油茶叶肿病变态叶片的形态特征和超微结构观察

利用扫描电镜(SEM)对油茶叶肿病变态叶叶片表面和横切面进行观察,利用透射电镜(TEM)对其细胞超微结构进行观察,以期探明油茶叶肿病变态叶的形态特征和细胞学特征。结果表明:(1)变态叶是受感染油茶幼叶组织增生形成的,肿大的叶片厚度比正常叶片厚度增加3~5倍,细胞体积增大3~8倍,细胞数增加1~2倍,叶片细胞形态和结构发生了变化。(2)叶片受细丽外担菌侵染后,菌丝存在于下表皮向内的4~7层细胞间隙中,感染后期叶片下表面脱落露出子实层。(3)变态叶细胞出现叶绿体膜破裂、类囊体片层膜数目减少及细胞器成分被破坏等异常现象。     

环境激素DBP对拟南芥体外培养叶片超微结构的影响

报道了酞酸酯类化合物DBP对拟南芥离体培养叶片超微结构的影响。在DBP(1.0mg·L-1)处理第3天即观察到拟南芥叶片叶绿体的超微结构受到破坏。不同浓度的DBP长期处理植株(40d)出现叶片白化、节间短缩等异常现象。在0.01mg·L-1DBP处理40d植株的叶细胞中,叶绿体出现解体,破碎部分呈颗粒状,散落其间,但细胞结构完整;0.1mg·L-1DBP处理后引起叶绿体的进一步解体,叶绿体中类囊体基粒和片层结构解体,细胞出现空洞现象,其它细胞器很少观察到;1.0mg·L-1DBP和2.0mg·L-1DBP处理植株叶片叶绿体中类囊体基粒和片层结构破碎,叶绿体结构也解体,细胞中其它细胞器数目极少。     

淹水对玉米叶片细胞超微结构的影响

对淹水过程中玉米（Ｚｅａ ｍａｙｓ Ｌ．）叶片细胞超微结构的变化进行连续观察。淹水２ｈ后,液泡膜发生明显内陷。淹水６ｈ后,液泡膜内陷加剧,呈极度松弛状态;叶发体被膜局部向外突出一个由单层膜包裹的泡状结构。淹水１２ｈ后,液泡膜局部破裂;叶绿体被膜破坏加剧,成为一松弛的单膜结构,同时,基质类囊体出现空泡化。淹水１８ｈ后,叶绿体的破坏进一步加剧：被膜完全消失,基质类囊体开始消化;同时,线粒体膜和核膜也开     

NaCl胁迫对宁夏枸杞叶和幼根显微及超微结构的影响

以宁夏枸杞为材料,采用超薄切片技术制备样品,应用光学显微镜和透射电镜分析了不同浓度NaCl胁迫条件下宁夏枸杞叶和幼根显微及超微结构的变化。结果表明：随着NaCl胁迫的加重,(1)叶片上表皮细胞增厚,栅栏组织细胞出现缩短现象,排列疏松且紊乱;幼根的初生结构无明显变化。(2)叶片栅栏组织中叶绿体不再紧靠在细胞膜上,叶绿体双层膜破坏,基粒片层松散排列,杂乱无章,出现膨胀和空泡现象,淀粉粒和嗜锇颗粒增多,叶肉细胞中线粒体发生轻微变化;幼根中皮层薄壁细胞线粒体形状发生改变,结构破坏,内膜和外膜模糊甚至破裂,大多数嵴模糊,出现空泡现象;细胞核解体,基质外溢。研究表明, 不同浓度的NaCl胁迫对宁夏枸杞叶片和幼根细胞的显微及超微结构影响不同,NaCl浓度大于200 mmol/L时,宁夏枸杞叶片和幼根细胞的显微及超微结构发生了明显变化,且叶肉细胞中线粒体的变化没有叶绿体的变化显著,推测叶肉细胞中线粒体的耐盐性比叶绿体强。     

