如何参与调控水杨酸转导途径应答基因的转录表达。     

Beta-cyanoalanine synthase as a molecular marker for induced resistance by fungal glycoprotein elicitor and commercial plant activators

ABSTRACT The biocontrol agent Pythium oligandrum produces glycoprotein elicitor in the cell wall fraction, designated CWP, and induces resistance to a broad range of pathogens. To understand the mechanism of CWP-induced resistance to pathogens, gene expression at the early stage of CWP treatment in tomato roots was analyzed using a cDNA array. At 4 h after CWP treatment, 144 genes were up-regulated and 99 genes were down-regulated. In the 144 up-regulated genes, nine genes exhibited about eightfold increased expression. Analysis of the response of these nine genes to three commercial plant activators indicated that a high level of one gene, beta-cyanoalanine synthase gene (LeCAS) encoding hydrogen cyanide (HCN) detoxification enzyme, was stably induced in tomato roots by such treatment. However, expression of LeCAS was not significantly induced in tomato roots at 4 h by abiotic stresses, whereas only a very low level of induction of such expression by cold stress was observed. This LeCAS expression was also induced after exogenous treatment with a low level of 1-amino-cyclopropane-1-carboxylate as the precursor of ethylene, but not with either salicylic acid or methyl jas-monate. The induction of LeCAS expression in CWP-treated and plant activator-treated roots is likely to be caused by the detoxification of HCN during ethylene production. Transient activation of LeCAS expression caused by ethylene production in tomato roots may be a general phenomenon in fungal elicitor-induced and synthetic plant activator-induced resistance. LeCAS seems to be useful for screening possible novel plant activators for plant protection against pathogens.     

Systemic resistance to bacterial leaf speck pathogen in Arabidopsis thaliana induced by the culture filtrate of a plant growth-promoting fungus (PGPF) Phoma sp. GS8-1

The culture filtrate (CF) from the plant growth-promoting fungus Phoma sp. GS8-1 was found to induce systemic resistance in Arabidopsis thaliana against the bacterial leaf speck pathogen Pseudomonas syringae pv. tomato DC3000 (Pst), and the underlying mechanism was studied. Roots of A. thaliana were treated with CF from GS8-1, and plants expressed a clear resistance to subsequent Pst infection; disease severity was reduced, and proliferation of pathogen was suppressed. Various mutants of A. thaliana were used to test whether the CF induced resistance through one of the known signaling pathways: salicylic acid (SA), jasmonic acid (JA), and ethylene (ET). The CF was fully protective against Pst in Arabidopsis mutants jar1 and ein2 similar to wild-type plants. However, its efficacy was reduced in plants containing transgene NahG. Examination of systemic gene expression revealed that CF modulates the expression of SA-inducible PR-1, PR-2 and PR-5 genes, the JA/ET-inducible ChitB gene, and the ET-inducible Hel gene. Moreover, the expression of these genes was further enhanced upon subsequent stimulation after attack by Pst. Our data suggest that in addition to a partial requirement for SA, the signals JA and ET may also play a role in defense signaling in Arabidopsis.     

Plant responses to plant growth-promoting rhizobacteria

Non-pathogenic soilborne microorganisms can promote plant growth, as well as suppress diseases. Plant growth promotion is taken to result from improved nutrient acquisition or hormonal stimulation. Disease suppression can occur through microbial antagonism or induction of resistance in the plant. Several rhizobacterial strains have been shown to act as plant growth-promoting bacteria through both stimulation of growth and induced systemic resistance (ISR), but it is not clear in how far both mechanisms are connected. Induced resistance is manifested as a reduction of the number of diseased plants or in disease severity upon subsequent infection by a pathogen. Such reduced disease susceptibility can be local or systemic, result from developmental or environmental factors and depend on multiple mechanisms. The spectrum of diseases to which PGPR-elicited ISR confers enhanced resistance overlaps partly with that of pathogen-induced systemic acquired resistance (SAR). Both ISR and SAR represent a state of enhanced basal resistance of the plant that depends on the signalling compounds jasmonic acid and salicylic acid, respectively, and pathogens are differentially sensitive to the resistances activated by each of these signalling pathways. Root-colonizing Pseudomonas bacteria have been shown to alter plant gene expression in roots and leaves to different extents, indicative of recognition of one or more bacterial determinants by specific plant receptors. Conversely, plants can alter root exudation and secrete compounds that interfere with quorum sensing (QS) regulation in the bacteria. Such two-way signalling resembles the interaction of root-nodulating Rhizobia with legumes and between mycorrhizal fungi and roots of the majority of plant species. Although ISR-eliciting rhizobacteria can induce typical early defence-related responses in cell suspensions, in plants they do not necessarily activate defence-related gene expression. Instead, they appear to act through priming of effective resistance mechanisms, as reflected by earlier and stronger defence reactions once infection occurs.     

