Phytotoxicity of selected herbicides to mung bean (Phaseolus aureus Roxb.)

Mung bean (Phaseolus aureus Roxb.) is grown after harvest of wheat during the fallow period. Herbicides such as metsulfuron, atrazine and isoxaflutole are recommended to control weeds in wheat–rice cropping system including weeds of fallow crop. The effects of three herbicides with different modes of action—atrazine, photosystem II inhibitor; metsulfuron, acetolactate synthase inhibitor; and isoxaflutole, 4-hydroxyphenylpyruvatedioxygenase inhibitor—on shoot height, chlorophyll concentrations and cellular damage in herbicide-treated mung bean were studied. While isoxaflutole inhibited shoot growth and chlorophyll concentration of mung bean, atrazine and metsulfuron did not cause reduction in the shoot growth of mung bean. Metsulfuron (226, 452, 1356 and 2260 μg/kg soil) and isoxaflutole (452, 1356 and 2260 μg/kg soil) in soil reduced the concentration of leaf chlorophyll of mung bean compared to the control. Atrazine in soil did not affect the total chlorophyll concentration of mung bean leaves. Electron micrographs showed that untreated mung bean had elongated chloroplasts, thylakoids organized as intact grana, distinct starch grains and a small number of plastoglubuli. Mesophyll cells of atrazine-treated mung bean leaves had swollen chloroplasts and thylakoids with disorganized grana. Leaves of metsulfuron-treated mung bean had swollen chloroplasts with a large number of starch grains. Starch grains were not observed in leaves of mung bean treated with either atrazine or isoxaflutole. Complete disruption of thylakoids was observed in isoxaflutole-treated mung bean leaves. Leaves of atrazine-treated mung bean showed detached microfibrils along with distorted and degenerated secondary walls. Metsulfuron-treated mung bean leaves showed aggregated microfibrils with completely dissolved secondary walls, while isoxaflutole-treated leaves had completely degenerated secondary walls with complete loss of microfibrils. We conclude that isoxaflutole at higher doses, influence mung bean at the morphological, physiological and cellular levels.     

Leaf anatomy and chloroplast ultrastructure of Mn-deficient orange plants

Leaf samples of Mn-deficient and Mn-sufficient (control) ‘Navelate’ orange plants grown in a greenhouse were taken to investigate the effects of Mn deficiency in leaf structure and chloroplast ultrastructure. Total leaf chlorophyll concentration was significantly lower in Mn-deficient plants than in control ones. Entire lamina thickness was not altered due to Mn deficiency. However, Mn deficiency resulted in disorganization of mesophyll cells, mainly of palisade parenchyma cells. The number of mesophyll chloroplasts per cellular area and their length were both affected negatively. The membranous system of chloroplasts was also disorganized. The percentages of starch grains and plastoglobuli per chloroplast of Mn-deficient leaves were significantly greater than those of control leaves.     

Experimental complexities in evaluating the comparative phytotoxicity of chemicals with different modes of action

Careful attention should be paid to bioassay experiments to examine the comparative phytotoxicity of chemicals with different modes of actions. Experimental complexities in examining comparative phytotoxicity of chemicals with differing modes of action are determined and discussed to appreciate the importance of relevant secondary effects that can be quickly measured. The chemicals selected were: benzoic acid, isoxaflutole and rimsulfuron. Data on shoot and root length of 7-day-old mustard (Brassica napus L.) seedlings, shoot height of 4-week-old mustard plants and total leaf chlorophyll concentrations were determined when plants were grown in soil treated with different concentrations of the three chemicals. Scanning electron microscope (SEM) studies were conducted to determine any damage to mustard root hairs after treatment. Root growth of 7-day-old mustard seedlings was reduced when treated with different concentrations of benzoic acid or rimsulfuron. Root growth of mustard seedlings, however, largely remained unaffected when plants were grown in soil treated with isoxaflutole. While no significant reduction in either chlorophyll concentration or shoot height of 4-week-old mustard plants was observed when treated with soil-applied benzoic acid, both parameters were inhibited when mustard plants were treated with isoxaflutole or rimsulfuron. SEM studies revealed significant damage to root hairs in a 7-day-old mustard seedlings when plants were grown in soil treated with 500, 1000 and 2000 mg/L benzoic acid, and 0.5 mg/L rimsulfuron. No such damage was observed when mustard was grown in soil treated with isoxaflutole.     

