Physiological response of Bacillus cereus C1L-induced systemic resistance in lily against Botrytis leaf blight

Bacillus cereus C1L has been demonstrated to induce systemic disease resistance against Botrytis elliptica in lily. The objective of this study was to investigate physiological responses of B. cereus C1L-triggered systemic resistance in lily cv. Star Gazer against B. elliptica. By histological and biochemical analyses, leaves inoculated with B. elliptica displayed cell death, H₂O₂ accumulation and lignin deposition. As plants were elicited with B. cereus C1L, cell death, H₂O₂ accumulation and lignin deposition in leaves caused by B. elliptica infection were suppressed, revealing that suppression of oxidative burst might be associated with B. cereus C1L-induced systemic resistance. In reactive oxygen species inhibitors assays, B. elliptica-caused lesion numbers and H₂O₂ accumulation in lily leaves were significantly reduced as leaves were pretreated with catalase or diphenylene iodonium. Furthermore, the expression of LsGRP1 and LsPsbR in leaves elicited with B. cereus C1L and inoculated with B. elliptica was decreased. The same expression pattern was also observed in leaves pretreated with catalase or diphenylene iodonium and inoculated with B. elliptica. These results suggest that B. cereus C1L-induced systemic resistance may be related to suppression or alleviation of oxidative stress and cell death of lily caused by B. elliptica.     

Identification of wheat blue dwarf phytoplasma effectors targeting plant proliferation and defence responses

The identification of effectors from pathogenic microbes is one of the most important subjects for elucidating infection mechanisms. Wheat blue dwarf (WBD) phytoplasma causes dwarfism, witches' broom, and yellow leaf tips in wheat plants, resulting in severe yield loss in northwestern China. In this study, 37 candidate effector proteins were transiently expressed in Nicotiana benthamiana. Plants expressing the SAP11‐like protein SWP1 exhibited typical witches' broom. Interestingly, another protein, SWP11, induced both cell death and defence responses, including H₂O₂ accumulation and callose deposition. Analysis by qRT‐PCR was used to show that a marker gene of the hypersensitive response, HIN1, and three pathogenesis‐related genes, PR1, PR2 and PR3, were significantly up‐regulated in leaves of N. benthamiana expressing SWP11. In addition, SWP12 and SWP21 (TENGU‐like) were shown to suppress SWP11‐, BAX‐, and/or INF1‐induced cell death. These results indicated that SWP21 has a distinct role in virulence compared with TENGU and that WBD phytoplasma possesses effectors that target plant proliferation and defence responses. The ability of these effectors to trigger or suppress plant immunity provides new insights into the phytoplasma–plant interaction.     

盐胁迫下交替呼吸对二斑叶螨侵染叶片及其间接防御的影响

为探索在遭受盐胁迫时二斑叶螨Tetranychus urticae Koch侵染叶片的交替呼吸途径的生理学功能,在实验室条件下研究了NaCl以及水杨基氧肟酸对二斑叶螨侵染叶片的间接防御反应,以及对过氧化氢含量和细胞死亡水平的影响。200 mmol/L NaCl胁迫没有明显影响二斑叶螨侵染叶片对智利小植绥螨Phytoseiulus persimilis的吸引作用,也没有提高二斑叶螨侵染叶片中过氧化氢含量和细胞死亡的水平,但显著提高了侵染叶片中交替呼吸途径的水平。200 mmol/L NaCl和1mmol/L水杨基氧肟酸的复合处理则显著降低了二斑叶螨侵染叶片对捕食螨的吸引作用,并提高了过氧化氢含量和细胞死亡的水平。研究表明,当二斑叶螨侵染叶片遭受盐胁迫时,交替呼吸途径不仅有助于维持植物的间接防御反应,而且缓解了植物体内的氧化压力和细胞死亡。     