Defense gene expression patterns of three SAR-induced tomato cultivars in the field

Plant activation is an appealing disease management tool, avoiding some of the challenges of traditional chemical control by not directly impacting the pathogen. This study examined effects of acibenzolar-S-methyl (ASM), a plant activator that induces systemic acquired resistance, on defense response activation in three field-grown tomato cultivars in New York. Salicylic acid, ethylene and jasmonic acid-mediated responses were monitored by following expression of a marker gene for each signaling pathway using quantitative real-time PCR over the course of two ASM applications. ASM induced salicylic acid and ethylene, but not jasmonic acid-regulated gene expression in all cultivars tested. All three cultivars demonstrated a significantly stronger gene expression response relative to the untreated control following the second ASM application. Implications of these findings on management practices are discussed.     

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.     

Bioluminescence reporter assay system to monitor Arabidopsis MPK3 gene expression in response to infection by Botrytis cinerea

The Arabidopsis MPK3 gene product participates in disease resistance mediated by the MAP kinase cascade. The expression of the MPK3 gene is induced by pathogen inoculation and treatment with chemicals such as salicylic acid (SA) and methyl jasmonate (JA), but the detailed expression pattern of the MPK3 gene has been largely unknown. To investigate MPK3 gene expression in response to disease stress, we fused the MPK3 promoter to the firefly luciferase gene to create a real-time monitoring system for regulated gene expression in planta. The results of an in vivo reporter assay using transgenic Arabidopsis plants harboring MPK3::Fluc showed that the MPK3 promoter activity was induced by treatment with chemicals such as SA and benzo(1,2,3)-thiadiazole-7-carbothioic acid S-methyl ester (BTH), that induce defense gene expression. Inoculation with the fungal pathogen Botrytis cinerea resulted in systemic induction of MPK3::Fluc.     

No role for bacterially produced salicylic Acid in rhizobacterial induction of systemic resistance in Arabidopsis

ABSTRACT The role of bacterially produced salicylic acid (SA) in the induction of systemic resistance in plants by rhizobacteria is far from clear. The strong SA producer Pseudomonas fluorescens WCS374r induces resistance in radish but not in Arabidopsis thaliana, whereas application of SA leads to induction of resistance in both plant species. In this study, we compared P. fluorescens WCS374r with three other SA-producing fluorescent Pseudomonas strains, P. fluorescens WCS417r and CHA0r, and P. aeruginosa 7NSK2 for their abilities to produce SA under different growth conditions and to induce systemic resistance in A. thaliana against bacterial speck, caused by P. syringae pv. tomato. All strains produced SA in vitro, varying from 5 fg cell(-1) for WCS417r to >25 fg cell(-1) for WCS374r. Addition of 200 muM FeCl(3) to standard succinate medium abolished SA production in all strains. Whereas the incubation temperature did not affect SA production by WCS417r and 7NSK2, strains WCS374r and CHA0r produced more SA when grown at 33 instead of 28 degrees C. WCS417r, CHA0r, and 7NSK2 induced systemic resistance apparently associated with their ability to produce SA, but WCS374r did not. Conversely, a mutant of 7NSK2 unable to produce SA still triggered induced systemic resistance (ISR). The possible involvement of SA in the induction of resistance was evaluated using SA-nonaccumulating transgenic NahG plants. Strains WCS417r, CHA0r, and 7NSK2 induced resistance in NahG Arabidopsis. Also, WCS374r, when grown at 33 or 36 degrees C, triggered ISR in these plants, but not in ethylene-insensitive ein2 or in non-plant pathogenesis- related protein-expressing npr1 mutant plants, irrespective of the growth temperature of the bacteria. These results demonstrate that, whereas WCS374r can be manipulated to trigger ISR in Arabidopsis, SA is not the primary determinant for the induction of systemic resistance against bacterial speck disease by this bacterium. Also, for the other SAproducing strains used in this study, bacterial determinants other than SA must be responsible for inducing resistance.     

Studies on regulation of plant physiology by pesticides

Some agrochemicals have unique activities on plant, which modes of actions differ from those of herbicides and plant growth regulators. Because these induce useful and important phenotypic characteristics by activating physiological mechanisms in plant cell, understanding the underlying mechanism of their activities should be crucial for plant physiology and agriculture. As examples of such agrochemicals, studies on agrochemicals that activate the plant immune systems or root elongation, are described. Plant activators, inducers of systemic acquired resistance, were divided into two types, acting on upstream and downstream of salicylic acid (SA) biosynthesis, respectively. They have been useful research tools to clarify the regulation mechanism of SA-mediated disease resistance and to investigate another type of disease resistance mechanism mediated by brassinosteroids. By analyzing the roles of phytohormones in the isoprothiolane-induced root elongation indicated a positive effect of jasmonic acid and ethylene on primary root elongation. These types of research, categorized to one of chemical biology, would provide novel insight into plant physiology, which also contribute to control of crops.     

