生物诱抗剂草酸对黄瓜叶片中过氧化物酶的系统诱导作用

 本文进一步证明草酸是一种有效的非生物诱抗剂,它能显著提高黄瓜对炭疽病的系统抗性,进而研究了草酸对黄瓜叶片中与抗病有关的过氧化物酶(POD)的系统诱导作用及其内在机理。结果表明:40mmol/L草酸诱导可溶态POD活性效果最佳;在测定时间范围内,草酸诱导的可溶态POD活性在第7d后逐渐下降,而细胞壁离子键结合态POD活性被诱导后保持稳定;草酸在快带区诱导出一种新的POD同功酶,其Rf值为0.63;自由基清除剂甘露醇和抗坏血酸可明显抑制草酸对可溶态POD活性的诱导作用;草酸诱导可溶态POD的作用信号可在5h左右从被处理的第1片真叶传导至未处理的第2片真叶,被处理叶的保留时间越长,其诱导效果越好。     

钙信使系统参与草酸对湖北海棠POD活性的诱导

 以EGTA、异博定(Ver)、氯丙嗪(CPZ)和Li⁺处理湖北海棠叶片,考察了钙信使系统在草酸诱导过氧化物酶(POD)中的作用。结果显示,4种抑制剂分别与草酸同时或先于草酸处理时,均明显抑制草酸对叶片POD总活性的诱导,表明Ca²⁺、Ca²⁺通道、钙调素(CaM)和磷酸肌醇均可能参与了草酸对POD的诱导。此外,草酸处理后一定时间再用抑制剂处理,也能够抑制草酸对POD的诱导作用。EGTA和Ver在草酸处理8h后应用的效果低于4h后应用;CPZ和Li⁺在草酸处理后4或8h应用,效果相近。     

光合细菌复配柠檬酸铜对移栽稻田水绵防效的研究

水绵是对水稻危害较重的低等藻类,本研究采用盆栽试验方法研究沼泽红假单胞菌复配柠檬酸铜对水绵的鲜重防效及施用后水绵中Mg²⁺、Fe²⁺、叶绿素含量和抗逆酶活性,同时测定了施用后水稻生长指标及抗逆酶活性。结果表明,施药后28 d,沼泽红假单胞菌复配柠檬酸铜对水绵的鲜重防效达92.17%,显著高于沼泽红假单胞菌或柠檬酸铜单独施用。与对照相比,水绵叶绿素、Mg²⁺和Fe²⁺含量分别显著降低60%,43%和39%,POD和SOD活性分别显著降低72%和71%,MDA含量显著升高50%。水稻叶片POD和SOD活性分别显著增加44%和104%,MDA含量显著降低51%,单株鲜重显著增加43%,分蘖数显著增加65%,株高增加5%。综上,沼泽红假单胞菌复配柠檬酸铜对水绵防效显著且对水稻安全,水绵叶绿素合成受到抑制和细胞膜过氧化作用是其对水绵防除的重要作用机理。     

草酸和BTH对黄瓜幼苗霜霉病抗性和胞间隙病程相关蛋白的诱导

 本文研究了草酸和BTH(苯并噻二唑)溶液对黄瓜幼苗霜霉病的抗性诱导及病程相关蛋白的积累。结果表明:10mmol/L草酸或0.5mmol/LBTH均能诱导黄瓜对霜霉病产生局部和系统抗性,且抗性的持久期均在15d以上。BTH处理或接种霜霉菌都可系统诱导黄瓜叶片胞间隙液产生分子量分别为33、27和22kD的蛋白,而草酸没有诱导这3种蛋白的表达。BTH或草酸处理均可引起POD活性的升高,但和对照相比并没有新的POD同工酶表达,POD活性升高与黄瓜对霜霉病的抗性有关。     