大豆花叶病毒引发大豆不同症状的生理特性比较

以接种3个SMV株系后分别表现无症状、花叶和坏死症状的大豆品系浙A8901为材料,研究3类症状叶片的生理指标差异,包括细胞超微结构、H_2O_2积累、水杨酸(SA)含量、光合作用及细胞病毒含量。结果表明:无症状叶片细胞中除叶绿体片层结构轻微扭曲外,其它细胞结构未见异常;CeCl_3标记H_2O_2电镜观察,发现H_2O_2仅在细胞壁外侧少量积累;在接种1d后SA含量较对照显著升高。花叶症状叶片细胞中出现髓鞘状结构和多泡体结构,核膜边缘有染色质凝集现象;细胞壁外侧观察到较多H_2O_2的积累;SA含量在接种后第2 d较对照显著增加。坏死症状叶片中细胞变形严重,细胞器解体,在细胞壁外侧和细胞质中均观察到H_2O_2的大量积累;接种后第2 d SA含量较对照显著增加。三类症状叶片的光合速率、最大光能转化效率(Fv/Fm)和实际光量子效率(ΦPSⅡ)的测定发现,与对照相比均无显著变化。     

The effect of hydrogen peroxide on the viability of tomato cells and of the fungal pathogen Cladosporium fulvum

An oxidative burst was previously demonstrated to be induced in tomato plants by race specific elicitors of the fungal pathogen Cladosporium fulvum . The in planta levels of H₂O₂estimated to occur during elicitor treatment, were compared with the levels required to show toxicity to host cells and to the fungal pathogen. Injection of Cf-9 tomato leaves with 100 m m H₂O₂caused an insignificant degree of necrosis and 1m H₂O₂was required to cause complete leaf necrosis comparable to that induced by the AVR9 elicitor. Assays with Cf-5 tomato cell suspensions confirmed the low toxicity of H₂O₂to tomato cells but, as expected, the addition of Fe²⁺with H₂O₂(or with intercellular fluids containing AVR5 elicitor) enhanced cell death as determined by the Evans Blue assay. Germination and germ tube growth of conidia of C. fulvum were significantly retarded by 4–5 m m H₂O₂, and at higher concentrations, death of germ tubes was observed (ED₅₀=22 m m), as determined by the fluorescein diacetate assay. The addition of Fe²⁺with H₂O₂had little effect on fungal growth or viability in vitro . These results suggest that the amount of H₂O₂accumulating during an elicitor-induced response in leaves may be sufficient to affect fungal colonization but not to affect viability of host cells unless the Fe²⁺status in the apoplast is in some way altered by the elicitor to facilitate OH^.production via the Fenton reaction.     

Avirulent strain of Colletotrichum induces a systemic resistance in strawberry

Strawberry plants exposed to an avirulent isolate of Colletotrichum fragariae acquired strong resistance against a virulent strain of C. acutatum. Biochemical, morphological and molecular markers indicated that the strong defence response was associated with an oxidative burst and a transient accumulation of salicylic acid (SA). A maximum accumulation of H₂O₂ and O₂ ^? was observed 8 h after inoculation (hai), callose was detected 48 hai, and a peak of SA was observed 48 hai. Biochemical and phytopathogenic analyses carried out in non-treated tissues revealed that the defence response was systemic and remained fully active 60 days after the first inoculation. Experiments also showed that the resistance acquired by mother plants after the inoculation with the avirulent isolate could be passed to daughter plants through runners. Further characterization of the induced systemic resistance showed that the resistance was not only effective against a virulent strain of C. acutatum but also against Botrytis cinerea.     

Activation of defence responses to Phytophthora infestans in potato by BABA

Late blight caused by Phytophthora infestans is one of the most devastating diseases of the potato crop. Resistance breeding and current fungicides are unable to control the rapidly evolving P. infestans and new control strategies are urgently needed. This study examined mechanisms of dl ‐β‐aminobutyric acid (BABA)‐induced resistance (IR) in the potato–P. infestans system. Leaves from two cultivars that differ in their degree of resistance, Bintje and Ovatio, were analysed after foliar treatment with BABA. Rapid activation of various defence responses and a significant reduction in P. infestans growth were observed in leaves treated with BABA. In the more resistant cultivar, Ovatio, the activation was both faster and stronger than in Bintje. Microscopic analysis of leaves treated with BABA revealed induction of small hypersensitive response (HR)‐like lesions surrounded by callose, as well as production of hydrogen peroxide (H₂O₂). Molecular and chemical analyses revealed soluble phenols such as arbutin and chlorogenic acid and activation of PR‐1. These results show a direct activation of defence responses in potato, rather than priming as reported for other plant species. They also show that the efficiency of BABA‐IR differs between cultivars, which highlights the importance of taking all aspects into consideration when establishing new methods for disease management.     