The effect of soil pH on chickpea (Cicer arietinum) genotype sensitivity to isoxaflutole

A glasshouse experiment was conducted to investigate the effect of soil pH on chickpea (Cicer arietinum) tolerance to isoxaflutole applied pre-emergence at 0, 75 (recommended rate) and 300 g a.i. ha⁻¹. For this study, the variables examined were two desi chickpea genotypes (97039-1275 as a tolerant line and 91025-3021 as a sensitive line) and four pH levels (5.1, 6.9, 8.1, and 8.9). The results demonstrated differential tolerances among chickpea genotypes to isoxaflutole at different rates and soil pH levels. Isoxaflutole applied pre-emergence resulted in increased phytotoxicity with increases in soil pH and herbicide rate. Even the most tolerant chickpea genotype was damaged when exposed to higher pH and herbicide rates, as indicated by increased leaf chlorosis and significant reductions in plant height, and shoot and root dry weight. The effects were more severe with the sensitive genotype. The susceptibility of chickpea to this herbicide depends on genotype and soil pH which should be taken into account in breeding new lines, and in the agronomy of chickpea production.     

Effect of Rice Straw Incorporation on Phytotoxicity of Isoxaflutole to an Exotic Weed Phalaris minor Retz.

Phalaris minorRetz. is a major exotic annual weed in the wheat (Triticum aestivum L.) crop. Unharvested rice (Oryza sativa L.) straw, unburned and burned, is often incorporated in the field prior to cultivating wheat. Isoxaflutole (Balance), a pre-emergent systemic soil applied herbicide, has potential to control P. minor. Glasshouse experiments were conducted to determine the phytotoxicity of isoxaflutole defined by reductions in relation to shoot length of P. minor when grown in unamended soil or soil amended with unburned or burned rice straw. A 120 g soil was amended with 0, 1, 2 and 4 g of unburned or burned rice straw, and placed in 150 mL styrofoam pots. Appropriate amount of isoxaflutole (75% active ingredient, ai) was added to pots to get final concentration of 0, 7.5, 30, 60 and 120 μg ai/pot. Unamended soil and soil amended with unburned or burned rice straw were analyzed for pH and organic matter; two important determinants of isoxaflutole activity. Results indicate a significant reduction in shoot length of P. minor when grown in soil treated with isoxaflutole at 30, 60 or 120 μg ai/pot. Inhibition in the shoot length of P. minor was observed when soil amended with unburned straw was treated with isoxaflutole at 7.5 and 30 μg ai/pot compared with unamended soil treated with similar amounts of isoxaflutole. No significant change in isoxaflutole toxicity was observed when soil amended with unburned straw was treated with isoxaflutole at 60 and 120 lg ai/pot compared with unamended soil treated with similar amounts of isoxaflutole. Isoxaflutole phytotoxicity to P. minor shoot length was eliminated when soil amended with burned straw was treated with isoxaflutole at 7.5 and 30 μg ai/pot. P. minor shoot length was greater when soil amended with burned straw was treated with isoxaflutole at 60 and 120 μg ai/pot relative to herbicide-treated unamended soils. We conclude that incorporation of burned rice straw greatly reduces the phytotoxicity of isoxaflutole toP. minor.     

Electron microscope observations on the differentiation of protocorm cells of Dactylorhiza maculata

The fine structure of aseptically cultivated protocorm cells of Dactylorhiza maculata (L.) Sod was studied with special reference to differentiation of leaf cells. In the young protocorm the characteristic structures are compound amyloplasts and smooth ER. In the older, the green shoot pole contains many proplastids storing starch, some membrane–bound protein crystalloids and lipid droplets attached to the nuclear membrane and plasmalemma. In the leaves of the young seedling, two types of chloroplasts are found. One contains little or no starch but many osmiophilic plasto–globuli; the other has only a few plastoglobuli but a large starch grain.     