Reduction of Bacterial Speck (Pseudomonas syringae pv. tomato) of Tomato by Combined Treatments of Plant Growth-promoting Bacterium, Azospirillum brasilense, Streptomycin Sulfate, and Chemo-thermal Seed Treatment

Inoculation of tomato seeds with the plant growth-promoting bacterium Azospirillum brasilense, or spraying tomato foliage with A. brasilense, streptomycin sulfate, or commercial copper bactericides, separately, before or after inoculation with Pseudomonas syringae pv. tomato, the casual agent of bacterial speck of tomato, had no lasting effect on disease severity or on plant height and dry weight. Seed inoculation with A. brasilense combined with a single streptomycin foliar treatment and two foliar bactericide applications at 5-day intervals (a third or less of the recommended commercial dose) reduced disease severity in tomato seedlings by over 90% after 4 weeks, and significantly slowed disease development under mist conditions. A. brasilense did not induce significant systemic resistance against the pathogen although the level of salicylic acid increased in inoculated plants. Treatment of tomato seeds that were artificially inoculated with P. syringae pv. tomato, with a combination of mild chemo-thermal treatment, A. brasilense seed inoculation, and later, a single foliar application of a copper bactericide, nearly eliminated bacterial leaf speck even when the plants were grown under mist for 6 weeks. This study shows that a combination of otherwise ineffective disease management tactics, when applied in concert, can reduce bacterial speck intensity in tomatoes under mist conditions.     

Comparison of induced resistance activated by benzothiadiazole, (2R,3R)‐butanediol and an isoparaffin mixture against anthracnose of Nicotiana benthamiana

Resistance in Nicotiana benthamiana against anthracnose caused by the hemibiotrophic fungus Colletotrichum orbiculare was activated by benzothiadiazole (BTH), (2R,3R)­butanediol or PC1, an isoparaffin‐based mixture. In inoculation experiments, BTH, (2R,3R)­butanediol and PC1 reduced the number of lesions per leaf area caused by C. orbiculare by 98%, 77% and 81%, respectively. Foliar application of BTH induced expression of genes for the acidic pathogenesis‐related (PR) proteins, NbPR‐1a, NbPR‐3Q and acidic NbPR‐5. In contrast, soil application of (2R,3R)­butanediol or PC1 primed expression of genes for the basic PR proteins, NbPRb‐1b, basic NbPR‐2 and NbPR‐5dB. These results are consistent with the activation of salicylic‐acid‐dependent systemic acquired resistance (SAR) by BTH and that of jasmonate/ethylene‐dependent induced systemic resistance (ISR) by (2R,3R)­butanediol or PC1, and show that (2R,3R)­butanediol and PC1 can affect gene expression similarly to plant growth‐promoting rhizobacteria. However, the effects of (2R,3R)‐butanediol and PC1 were not identical. In addition to priming, (2R,3R)‐butanediol induced expression of basic NbPR‐2, whereas PC1 treatment induced expression of both NbPRb‐1b and basic NbPR‐2. Although a number of microbial products, such as (2R,3R)­butanediol, have been shown to produce ISR, this is the first demonstration that an isoparaffin‐based mixture, not derived from a microorganism, can produce ISR.     

Transcriptome profile of acibenzolar-S-methyl-induced genes in tomato suggests a complex polygenic effect on resistance to Phytophthora infestans

Induced resistance by chemicals such as acibenzolar-S-methyl -ASM (commercialized as Actigard by Syngenta Inc) mimics the biological activation of systemic acquired resistance (SAR). ASM takes the place of salicylic acid (SA) in the SAR signal pathway inducing the same molecular markers and range of resistance. The goal of our work was to understand the downstream molecular events by which ASM confers resistance to Phytophthora infestans in tomatoes. To accomplish this goal we assayed gene expression in ASM-treated plants using a microarray with more than 12,000 tomato ESTs. As many as 300 genes were responsive to ASM. Of these, 117 were detected in most of the biological replications. Basal defense associated genes as well as SAR and disease resistance genes (R-like) involved in induced resistance and effector-triggered immunity were highly expressed. We attempted to determine the phenotype of 13 of these genes by virus induced gene silencing (VIGS). These 13 genes were selected on the basis of previous implication in plant defense response and by reliability of induction by ASM. VIGS was partially successful for three of the 13 genes, but this partial silencing did not lead to a significant reduction in the effect of ASM. The ethylene pathway was also activated in response to ASM, but a tomato mutant not responsive to ethylene remained responsive to ASM. It seems most likely that the ASM effect is complex and polygenic, depending on the effect of several genes.     