Ca2+,Mn2+等8种金属离子对软腐欧文氏菌一些致病因子的影响

 在离体试验中,不同金属离子对马铃薯软腐欧文氏菌(Erwinia carotovora subsp.carotovora)菌株StEcc-12生长,产生胞外酶和胞外酶活性具有不同的影响。在含果胶酸钠的培养液中,Ca²⁺促进生长,并使培养液中的果胶裂解酶(PL)、果胶水解酶(PG)和蛋白酶3种胞外酶分别提高1.4~23.4倍,0.3~2.9倍和0.7~6.5倍。Mn²⁺对StEcc-12生长及产生3种胞外酶的影响与Ca²⁺相似。Ni²⁺显著抑制细菌的生长,但在含Ni²⁺培养液中3种胞外酶的单位活性比对照高1.7倍以上。在酶粗提液中分别加入上述3种离子对PL酶的活性均有抑制作用。Zn²⁺对细菌生长没有显著影响,高浓度Al³⁺抑制细菌生长,这2种离子均能促进3种胞外酶的产生。随着Fe²⁺和K⁺浓度的增高,细菌生长减少,两种果胶酶产量降低,蛋白酶产量增加,粗提液中PL酶活性下降。Mg²⁺对细菌生长和胞外PG产生没有明显影响,提高Mg²⁺浓度有利于PL的产生。酶粗提液中加入Mg²⁺后PL活性增高。所有以上离子都能使聚半乳糖醛酸发生不同程度的凝聚,但只有Ca²⁺和Mg²⁺的有效凝聚浓度与薯块的实际浓度接近。     

拮抗细菌诱导番茄植株抗灰霉病机理研究

 拮抗细菌多粘类芽孢杆菌(Paenibacillus polymyxa) W3菌株悬浮液及其滤液可以诱导番茄叶片对灰霉病(Botrytis cinerea)的系统抗性。W3及其滤液诱导处理后,植株叶内苯丙氨酸解氨酶(PAL)、过氧化物酶(POD)、多酚氧化酶(PPO)和超氧化物歧化酶(SOD)活性明显增强。诱导后1 d,PAL活性最大,是对照的3.8~3.9倍,6 d后仍为对照的2.5倍;POD和PPO诱导后3 d活性最高,分别比对照增加34.7%~54.1%和78.5%~78.7%,6 d后仍比对照高;SOD活性诱导后2d达高峰,6 d后稍高于对照。活性氧(O₂^-)产生速率诱导后1 d最大,比对照增加85.6%~88.6%,以后急剧下降,6 d后接近对照。此外,W3诱导后1 d或2 d,处理叶和上一叶位叶片水杨酸含量明显上升,分别是对照的2.6倍和1.6倍,这表明该拮抗细菌诱导的系统抗性可能与水杨酸介导有关。     

钙离子和茉莉酸灌根对菜豆叶片防御酶活性和西花蓟马取食偏好性的影响

为探明外源物质对菜豆植株防御酶活性和西花蓟马取食偏好性的影响。本试验分别采用CaCl₂和茉莉酸(JA)进行灌根,研究了西花蓟马取食前后菜豆叶片脂氧合酶(LOX)、丙二烯氧化物合酶(AOS)、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、过氧化物酶(POD)和多酚氧化酶(PPO)活性的变化,并研究了西花蓟马对JA和CaCl₂处理后的菜豆植株的取食偏好性。结果表明：不论CaCl₂还是JA灌根均可诱导菜豆防御酶AOS、LOX、CAT和SOD活性的上升,其中对CAT活性影响最大,其活性分别上升至对照的2.83倍和3.57倍,但对POD和PPO活性没有影响。西花蓟马取食CaCl₂处理的菜豆植株,能诱导LOX、POD和PPO活性分别上升1.40、2.03倍和2.26倍,而SOD活性则被显著抑制。西花蓟马取食JA灌根的菜豆植株能明显诱导POD和PPO活性的升高,二者活性分别是未取食植株的1.67倍和1.45倍,但AOS、LOX、CAT和SOD活性被抑制。西花蓟马2龄若虫取食CaCl₂和JA...     

H2O2、NO和Ca2+参与疫病菌激发子PB90诱导烟草气孔的关闭

 本文研究了过氧化氢(H₂O₂)、一氧化氮(NO)和Ca²⁺在棉疫病菌激发子PB90诱导烟草气孔运动中的作用。1 nmol/L激发子PB90可诱导野生型烟草Bel-W3气孔关闭,且在相同条件下该激发子诱导反义抑制抗坏血酸过氧化物酶anti-APX烟草气孔关闭的孔径比野生型的更小;进一步药理学证明,抗氧化剂DTT和NADPH氧化酶抑制剂DPI(diphenylene iodonium)、一氧化氮合酶(NOS)抑制剂L-NAME、Ca²⁺螯合剂EGTA和Ca²⁺信号途径中CaMPK Ⅱ的竞争抑制剂KN93与磷脂酶C抑制剂U73122都能抵消PB90诱导气孔关闭的效应。推测该激发子PB90通过NADPH氧化酶途径形成H₂O₂、NOS途径形成NO和Ca²⁺信号途径进而诱导气孔关闭。表明H₂O₂、NO、Ca²⁺在激发子PB90诱导气孔关闭信号传递中作为第二信使起着重要的作用。     