Ultrastructural and Morphological Characteristics of Cultivated Wheat Growing on Copper-Polluted Fields

A cultivated Greek variety of wheat (Triticum aestivum L. cv. Vergina) growing in fields naturally polluted by outcrops of copper ores was investigated. Wheat plants show a negative response to increasing quantities of soil copper, including reduced growth and chlorosis. Copper toxicity was demonstrated in the laboratory by a rooting test; the frequency of mitoses declines sharply with increasing copper concentration in the nutrient solution. The mesophyll cells of polluted plants display a circular shape (in transverse sections) with a few chloroplasts parietally distributed, in contrast with the elongate or pleomorphic shape of control leaves that contain numerous chloroplasts crowded at the cell periphery. Ultrastructurally, the chloroplasts of polluted plants contain a poorly developed internal membrane system consisting of thylakoids arranged parallel to each other with only a few, rudimentary grana. In addition, a number of statistically significant differences were found, including the number of starch grains and plastoglobuli, chloroplast surface area, volume fraction of starch grains and, most important, the volume fraction of the internal membrane system. All ultrastructural changes are attributed to the toxic effect of high concentrations of soil copper.     

Morphometric Analysis of Chloroplasts of Cotton Leaf and Fruiting Organs

We examined morphological and ultrastructural differences in chloroplasts of cotton leaves and the fruiting organs, bract, and capsule wall to advance our understanding of their commonly observed differences in photosynthetic efficiency. Chloroplasts from leaves were large (7.1 μm long in cross section), lens shaped with a well developed membrane system differentiated into grana and stroma lamellae that occupied the large cross-sectional area (12.3 μm²) of the organelle. A few small plastoglobuli and starch grains were scattered in the stroma region. The bract chloroplasts were correlative of leaf chloroplasts in size (6.8 μm in length) and shape with the exception that the bract chloroplasts exhibited greater thylakoid number per granum (15.8) than the leaf chloroplasts (10.5). In contrast to leaf and bract, the capsule wall chloroplasts were smaller in size (4.3 μm) and cross sectional area (6.8 μm²) than either the leaf or bract. The most intriguing feature of the capsule wall chloroplasts was its domination by large starch granules (5.3 μm²) in the stroma which filled the whole chloroplast coercing the membrane system to move towards the periphery of the organelle. Grana number and thylakoids per granum were lowest in the capsule wall chloroplasts. This revised version was published online in July 2006 with corrections to the Cover Date.     

油菜类型和品种对外来入侵杂草小子虉草的替代控制作用

外来入侵植物小子虉草（Phalaris minor Retz.）是世界公认的冬季农田恶性杂草,掌握农作物对其替代控制作用具有重要的研究价值。前期研究表明,油菜是替代控制小子虉草的优良农作物,然而,目前尚不清楚油菜类型与品种对其控制能力的影响。为此选取与小子虉草同域发生的不同类型（白菜型油菜、芥菜型油菜和甘蓝型油菜）油菜品种各3种,通过田间小区实验和室内化感作用测定,对比研究其对小子虉草的生长、繁殖、表型以及化感作用的影响。田间实验显示：竞争方式（种内或种间竞争）和油菜类型对小子虉草的地上生物量、种子数、株高、分枝数、叶面积和比叶面积存在极显著（P=0.0001）影响;而油菜品种对小子虉草的地上生物量（P=0.6064）、种子数（P=0.3577）、株高（P=0.4279）、分枝数（P=0.6357）、叶面积（P=0.8839）和比叶面积（P=0.3424）均无显著影响。3种类型油菜对小子虉草生长、繁殖以及表型的影响存在明显差异,其中芥菜型油菜对小子虉草的上述指标的影响最强,而白菜型油菜的影响最弱。室内生物测定显示,油菜对小子虉草具有化感抑制作用,当供试油菜叶片水提液浓度为0.1 g/mL时,小子虉草种子的萌发和幼苗的株高、根长、生物量均被显著抑制;研究也表明不同类型油菜对小子虉草的化感作用显著不同,同等条件下,芥菜型油菜对小子虉草的化感抑制作用最强。综上所述,油菜类型对外来入侵小子虉草的控制作用存在显著差异,其中芥菜型油菜对植物小子虉草的替代控制作用明显优于白菜型油菜和甘蓝型油菜,而其强的化感抑草特性或许是其强控草能力的原因之一。另外,本研究也为进一步利用油菜替代控制入侵植物小子虉草提供了参考。     