枫香变红过程中叶片组织结构、光合特性及色素含量变化研究

枫香(Liquidambar formosana)因其叶片入秋后逐渐变红而极具观赏价值,是优良的景观生态树种。为了解枫香叶片结构变化与叶色的关系,该文通过连续监测枫香叶片变红过程中组织结构、光合特性及色素含量的变化,分析叶片结构与其光合特性和色素的关系。结果表明:(1)叶片变色过程中,表皮细胞均为椭圆形,紧密排列,未观察到明显的细胞变异,表面未附着绒毛和蜡质,且上表皮细胞与栅栏组织细胞间排列紧密,未出现较大的气室。(2)随着叶片逐渐变红,叶片结构变化显著,其中叶片、上表皮、栅栏组织和海绵组织厚度及气孔开度均逐渐减小,而气孔器长和宽、单个气孔器面积则逐渐增大。(3)随着叶片结构的变化,其叶绿素含量逐渐减少,致使净光合速率逐渐减小,在出现光破坏时,叶片通过在栅栏组织细胞液泡内合成花色苷来自我保护,而大量的花色苷致使叶片表面呈现红色。综上认为,叶绿素含量降低,花色素苷大量积累是导致枫香叶片变红的直接原因,而枫香叶色变红则是其一系列生理结构特征综合作用的结果。     

噬菌体脱毒机理的研究Ⅰ

用噬菌体处理河流弧菌Ⅱ的时间不同,细菌菌落的透明度和超微结构也不一样。在吸附后７ｈ开始菌落出现透明度的变化,首先是不透明（乳黄色）的菌落占绝对优势,但已出现半透明和透明菌落,培养到４８ｈ时,几乎全部变为透明菌落。透明菌落不再能与其噬菌体吸附而产生噬斑,也不会再感染鲍而产生脓疱病。透射电镜观察发现,透明菌落的菌细胞形状和结构大都发生变化。细胞壁薄,细胞质浓缩集中在细胞的一侧或一端。不透明菌落细胞形状正常,细胞壁完整,细胞质外延使细胞横切面呈指环状。电镜观察不同时间取样发现,１号样品细胞结构无明显变化。２号样品多数细胞产生外膜泡。３号样品许多细胞结构与１号相似,但在一些细胞的附近出现成簇的外膜泡和噬菌体。４号样品细胞结构变化较大,细菌的核区被破坏,在细胞的周围发现许多噬菌体。５号样中发现许多菌细胞被破坏,也在一些细胞内发现噬菌体。６号样中透明菌落和不透明菌落细胞结构变化明显。７号样几乎所有菌落都变为透明状,许多细胞破裂等。     

基于叶片解剖特征分析三种无患子科果树的亲缘关系

采用石蜡切片和组织离析法,对3种无患子科果树的10份种质叶片横切及表皮解剖特征进行观察,并采用聚类分析对其亲缘关系进行初步研究。结果表明：10份试材均为异面叶,叶横切结构分为表皮、叶肉和叶脉3部分。中脉厚度总体上龙眼最大、龙荔次之、荔枝最小,差异显著。荔枝的中脉横切面为圆三角形,龙眼近似半圆形,龙荔则近似扁圆形。不同试材的叶片、上表皮和下表皮、栅栏组织及海绵组织的厚度分别为175.23~318.84、11.18~25.13、7.49~20.43、50.01~124.59和84.0~173.64μm,栅栏组织细胞有2~3层。此外,叶脉突起度、栅栏组织与海绵组织厚度之比、叶片组织结构紧密度和叶片组织结构疏松度分别为2.65~5.77、0.52~0.82、28.89％~39.95％和44.89％~55.57％。荔枝的表皮细胞较小、多边形,垂周壁为弧形,下表皮无表皮毛,气孔器呈长椭圆形;龙眼的表皮细胞较大、不规则形,垂周壁深波状,下表皮具表皮毛,气孔椭圆形或近圆形;龙荔的表皮细胞与龙眼相近,但垂周壁为波状。聚类分析显示,10份试材首先聚为两大类,其中第一类是荔枝属,第二类是龙眼属;然后又分为4个亚类,三月红和龙荔各自单独聚为一个亚类。     