Comparative analysis of early H2O2 accumulation in compatible and incompatible wheat–powdery mildew interactions

Comparative analysis was carried out for hydrogen peroxide (H₂O₂) accumulation in response to powdery mildew (Blumeria graminis[Erysiphe graminis] f.sp. tritici) primary germ tube (PGT) and appressorial germ tube (AGT) contact, and in papilla (cell wall apposition) and hypersensitive responses (HR) in wheat (Triticum aestivum). Using primary leaves of three susceptible wheat lines (Bainong 3217; Beijing 837; Jingshuang 16) and five resistant lines [Mardler (Pm2 + Pm6); Ulka/8 × Cc (Pm2); Mardler/7 × Bainong 3217 (Pm2); Pm16; Pm16/7 × Beijing 837 (Pm16)], early H₂O₂ accumulation between 10 and 48 h after inoculation was studied. Strong H₂O₂ accumulation was found in effective papillae and associated cytosolic vesicles in both susceptible and resistant wheat lines, suggesting the important role of H₂O₂ in effective papillae formed as a general plant defence against powdery mildew. High frequency of effective papilla formation was observed in all five resistant lines. Among resistant lines, hypersensitive cell death was detected earlier in Mardler and Ulka/8 × Cc than in Pm16 and Pm16/7 × Beijing 837. In all cases this was associated with H₂O₂ accumulation in attacked epidermal cells. Interestingly, penetration resistance but not HR appeared to be mediated by the Pm2 gene in the Mardler/7 × Bainong 3217 line, suggesting that Pm2 may govern an HR-independent defence pathway in this genetic background. As effective papillae and HR did not occur in the same cells, papilla deposition may be independent of the HR response, despite the fact that both defence mechanisms were associated with high H₂O₂ accumulation. In wheat powdery mildew-incompatible interactions, HR acts as a second line of defence to contain infection when the papilla defence fails.     

葡萄应答霜霉病过程中多磷酸肌醇激酶基因与过氧化氢的作用机制

以对霜霉病不同抗性的葡萄品种左优红和霞多丽为材料,利用分子生物学和植物生理学试验手段,结合药理学试验,探讨葡萄在应答霜霉病过程中葡萄多磷酸肌醇激酶基因（VvIPK2）和H2O2的作用机制。接种霜霉病菌后15 h葡萄叶片VvIPK2表达量是正常水平的12倍,接种后3 hH2O2含量达最大值,同时苯丙氨酸解氨酶和几丁质酶活性升高;多磷酸肌醇激酶（IPK2）抑制剂、外源H2O2及H2O2清除剂均能改变霜霉病菌所引起的抗性葡萄品种左优红叶片PAL和几丁质酶活性的变化,同时可以影响不同抗性品种叶片的感病情况;IPK2抑制剂对葡萄霜霉病菌引起的H2O2水平变化没有影响;清除H2O2可减弱葡萄霜霉病菌对VvIPK2表达量的诱导效应。研究表明H2O2位于IPK2的上游,通过调控PAL和几丁质酶活性参与葡萄应答霜霉病过程。     

Agrobacterium induces plant cell death in wheat (Triticum aestivum L.)