Effect of Glycinebetaine on Chloroplast Ultrastructure, Chlorophyll and Protein Content, and RuBPCO Activities in Tomato Grown under Drought or Salinity

The effect of foliar-applied glycinebetaine (GB) on chloroplast ultrastructure, the amount of chlorophyll and proteins and ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPCO) activity in stressed tomato leaves were analysed. Initial, and total activity and activation state state of RuBPCO were also measured. RuBPCO activities, chloroplast area, and the number of plastoglobuli remained unaffected, while the relative area of starch granules increased in GB-treated, salt-stressed tomato leaflets. Under drought-stress, the relative area of plastoglobuli increased upon GB application. The primary effect of GB was, however, the increased protein and chlorophyll content.     

Cultivar Differences in Growth and Chloroplast Ultrastructure in Rice as Affected by Nitrogen

Plant growth, leaf protein and chlorophyll content, and chloroplastultrastructure as affected by nitrogen (N) were examined infour rice (Oryza sativa L.) cultivars grown in culture solutionunder controlled conditions. Increasing N concentration generallyincreased height and shoot dry weight of all cultivars. Cultivardifferences were significant at normal N level (40 ppm). Amongcultivars, IR8 was most responsive to increasing N, having thesignificantly highest shoot dry weight and protein content.Total chlorophyll and protein contents varied with cultivarand N, but chlorophyll a/b ratio remained constant. At the ultrastructurallevel, chloroplasts had generally well-developed grana and stromalamellae at 40 ppm.N. Chloroplasts at high N had from one tofour times as many grana as the N-deficient chloroplasts. Nitrogendeficiency reduced the size of the chloroplast, grana-stromalamellae and resulted in fewer poorly stacked grana. Increasingthe N level (120 ppm) above the normal level did not significantlyaffect chloroplast size of any cultivar, except for IR8 whichhad the largest chloroplast. A reduction in the number of starchgrains was observed in IR8, but more were present in ER36 underN-deficient conditions. The size of starch grains was not affectedby N and did not differ among cultivars. Plastoglobuli appearedto be larger under N-deficient conditions. Nitrogen had no effecton the number of plastoglobuli but cultivar differences existed.The highly N-responsive IR8 (based on dry weight) had the largestchloroplast which increased with N level. The increase in chloroplastsize accounted for the increase in both chlorophyll and proteincontents and, consequently, dry weight. Key words: Oryza sativa L., chloroplast, chlorophyll, protein     

锌胁迫对不知火杂柑和椪柑叶片光合特性及超微结构的影响

采用砂基培养法,运用CI－340便携式光合测定仪、H-7650透射电子显微镜研究了不同锌浓度如0mg／L（锌缺乏）、0．05mg／L（对照）和0．5mg／L（锌过量）处理下不知火杂柑（简称不知火）和槿柑叶片叶绿体色素含量、净光合速率及超微结构的变化。结果表明：（1）锌缺乏处理的不知火及锌过量处理的槛柑叶片叶绿素（a＋b）和类胡萝卜素含量、锌缺乏与锌过量处理的两者叶片净光合速率均相对低于对照。（2）锌缺乏处理下,两者叶片叶绿体出现变形或空室化,其中碰柑叶绿体内淀粉粒和质体小球增多,细胞核及线粒体正常,而不知火淀粉粒少,细胞核中出现许多黑色小颗粒物质。锌过量处理下,两者叶片叶绿体出现空室化或叶绿体膜模糊,线粒体解体,其中槿柑叶绿体内质体小球增多,淀粉粒少;而不知火淀粉粒明显增多、增大,基粒片层膨胀、松弛。可见,锌胁迫对两者叶片叶绿体色素、净光合速率及超微结构有着明显的影响,其影响程度因品种而异。     