小麦矮腥黑粉菌在小麦体内侵染过程的显微观察

小麦矮腥黑粉菌可导致小麦矮腥黑穗病,是麦类黑粉病中危害最大、极难防治的国际重要检疫性病害之一.本研究结合扫描电子显微镜、透射电子显微镜及激光共聚焦显微镜观察该真菌在小麦(Triticum aestivum)体内的侵染过程.经观察发现,被该真菌侵染后的小麦叶片细胞超微结构发生了显著变化,如叶肉细胞畸形、质膜内陷和断裂、细胞核结构破坏及细胞器的基质电子密度下降;细胞间隙出现空细胞和纤维状膜状物等;菌丝随生长点移动;寄主小麦子房及花药被侵染导致无法成功受精.该真菌侵染小麦后不但影响小麦的正常生理,且在寄主小麦的根、茎、旗叶以及看似正常的成熟籽粒中均发现冬孢子.     

高压电场灭菌效果研究

本文以乳酸杆菌、枯草杆菌及酵母菌为研究对象,研究了高压交流电场作用对它们的致死率影响及细胞结构的变化。实验表明：在22．5kv／cm的场强下处理ls能导致乳酸杆菌活菌数降低近6个数量级,通过显微摄影可明显观察到枯草杆菌细胞结构的破裂和酵母细胞的死亡。     

盐胁迫对芦苇细胞超微结构的影响

于2009年3月从辽宁盘锦双台河口湿地挖取芦苇根茎并人工桶栽,待缓苗成功后进行不同浓度的盐胁迫处理,用透射电镜观察芦苇细胞超微结构对不同盐度胁迫的响应,以明确芦苇细胞的耐盐性。结果表明:芦苇细胞可承受4.0%以下浓度的盐胁迫。当盐度介于0%～4.0%时,芦苇细胞膜系统开始遭到破坏,使芦苇细胞受损的膜结构发生局部内陷或萎缩变形,细胞器表面变得凹凸不平,或将功能丧失的细胞器清理出细胞外,出现破裂和解体,以响应结构损伤的膜系统的修复,使细胞功能得到修复;当盐度为4.0%时,芦苇细胞叶绿体、线粒体、细胞核等具有膜结构细胞器及细胞壁遭到破坏,造成芦苇细胞膜系统的不可逆损伤,使细胞正常的物质代谢与能量转换和信息传递无法完成,导致芦苇细胞新陈代谢过程的中断,芦苇细胞生命活动趋于停止;在8.0%浓度盐胁迫下,芦苇细胞膜系统结构完全消失解体,导致芦苇细胞直接死亡。     

Cell-Wall Changes and Cell Tension in Response to Cold Acclimation and Exogenous Abscisic Acid in Leaves and Cell Cultures

Freeze-induced cell tensions were determined by cell water relations in leaves of broadleaf evergreen species and cell cultures of grapes (Vitis spp.) and apple (Malus domestica). Cell tensions increased in response to cold acclimation in leaves of broadleaf evergreen species during extracellular freezing, indicating a higher resistance to cell volume changes during freezing in cold-hardened leaves than in unhardened leaves. Unhardened leaves, typically, did not develop tension greater than 3.67 MPa, whereas cold-hardened leaves attained tensions up to 12 MPa. With further freezing there was a rapid decline and a loss of tension in unhardened leaves of all the broadleaf evergreen species studied. Also, similar results were observed in cold-hardened leaves of all of the species except in those of inkberry (Ilex glabra) and Euonymus fortunei, in which negative pressures persisted below -40[deg]C. Abscisic acid treatment of inkberry and Euonymus kiautschovica resulted in increases in freeze-induced tensions in leaves, suggesting that both cold acclimation and abscisic acid have similar effects on freezing behavior[mdash] specifically on the ability of cell walls to undergo deformation. Decreases in peak tensions were generally associated with lethal freezing injury and may suggest cavitation of cellular water. However, in suspension-cultured cells of grapes and apple, no cell tension was observed during freezing. Cold acclimation of these cells resulted in an increase in the cell-wall strength and a decrease in the limiting cell-wall pore size from 35 to 22 A in grape cells and from 29 to 22 A in apple cells.     