The symptoms of gall or hairy root do not occur in the interactions between wheat (Triticum aestivum L.) and other monocotyledonous plants, with Agrobacterium tumefaciens or Agrobacterium rhizogenes. However, both bacteria colonized wheat root surfaces at similar levels (2.0 × 10⁷ colony forming U g⁻¹ root) and grew without inhibition in suspension with intact or wounded wheat embryos or root segments present. Suspension-cultured wheat embryo cells grown in 7.4 m M O₂ displayed 23% cell death after 1 h exposure to Agrobacterium cells, while the extent of cell death with 2.1 m M O₂ averaged 8%. Cultured wheat embryo and root cells rapidly produced hydrogen peroxide (H₂O₂) when contacted with A. tumefaciens or A. rhizogenes. Production of H₂O₂ was lower at 2.1 m M O₂ than 7.4 mM O₂. Browning and autofluorescence of epidermal cells of callus derived from wheat embryos and wheat roots was observed after inoculation with Agrobacterium. An increase in ferulic acid was detected in the walls of roots exposed to Agrobacterium. However, neither lignin nor callose was detected by diagnostic staining methods. These findings suggest that Agrobacterium induced a resistance-like response in wheat that may reduce the efficacy of transformation and limit the normal symptom formation.     

Riboflavin (Vitamin B2) induces defence responses and resistance to Plasmopara viticola in grapevine

Grapevine (Vitis vinifera) is susceptible to a variety of pathogenic fungi that affect yield and wine quality. Their control is generally achieved by widespread application of fungicides in the vineyards. The economic costs and negative environmental impact associated with these applications has led to a quest for alternative strategies, focusing on manipulation of host defence mechanisms. Here, we evaluated the ability of riboflavin (i.e. Vitamin B₂) to induce resistance against downy mildew in grapevine. Our results showed that 2 mM riboflavin applied 1–3 days before pathogen inoculation provided a disease reduction efficiency of 86 %. A microscopic analysis of the time course of P. viticola colonization in riboflavin-treated leaf discs suggests early inhibition of hyphae spreading in the intercellular space. This resistance does not result from a direct fungitoxic effect of riboflavin. However, this vitamin activates host-defence responses including H₂O₂ generation, upregulation of an array of defence-related genes and synthesis of callose in stomata cells, while stilbene synthesis is not affected. Using a pharmacological approach, we deduced that both jasmonic acid and callose biosynthesis pathways are significantly involved in riboflavin-induced resistance against downy mildew in grapevine.     

吲哚乙酸引导下三唑醇在大豆植株中的传导与积累研究初报

将吲哚乙酸与三唑醇发生酯化反应后生成的吲哚乙酸三唑醇酯 ,在0.5 mmol/L浓度下水培和喷雾处理6~8叶期大豆植株,同时以相同浓度的三唑醇及其与吲哚乙酸的混合物为对照,色谱法测定不同时间植株不同部位吲哚乙酸三唑醇酯和三唑醇的含量。结果表明,与对照相比,吲哚乙酸三唑醇酯在大豆中具有明显的双向传导和向根部积累的特点。喷雾处理后,在12~60 h之间内吸量和根部积累量均出现最大值。     

Mechanisms of induced resistance in lettuce against <Emphasis Type="Italic">Bremia lactucae</Emphasis> by DL<Emphasis Type="Bold">-</Emphasis>β<Emphasis Type="Bold">-</Emphasis>amino<Emphasis Type="Bold">-</Emphasis>butyric acid (BABA)

BABA induced local and systemic resistance in lettuce (Lactuca sativa) against the Oomycete Bremia lactucae. Structure-activity analysis showed no induced resistance by related amino-butanoic acids or β-alanine. The R-enantiomer of BABA induced resistance whereas the S-enantiomer did not, suggesting binding to a specific receptor. Other compounds known to be involved in SAR signaling, including abscisic acid, methyl-jasmonate, ethylene, sodium-salicylate and Bion® (BTH) did not induce resistance. Systemic translocation of ¹⁴C-BABA and systemic protection against downy mildew were tightly correlated. BABA did not affect spore germination, appressorium formation, or penetration of B. lactucae into the host. Epifluorescence and confocal microscopy revealed that BABA induced rapid encasement with callose of the primary infection structures of the pathogen, thus preventing it from further developing intercellular hyphae and haustoria. Invaded host cells treated with BABA did not accumulate phenolics, callose or lignin, or express HR. In contrast, cells of genetically-resistant cultivars accumulated phenolics, callose and lignin and exhibited HR within one day after inoculation. The callose synthesis inhibitor DDG did not inhibit callose encasement nor compromised the resistance induced by BABA. PR-proteins accumulated too late to be responsible for the induced resistance. DAB staining indicated that BABA induced a rapid accumulation of H₂O₂ in the penetrated epidermal host cells. Whether H₂O₂ stops the pathogen directly or via another metabolic route is not known.     