BTH诱导花椰菜对菌核病的抗性研究

 利用苯并噻二唑BTH处理菌核病抗性不同的花椰菜品种幼苗, 采用营养生长期活体叶片菌丝块接种鉴定法评价菌核病抗性诱导效果,结果表明经BTH处理的植株菌核病病情指数明显下降, 对感病品种和抗病品种的诱抗效果分别达到81.5%和63.8%。对于花椰菜重要的防御酶活性变化研究结果表明,BTH诱导处理的花椰菜植株过氧化物酶（POD）、抗坏血酸酶( SOD )、过氧化氢酶（CAT）、苯丙氨酸解氨酶（PAL）和多酚氧化酶( PPO)的活性均有所提高。同时病程相关蛋白几丁质酶和β-1,3-葡聚糖酶的活性也增加。 利用半定量RT-PCR方法检测防御反应基因表达,结果表明BTH诱导首先激发了植株 PR-1等基因参与的水杨酸信号传导防御反应途径的发生,同时PDF1.2 基因的上调表达说明BTH诱导也影响了茉莉酸信号传导途径。     

Menadione sodium bisulphite: a novel plant defence activator which enhances local and systemic resistance to infection by Leptosphaeria maculans in oilseed rape

Pretreatment of the first true leaves of oilseed rape plants ( Brassica napus cv. Bristol) with menadione sodium bisulphite (MSB) locally and systemically induced resistance, as shown by reduced lesion size and number, to infection by the fungal pathogen Leptosphaeria maculans , the causal agent of stem canker. Using a known systemic activator of salicylic acid-dependent PR-1 induction, acibenzolar- S -methyl (BTH; S -methylbenzo[1,2,3]thiadiazole-7-carbothiate) as a comparison, real-time PCR expression analysis of genes encoding a pathogenesis-related protein 1 ( PR-1 ) and an ascorbate peroxidase ( APX ) demonstrated a systemic enhancement of APX expression in MSB-pretreated plants, with no effect on PR-1 expression, suggesting augmented reactive oxygen species production in MSB-pretreated plants. The results demonstrate MSB to be an effective resistance activator in oilseed rape, and potentially useful for the control of stem canker.     

The tomato Pto gene confers resistance to Pseudomonas floridensis,an emergent plant pathogen with just nine type III effectors

A newly discovered bacterial species, Pseudomonas floridensis, has emerged as a pathogen of tomato in Florida. This study compares the virulence and other attributes of P. floridensis to Pseudomonas syringae pv. tomato, which causes bacterial speck disease of tomato. Pseudomonas floridensis reached lower population levels in leaves of tomato as compared to the P. syringae pv. tomato strains DC3000 and NYT1. Analysis of the genome sequence of the P. floridensis type strain GEV388 revealed that it has just nine type III effectors including AvrPtoB_GEV388, which is 66% identical to AvrPtoB in DC3000. Five of these effectors have been previously reported to be members of a ‘minimal effector repertoire’ required for full DC3000 virulence on Nicotiana benthamiana; however, GEV388 grew poorly on leaves of this plant species compared to the DC3000 minimal effector strain. The tomato Pto gene recognizes AvrPtoB in race 0 P. syringae pv. tomato strains, thereby conferring resistance to bacterial speck disease. Pto was also found to confer resistance to P. floridensis, indicating this gene will be useful in the protection of tomato against this newly emerged pathogen.     

Activation of Defense Pathways: Synergism between Reactive Oxygen Species and Salicylic Acid and Consideration of Field Applicability

Treatment of tobacco with a mixture containing reactive oxygen species (ROS) and salicylic acid (SA) provided greater protection of tobacco against infection by Pseudomonas syringae pv. tabaci than either treatment alone. Synergism in expression from the promoter of the defense gene PR-1a was also observed. Although the ROS hydrogen peroxide and peracetic acid were poor inducers alone, they enhanced the level of -glucuronidase (GUS) activity expressed from the PR-1a promoter when applied with SA to a transgenic plant bearing a PR-1a::GUS fusion. PR-1a expression was not correlated with increased cell death as determined by Evans blue staining. There was no effect on the timing at which expression was increased by the mixture compared with the separate treatments. The mixture of hydrogen peroxide and SA partially mimicked the effect of a commercial product Oxycom that has field efficacy in improving plant performance. Repetitive applications of Oxycom enhanced expression from the PR-1a promoter and the production of the PR-1 protein. Enhanced activity occurred systemically both from aerial applications to single leaves and from root drenches. Root application strongly promoted veinal expression for the PR-1a promoter compared with confluent production in leaves of sprayed seedlings. Application methods and timing may aid in the success of activators of systemic acquired resistance in field conditions.