Effect of microtubule stabilization on the freezing tolerance of mesophyll cells of spinach

Freezing, dehydration, and supercooling cause microtubules in mesophyll cells of spinach (Spinacia oleracea L. cv Bloomsdale) to depolymerize (ME Bartolo, JV Carter, Plant Physiol [1991] 97: 175-181). The objective of this study was to determine whether the LT₅₀ (lethal temperature: the freezing temperature at which 50% of the tissue is killed) of spinach leaf tissue can be changed by diminishing the extent of microtubule depolymerization in response to freezing. Also examined was how tolerance to the components of extracellular freezing, low temperature and dehydration, is affected by microtubule stabilization. Leaf sections of nonacclimated and cold-acclimated spinach were treated with 20 micromolar taxol, a microtubule-stabilizing compound, prior to freezing, supercooling, or dehydration. Taxol stabilized microtubules against depolymerization in cells subjected to these stresses. When pretreated with taxol both nonacclimated and cold-acclimated cells exhibited increased injury during freezing and dehydration. In contrast, supercooling did not injure cells with taxol-stabilized microtubules. Electrolyte leakage, visual appearance of the cells, or a microtubule repolymerization assay were used to assess injury. As leaves were cold-acclimated beyond the normal period of 2 weeks taxol had less of an effect on cell survival during freezing. In leaves acclimated for up to 2 weeks, stabilizing microtubules with taxol resulted in death at a higher freezing temperature. At certain stages of cold acclimation, it appears that if microtubule depolymerization does not occur during a freeze-thaw cycle the plant cell will be killed at a higher temperature than if microtubule depolymerization proceeds normally. An alternative explanation of these results is that taxol may generate abnormal microtubules, and connections between microtubules and the plasma membrane, such that normal cellular responses to freeze-induced dehydration and subsequent rehydration are blocked, with resultant enhanced freezing injury.     

冰冻对珊瑚树(Viburnum odoratissimum)叶片光合效率的影响

光合作用的冰冻伤害研究大多利用离体叶片、原生质体,甚至叶绿体类囊体进行人工冰冻预处理,关于室外自然温度下出现的植物光合作用冰冻伤害,特别是冰冻影响光合量子效率的研究报告还很少。虽然光合碳同化受抑是最早可以测到的冰冻伤害征状,但其机理迄今不清楚(Krause等1982,Krause和Klosson1983)。     

Low temperature-induced modifications in cell ultrastructure and localization of phenolics in winter oilseed rape (Brassica napus L. var. oleifera L.) leaves

Acclimation of winter oilseed plants in the cold (i.e. at temperatures >0 degrees C) followed by short exposure to sub-lethal freezing temperatures resulted in pronounced ultrastructural changes of leaf epidermal and mesophyll cells. The following major changes were observed upon acclimation at 2 degrees C: increased thickness of cell walls; numerous invaginations of plasma membranes; the appearance of many large vesicles localized in the cytoplasm in close proximity to the central vacuole; the occurrence of abundant populations of microvesicles associated with the endoplasmic reticulum (ER) cisternae or located in the vicinity of dictyosomes; and the occurrence of paramural bodies and myelin-like structures. In addition, large phenolic deposits were observed in the vicinity of the plasma membrane and membrane-bound organelles such as chloroplasts, large vesicles or cytoplasm/tonoplast interfaces. Transient freezing (-5 degrees C for 18 h) of the cold-acclimated leaves led to reversible disorganization of the cytoplasm and to pronounced structural changes of the cellular organelles. Chloroplasts were swollen, with the stroma occupying one half of their volume and the thylakoid system being displaced to the other half. Large phenolic aggregates disappeared but distinct layers of phenolic deposits were associated with mitochondrial membranes and with chloroplast envelopes. In frost-thawed cells recovered at 2 degrees C for 24 h, dictyosomes and dictyosome- or ER-derived small vesicles reappeared in the ribosome-rich cytoplasm. Aberrations in the structure of chloroplasts and mitochondria were less pronounced. Few phenolic deposits were seen as small grains associated with chloroplast envelopes and vesicle membranes. These observations demonstrate that plants undergo different changes in cell ultrastructure depending on whether they are subjected to chilling or freezing temperatures. Results are discussed in relation to membrane recycling and the possible role of phenolics during the first and second stages of plant acclimation at low temperature.     