Induced defense dynamics in plant parts is requisite for resistance to <Emphasis Type="Italic">Helicoverpa armigera</Emphasis> (Hubner) infestation in chickpea

Helicoverpa armigera is the most serious insect pest in chickpea that causes significant yield losses due to its feeding on vegetative (leaves) and reproductive (developing pods and seeds) parts of plants. The present aim of study was to explore response dynamics of induced defence mechanism in leaves, podwall and seeds of ten chickpea genotypes (ICC 506, ICCV 10, ICC 10393, 5283, RSG 963, GL 25016, GL 26054, ICCL 86111, ICC 3137, L 550) after insect infestation. Two chickpea genotypes namely ICC 3137 and L 550 were found to be highly susceptible to Helicoverpa armigera infestation due to higher leaf and pod damage in them as compared to rest of eight genotypes which are found to be considerably resistant due to lower damage. Insect infestation induced decreased activities of defensive enzymes such as peroxidase (POD), catalase (CAT), glutatione reductase (GR) and polyphenol oxidase (PPO), decreased free radical scavenging activities in terms of 2,2-diphenyl-1-picryl hydrazyl (DPPH), decreased contents of signaling molecules such as nitric oxide ((NO), hydrogen peroxide (H₂O₂), reduced content of insect feeding behaviour regulating molecules such as total phenols, trypsin inhibitor and accumulation of membrane damage marker such as malondialdehyde (MDA) in leaves of ICC 3137 and L 550; decreased POD activity, nitric oxide content and H₂O₂ in podwall of L550; decreased SOD, GR, nitric oxide content and H₂O₂ in seeds of L550 resulted in aggravation of infestation induced oxidative stress and makes these genotypes more vulnerable to insect damage. The resistance of rest eight chickpea genotypes to insect infestation was due to the integrative effect of up regulated defensive components in leaves, podwall and seeds such as enhanced activities of CAT, POD, GR, PPO and PAL along with accumulation of H₂O₂` and total phenols in leaves, increased SOD, POD, GR and PPO activities along with increased contents of trypsin inhibitor and total phenols in podwall; increased SOD, GR, PPO activities and accumulated total phenols in seeds of resistant chickpea genotypes might be responsible for causing significant shift in oxidative status of these genotypes due to scavenging of free radicals, maintenance of membrane integrity and deterrent to insect feeding. Induced glycine betaine after herbivory was found to be positively correlated with superoxide dismutase and trypsin inhibitors. H₂O₂ content was positively correlated with trypsin inhibitor, DPPH, ferric reducing antioxidant power (FRAP) and total phenols in leaves and with FRAP, DPPH and total phenols in pod wall indicating that H₂O₂ might be stimulating the cascade that will be helping to scavenge free radical species and correlation with phenols and trypsin inhibitor indicated that it act as toxicant to insect feeding.     

Arabidopsis ethylene receptors have different roles in Fumonisin B1-induced cell death

Ethylene is a central signalling agent in mediating plant defence against pathogens. Mutations to the ethylene receptor ETR1 have been shown to alter susceptibility of plants to mycotoxin-induced cell death. Using Fumonisin B₁ (FB₁) to induce cell death, we demonstrate that the receptor mutant ein4-1 has a reduced rate of necrosis, potentially due to an upregulation of ETHYLENE RESPONSE FACTOR1. Mutations in other ethylene receptors differentially affected the expression of genes in the jasmonic and salicylic acid defence pathways. Together these data indicate that ethylene receptors do not have redundant roles in mediating FB₁-induced cell death.     