重寄生真菌盾壳霉胞外蛋白酶产生条件及酶活影响因子

重寄生真菌盾壳霉Coniothyrium minitans是核盘菌Sclerotinia sclerotiorum的重要生防菌。为了探讨盾壳霉胞外蛋白酶在寄生核盘菌过程中的作用,采用明胶平板法对盾壳霉寄生核盘菌菌核产生的蛋白酶活性进行了检测,并进一步采用福林酚法定量测定蛋白酶活性,研究盾壳霉产生胞外蛋白酶的培养条件及影响蛋白酶活性的因子。试验结果表明,在被盾壳霉寄生的核盘菌菌核中检测到蛋白酶活性,表明蛋白酶可能参与盾壳霉重寄生作用。发现核盘菌菌核浸出液培养基适合盾壳霉产生胞外蛋白酶,摇培(20℃、200r/min)5d时蛋白酶活性最高,达到0.22U/mL。盾壳霉胞外蛋白酶酶促反应的最适温度为60℃,最适pH7.0。当温度不高于40℃时,蛋白酶酶活较稳定。5mmol/L的金属离子Mg²⁺、Zn²⁺、Ca²⁺、Cu²⁺、Mn²⁺、Li⁺和K⁺等对蛋白酶酶活没有显著影响(P>0.05),而Fe²⁺(5mmol/L)显著(P<0.05)提高了蛋白酶活性。盾壳霉蛋白酶对苯甲基磺酰氟(PMSF)敏感,说明盾壳霉产生的胞外蛋白酶可能主要是丝氨酸蛋白酶。这些结果为盾壳霉胞外蛋白酶的分离纯化和功能研究奠定了基础。     

几种化学物质诱导彩色马蹄莲对软腐病抗性的研究

本试验测定了纳米硅、草酸、硅酸钠和硫酸亚铁4种化学物质在不同浓度下对彩色马蹄莲软腐病菌的室内抑菌活性和诱导抗病效果。结果表明,硫酸亚铁3种浓度对彩色马蹄莲软腐病菌均表现较强的抑制作用;1 g/L纳米硅、0.15 g/L草酸和0.2 g/L硅酸钠3种化学物质对软腐病菌无明显抑制作用且诱导抗病效果较好,分别达到62.28%、77.56%、88.46%;经3种化学物质诱导处理后再进行接种,诱导处理和接种期间彩色马蹄莲叶片组织内过氧化物酶(POD)、苯丙氨酸解氨酶(PAL)、多酚氧化酶(PPO)酶活性均高于对照,其中硅酸钠处理后彩色马蹄莲叶片组织内POD、PAL活性高于纳米硅和草酸处理,草酸处理后叶片组织内PPO活性高于纳米硅和硅酸钠处理。     

不同化学诱抗剂对金银花叶片防御酶系的影响

用4种化学诱抗剂壳聚糖、草酸、硫酸亚铁和硅酸钠处理金银花,通过计算酶活性发展曲线下面积AUEAPC,分析了叶片防御酶系3种主要酶的活性变化情况。结果表明：4种诱抗剂处理中,壳聚糖处理的苯丙氨酸解氨酶（PAL）活性最高,其次是草酸处理,两者差异极显著,并且极显著高于硅酸钠、硫酸亚铁处理和对照,而后3种处理间差异不显著;硫酸亚铁处理后,过氧化物酶（POD）的活性最高,极显著高于壳聚糖、草酸和硅酸钠处理,并且这4种诱抗剂处理POD的活性均极显著高于对照;但只有壳聚糖处理,多酚氧化酶（PPO）的活性极显著高于对照,其他3种诱抗剂处理的PPO活性极显著低于对照。壳聚糖、草酸、硫酸亚铁和硅酸钠处理后,3种酶的综合活性分别比对照提高71.1%、31.1%、11.7%和2.5%,与前期对金银花白粉病诱抗效果研究结论相一致。本研究初步从防御酶学角度分析了4种化学诱抗剂在金银花上的诱导抗病机制。     