Extracellular freezing in leaves of freezing-sensitive species

Low-temperature scanning-electron microscopy was used to study the freezing of leaves of five species that have no resistance to freezing: bean (Phaseolus vulgaris L.), tobacco (Nicotiana tabacum L.), tomato (Lycopersicon esculentum L.), cucumber (Cucumis sativus L.), and corn (Zea mays L.). In the leaves of the four dicotyledonous species, ice was extracellular and the cells of all tissues were collapsed. In contrast, in maize leaves ice was extracellular in the mesophyll, and these cells were collapsed, but the epidermal and bundle-sheath cells apparently retained their original shapes and volume. It is concluded that the leaves of the freezing-sensitive dicotyledonous species tested were killed by cellular dehydration induced by extracellular freezing, and not by intracellular freezing. Freezing injury in maize leaves apparently resulted from a combination of freezing-induced cellular dehydration of some cells and intracellular ice formation in epidermal and bundle-sheath cells.     

The effect of water, sugars, and proteins on the pattern of ice nucleation and propagation in acclimated and nonacclimated canola leaves

Infrared video thermography was used to observe ice nucleation temperatures, patterns of ice formation, and freezing rates in nonacclimated and cold acclimated leaves of a spring (cv Quest) and a winter (cv Express) canola (Brassica napus). Distinctly different freezing patterns were observed, and the effect of water content, sugars, and soluble proteins on the freezing process was characterized. When freezing was initiated at a warm subzero temperature, ice growth rapidly spread throughout nonacclimated leaves. In contrast, acclimated leaves initiated freezing in a horseshoe pattern beginning at the uppermost edge followed by a slow progression of ice formation across the leaf. However, when acclimated leaves, either previously killed by a slow freeze (2 degrees C h(-1)) or by direct submersion in liquid nitrogen, were refrozen their freezing pattern was similar to nonacclimated leaves. A novel technique was developed using filter paper strips to determine the effects of both sugars and proteins on the rate of freezing of cell extracts. Cell sap from nonacclimated leaves froze 3-fold faster than extracts from acclimated leaves. The rate of freezing in leaves was strongly dependent upon the osmotic potential of the leaves. Simple sugars had a much greater effect on freezing rate than proteins. Nonacclimated leaves containing high water content did not supercool as much as acclimated leaves. Additionally, wetted leaves did not supercool as much as nonwetted leaves. As expected, cell solutes depressed the nucleation temperature of leaves. The use of infrared thermography has revealed that the freezing process in plants is a complex process, reminding us that many aspects of freezing tolerance occur at a whole plant level involving aspects of plant structure and metabolites rather than just the expression of specific genes alone.     