Cellular reactions in Arabidopsis following challenge by strains of Pseudomonas syringae: From basal resistance to compatibility

Reactions to strains of the bean halo-blight pathogen Pseudomonas syringae pv. phaseolicola (Pph) strain 1448A and the Arabidopsis and tomato pathogen P.s. pv. tomato (Pst) strain DC3000 were examined by transmission electron microscopy. Wild-type and hrpL mutant strains of Pph failed to multiply in the accession Columbia (Col)-5, but did not cause a hypersensitive reaction (HR). Symptomless non-host resistance to 1448A and the hrpL mutant was associated with the progressive alteration of the plant cell wall adjacent to bacteria, following the accumulation of membrane bound vesicles within the cytoplasm at reaction sites. Large papillae containing callose accumulated within challenged plant cells. Papillae also formed in the pmr4–1 mutant of Col-0 which lacks an inducible callose synthase but immunocytochemical labelling demonstrated that they contained very little β-1, 3 glucan. Some papillae formed in Col-5 in response to the virulent pathogen DC3000, but they dispersed during cell collapse and lesion formation. Transconjugants of Pph expressing the avirulence genes avrPpiA and avrPphB matching the RPM1 and RPS5 resistance genes, caused rapid and slow HR development, respectively. Although corpse morphology was observed our observations suggest that in Arabidopsis, plant cell death during the HR is programmed but represents a variant of necrosis rather than apoptosis. Cerium chloride staining revealed the accumulation of H₂O₂ at reaction sites. The strongest H₂O₂ response was found during the HR activated by avrPpiA but localised generation of peroxide was also found at sites of papilla deposition next to 1448A or the hrpL mutant. Accumulation of H₂O₂ during the HR, but not during wall alterations, was strongly suppressed by inhibition of NADPH oxidase. The differential effect of the inhibitor suggests an alternative source of H₂O₂ to modify the plant wall. Extension of peroxide-driven cross-linking reactions to bacterial cell walls may contribute to the restriction of bacterial multiplication. The lowest level of H₂O₂ occurred during the compatible reaction to DC3000. Characterisation of the cellular co-ordination of basal (non-host) resistance has revealed several potential targets for bacterial effector proteins.     

Oligochitosan induces cell death and hydrogen peroxide accumulation in tobacco suspension cells

Oligochitosan has been shown to induce several plant defense responses. In the present work, the effect of oligochitosan on tobacco cell survival was investigated. The results showed that oligochitosan caused tobacco cell death in a dose-dependent manner. About 40.6 % tobacco cells died when cultured for 72 h after 500 μg ml⁻¹ oligochitosan treatment. Certain aspects of this cell death process appeared to be similar to apoptosis in animal cells. These included shrinkage of cytoplasm and condensation of chromatin. Oligochitosan also induced H₂O₂ accumulation in tobacco cell suspension culture. The role of H₂O₂ in the signal transduction that leads to cell death was investigated. Co-treatment of tobacco cells with oligochitosan and catalase inhibited H₂O₂ accumulation but did not inhibit the induction of cell death. The results suggested that apoptosis-like cell death of tobacco cells induced by oligochitosan is independent of H₂O₂ signal pathway.     

BTH处理对甜瓜叶片活性氧代谢的诱导作用

为了明确活性氧（reactive oxygen species,ROS）代谢在甜瓜抗病性诱导中的作用,以抗白粉病甜瓜品种Tam Dew和感病品种卡拉克赛幼苗为材料,通过盆栽试验研究了苯丙噻二唑（BTH）喷雾或白粉菌接种后甜瓜叶片超氧阴离子（O2.-）产生速率、过氧化氢（H2O2）含量及超氧化物歧化酶（SOD）、过氧化氢酶（CAT）、苯丙氨酸解氨酶（PAL）活性的变化。BTH处理或白粉菌接种均可诱导甜瓜叶片SOD、PAL活性升高,抑制CAT活性,导致叶组织O2.-产生速率和H2O2含量增加,BTH喷雾＋白粉菌接种比二者单独处理效果更好。结果表明,BTH处理后叶片O2.-产生速率提高和H2O2积累是甜瓜抗白粉病能力提高的重要机制,BTH通过诱导ROS代谢酶活性调节H2O2含量,且BTH诱导的甜瓜抗病性与品种的基础抗性有关。