水杨酸诱导水稻幼苗抗瘟性的生化机制

 以0.01mM水杨酸(SA)喷雾处理或稻瘟菌孢子悬浮液接种稻苗后,叶片中苯丙氨酸解氨酶(PAL)和定位于细胞不同部位的过氧化物酶(POD),特别是与细胞壁结合的POD和细胞间隙POD的活性迅速升高,但没有明显的新POD同功酶带出现。同时叶片中木质素含量迅速增加。在SA处理后的稻叶中提取到能抑制稻瘟菌分生孢子萌发、含有Momilactone A的抑菌物质,并检测到分子量分别为48、52、59、106.5KDa的碱性病程相关蛋白(PRs)。这些生化指标的时序变化与SA诱导抗瘟性的表现相吻合。对SA诱导水稻幼苗抗瘟性的可能生化机理作了归纳。     

Degradation of anilinopyrimidine fungicides photoinduced by iron(III)-polycarboxylate complexes

The photoinduced degradation of three anilinopyrimidine fungicides (cyprodinil, pyrimethanil and mepanipyrim) by Fe(III)-polycarboxylate complexes in aqueous solution was investigated. A photochemical redox reaction of Fe(III) complexes of oxalate and citrate occurs during irradiation (simulating sunlight) and is an important source of Fe(II) and a series of oxidants such as H2O2 and O radicals. The mechanism involves the formation of polycarboxylate radicals and/or carbon-centred radicals derived from decarboxylation, whereas the contribution of Fe(OH)2+ to O radical formation is negligible. The attack of O radicals on the fungicide molecules produces numerous photodegradation products, which were identified by means of LC-ES-MS and turned out to be mono- or dihydroxylated derivatives of the active ingredients, except for 2-amino-4,6-dimethylpyrimidine, which is only formed by pyrimethanil. The half-lives of the active ingredients, when submitted to irradiation in the presence of iron(III)-polycarboxylate complexes, were estimated to vary between 28 and 79 min (photodegradation rates in the same conditions: mepanipyrim > cyprodinil > pyrimethanil), and photodegradation is slower in citrate than in oxalate solutions. Photoproducts and their kinetics of formation are very similar for the three fungicides. The OH substitution involves the aromatic and the heterocyclic ring and the nitrogen bridge between the two rings, except for mepanipyrim when the hydroxylation also involves the propynylic side chain.     

A binding protein for fungal signal molecules in the cell wall of Pisum sativum

In the plant cell wall of Pisum sativum seedlings, we found an NTPase (E.C. 3.6.1.5.) with ATP-hydrolyzing activity that was regulated by an elicitor and suppressors of defense from pea pathogen Mycosphaerella pinodes. The ATPase-rich fraction was purified from pea cell walls by NaCl solubilization, ammonium sulfate precipitation, and chromatography with an ATP-conjugated agarose column and an anion-exchange column. The specific activity of the final ATPase-rich fraction increased 600-fold over that of the initial NaCl-solubilized fraction. The purified ATPase-rich fraction also had peroxidase activity and generated superoxide, both of which were regulated by the M. pinodes elicitor and suppressor (supprescins). Active staining and Western blot analysis also showed that the ATPase was copurified along with peroxidases. In this fraction, a biotinylated elicitor and the supprescins were bound primarily and specifically to ca. 55-kDa protein (CWP-55) with an N-terminal amino acid sequence of QEEISSYAVVFDA. The cDNA clone of CWP-55 contained five ACR domains, which are conserved in the apyrases (NTPases), and the protein is identical to a pea NTPase cDNA (GenBank accession AB071369). Based on these results, we discuss a role for the plant cell wall in recognizing exogenous signal molecules.     

Metal chelate molluscicides: The redistribution of iron diazaalkanolates from the gut lumen of the slug,Deroceras reticulatum (Müller) (Pulmonata: Limacidae)

Two related iron chelates, one toxic to slugs by ingestion, the other not, were introduced into the foregut of D. reticulatum. The subsequent movement and redistribution of the metal within the slug tissues was studied by labelling the chelates with the radioactive isotope ⁵⁵Fe. In slugs which survived treatment approximately half of the ⁵⁵Fe was voided in faeces. The iron retained became unevenly distributed, the highest concentration occurring in the digestive gland, irrespective of the chelate used. At high doses, slugs treated with tris(1-oxo-1,2-diazabutan-2-oxido)Fe(III) were fatally poisoned while those treated with the homologue, tris(1-oxo-1,2-diazaoctan-2-oxido)Fe(III) were not. Slugs killed by the toxic chelate consistently contained proportionally less iron in the digestive gland and proportionally more in the body wall and reproductive system. Dosing slugs already killed by carbon dioxide asphyxiation gave a similar pattern, suggesting that the greater mobility of the iron from the toxic chelate was not a function of the slugs' metabolism.     