Lipid and protein changes due to freezing in Dunning AT-1 cells

Defining the process of cellular injury during freezing, at the molecular level, is important for cryosurgical applications. This work shows changes to both membrane lipids and protein structures within AT-1 Dunning prostate tumor cells after a freezing stress which induced extreme injury and cell death. Cells were frozen in an uncontrolled fashion to -20 or -80 degrees C. Freezing resulted in an increase in the gel to liquid crystalline phase transition temperature (T(m)) of the cellular membranes and an increase in the temperature range over which the transition occurred, as determined by Fourier transform infrared spectroscopy (FTIR). Thin layer chromatography (TLC) analysis of total lipid extracts showed free fatty acids (FFA) in the frozen samples, indicating a change in the lipid composition. The final freezing temperature had no effect on the thermotropic response of the membranes or on the FFA content of the lipid fraction. The overall protein secondary structure as determined by FTIR showed only slight changes after freezing to -20 degrees C, in contrast to a strong and apparently irreversible denaturation after freezing to -80 degrees C. Taken together, these results suggest that the decrease in viability between control and frozen cells can be correlated with small changes in the membrane lipid composition and membrane fluidity. In addition, loss of cell viability is associated with massive protein denaturation as observed in cells frozen to -80 degrees C, which was not observed in samples frozen to -20 degrees C.     

Protoplasmic Swelling as a Symptom of Freezing Injury in Onion Bulb Cells : Its Simulation in Extracellular KCl and Prevention by Calcium

Freezing injury, in onion bulb tissue, is known to cause enhanced K⁺ efflux accompanied by a small but significant loss of Ca²⁺ following incipient freezing injury and swelling of protoplasm during the postthaw secondary injury. The protoplasmic swelling of the cell is thought to be caused by the passive influx of extracellular K⁺ into the cell followed by water uptake. Using outer epidermal layer of unfrozen onion bulb scales (Allium cepa L. cv Big Red), we were able to stimulate the irreversible freezing injury symptoms, by bathing epidermal cells in 50 millimolar KCl. These symptoms were prevented by adding 20 millimolar CaCl₂ to the extracellular KCl solution. Our results provide evidence that loss of cellular Ca²⁺ plays an important role in the initiation and the progression of freezing injury.     

The role of calcium ions in freezing injuries of winter oilseed rape leaves. Effects of calcium channel blockers and mimesis by calcium ionophore

Calcium ionophore A23187 allowing for a calcium ion influx from an apoplast to a cytoplasm, mimicked symptoms of the frost-induced injuries in winter oilseed rape leaves, as estimated by the conductivity method. Both calcium ionophore and freezing treatment induced degradation of phosphatidylcholine. On the other hand lanthanum and gadolinum ions as well as verapamil, the inhibitors of calcium ion channels, decreased the degree of the frost-induced injuries. Lanthanum ions prevented the frost-induced degradation of PC. It is proposed that freezing alters the functioning of calcium ion channels which results in calcium ion influx into a cytosol. This in turn may lead to a degradation of cell membranes.     

Carbohydrate distribution and cellular injuries in acid rain and cold-treated spruce needles

Summary The cellular structures of acid rain-irrigated needles of several provenances of Norway spruce (Picea abies L. Karst) seedlings were studied after winter experimental freezing. Frost injuries and recovery were characterized by visual damage scoring and classification of mesophyll cell alterations, also using histochemical methods for carbohydrate fluorescent staining. The treatment with-30° C during the late dormancy period was sufficient to cause significant injuries and intracellular degradation in the tissues of the green needles. The most affected seedlings in terms of visual injury scoring were found among those treated with clean water or at pH 3, while freezing injury, defined as an occlusion of phenolic substances in the central vacuole of the mesophyll cells, was most abundant in the needles from spruces irrigated either with clean water or at pH 4 or pH 3. Electron microscopy revealed the details of the injury, e. g. thinning out of the cytoplasm and chloroplast stroma, darkening of the chloroplasts and eventually swelling of the chloroplasts and protoplast. PAS and ConA reactions in the needle tissue revealed intense starch accumulation in the mesophyll and transfusion tissues as early as in March, with a tendency to increase, especially in the untreated needles during the recovery period. Plasma membrane disturbances were indicated by histochemical identification of callose deposits in the mesophyll cell walls, these being most abundant in the acid rain-treated needles. All these findings suggest that freezing at –30° C was more deleterious to the seedlings pretreated with acid or clean water than to those not given additional irrigation.