Potential role for salicylic acid in induced resistance of asparagus roots to Fusarium oxysporum f.sp. asparagi

Pre-inoculation of asparagus ( Asparagus officinalis ) roots with selected nonpathogenic isolates of Fusarium oxysporum (np Fo ) has previously been shown to induce systemic resistance against infection by F. oxysporum f.sp. asparagi ( Foa ) through activation of plant-defence mechanisms. To elucidate the putative np Fo -mediated defence pathways, the effect of salicylic acid (SA) was examined in a split-root system of asparagus where one half of the seedling root system was drenched with SA and the activation of defence responses was measured subsequently on the remaining roots. SA-treated plants exhibited enhanced systemic resistance, with a significant reduction in disease severity of the roots inoculated with Foa , compared with untreated plants. SA activated peroxidase and phenylalanine ammonia-lyase, as well as lignification, upon Foa attack, in a manner similar to that observed with np Fo pretreatment. In addition, application of diphenyleneiodonium, an SA biosynthesis inhibitor, led to failure of np Fo to induce lignin deposition and systemic resistance. Treatment of fungal spores with SA did not affect germination and growth of either np Fo or Foa in in vitro antifungal assays. Production of SA at the site of np Fo infection may be involved in the induction of Foa resistance in asparagus roots.     

Benzothiadiazole and BABA improve resistance to <Emphasis Type="Italic">Uromyces pisi</Emphasis> (Pers.) Wint. in <Emphasis Type="Italic">Pisum sativum</Emphasis> L. with an enhancement of enzymatic activities and total phenolic content

Benzothiadiazole (BTH) and DL-β-aminobutyric acid (BABA) induced systemic resistance was investigated in susceptible and resistant pea genotypes against Uromyces pisi. Resistance was characterized by reduced infection frequency mainly due to decreases in appressorium formation, stomatal penetration, growth of infection hyphae and haustorium formation. Changes in β-1,3-glucanase, chitinase, phenylalanine ammonia-lyase and peroxidase activities and in total phenolics content, demonstrate that U. pisi resistance is induced by BTH and BABA treatments at early and late stages of the fungal infection process, but that the chemicals operate via different mechanisms. In fact, our study showed that BTH treatment primed the activity of pathogenesis related-proteins such as β-1,3-glucanase, chitinase and peroxidase in both susceptible and resistant genotypes. On the other hand, BABA treatment did not increase the enzymatic activities in the studied genotypes, but significantly increased their total phenolic contents.     

H2S位于H2O2下游介导葡萄抗霜霉病过程

 以霜霉病抗性葡萄品种左优红和感病品种霞多丽为材料,研究硫化氢(H₂S)在葡萄应答霜霉病过程中的作用及其与过氧化氢(H₂O₂)的相互关系。结果表明,接种霜霉病菌后,抗性品种左优红和感病品种霞多丽叶片均出现H₂S含量的猝发、H₂S合成酶基因表达量增加、抗病相关蛋白多酚氧化酶(PPO)和β-1,3葡聚糖酶(Glu)活性显著升高现象;且H₂S清除剂次牛磺酸(HT)可以显著抑制霜霉病菌诱导的PPO和Glu活性升高,并导致葡萄的感病率和病情指数显著升高;同时外源H₂S可降低两个葡萄品种的感病率和病情指数。说明,H₂S是参与葡萄抗霜霉病过程的新的信号物质。受霜霉病菌侵染后,葡萄叶片H₂O₂含量猝发早于H₂S,并且H₂O₂清除剂抗坏血酸(AsA)可显著抑制霜霉病菌所诱导的H₂S含量的上升,而H₂S清除剂HT对霜霉病菌诱导的H₂O₂含量变化影响不显著,推测,H₂S作用于H₂O₂的下游参与葡萄抵御霜霉病过程。