Changes in the plastid ultrastructure duringSedum rotundifolium leaf development

The ultrastructure of plastids was investigated in succulent leaves ofSedum rotundifolium to examine their changes during development. Leaves were categorized as etiolated, immature, young, and mature, based on their developmental stage and size. Of particular interest were the features of the tubular inclusion bodies (TIBs) and starch grains. These, along with vacuole size, showed remarkable changes over time. Etioplasts of unexposed leaves had prolamellar bodies, abundant starch grains, large TIB, few plastoglobuli, and thylakoid systems. Membranes of the thylakoids were still continuous with those of the prolamellar body. The plastids were often influenced by the presence and profile of inclusion bodies and starch grains throughout the early stages. Morphology was highly variable in the etioplasts but consistently hemispherical or ovoid in mature chloroplasts. TIB was most abundant in the etiolated leaves, but disappeared completely with development. Starch grains also became significantly reduced in size. Both young and mature mesophyll cells exhibited a normal chloroplast ultra-structure and huge central vacuoles, with an extremely thin peripheral cytoplasm. Grana were extensive and comprised a large portion of the chloroplasts. Traces of peripheral reticulum were also discovered in the chloroplasts of expanded leaves. The implications of these ultrastructural changes in the tubular inclusions and starch grains are discussed with relevance to Crassulacean acid metabolism (CAM).     

Effects of drought and waterlogging on ultrastructure of Scots pine and Norway spruce needles

Effects of water stress on needle ultrastructure of 2-year-old Scots pine (Pinus sylvestris L.) and 5-year-old Norway spruce [Picea abies (L.) Karst.] seedlings were studied in greenhouse experiments. Drought stress was induced by leaving seedlings without watering, and waterlogging stress was produced by submerging the seedling containers in water. Needle samples for ultrastructural analyses were collected several times during the experiments, and samples for nutrient analyses at the end of the experiments. In drought stress, plasmolysis of mesophyll and transfusion parenchyma tissues, aggregation of chloroplast stroma and its separation from thylakoids and decreased size and abundance of starch grains in needles of both species were observed. The concentration of lipid bodies around the chloroplasts were detected in pine needles. Calcium and water concentrations in spruce needles were lower by the end of the experiments compared to controls. In waterlogging treatment, swelling of phloem cells in pine needles and large starch grains, slight swelling of thylakoids and increased translucency of plastoglobuli in chloroplasts of both species studied were observed. The phosphorus concentration in pine needles was higher while phosphorus, calcium and magnesium concentrations in spruce needles were lower in the waterlogging treatments compared to controls. Typical symptoms induced by drought stress, e. g. aggregation of chloroplast stroma and its separation from thylakoids, were detected, but, in waterlogging stress, ultrastructural symptoms appeared to be related to the developing nutrient imbalance of needles.     

葡萄属植物叶片显微和超微结构观察

利用光学显微镜和透射电镜,对葡萄属(Vitis L.)15种、1亚种和4栽培品种植物的叶片显微和超微结构进行了观察研究。结果显示:(1)东亚种群中桦叶葡萄的叶片总厚度和各组织厚度最大;北美种群的河岸葡萄和沙地葡葡叶片总厚度、上表皮厚度、栅栏组织厚度、海绵组织厚度和下表皮厚度较大,但叶片组织紧密度最小。(2)各种类间栅栏组织中叶绿体数目均多于海绵组织,且刺葡萄、桦叶葡萄、葛藟葡萄和秋葡萄叶肉细胞中叶绿体数目最多,北红、蘡薁、网脉葡萄和菱叶葡萄叶片中叶绿体数目最少,巨峰和桦叶葡萄叶绿体中含有大量的淀粉粒,桦叶葡萄、网脉葡萄、秋葡萄、葛藟葡萄、刺葡萄和华东葡萄含有较多的嗜锇颗粒。研究表明,中国野生葡萄的许多种类在叶片组织紧密度、叶绿体数量、淀粉粒数量、嗜锇颗粒数量方面均高于北美种群的河岸葡萄和沙地葡萄,具有更高的结构抗性基础。     

Fatty acid synthesis by slices from developing leaves

Fatty acid synthesis was studied in successive leaf sections from the base to the tip of developing barley (Hordeum vulgare L.), maize (Zea mays L.), rye grass (Lolium perenne L.) and wheat (Triticum aestivium L.) leaves. The basal regions of the leaves had the lowest rates of fatty acid synthesis and accumulated small amounts of very long chain fatty acids. Fatty acid synthesis was highest in the middle leaf sections in all four plants. Linolenic acid synthesis from [1-¹⁴C]acetate was highest in the distal leaf sections of rye grass. The labelling of the fatty acids of individual lipids of rye grass was examined and it was found that [¹⁴C]linolenic acid was highest in the galactolipids. Synthesis of this acid in the galactolipids was most active in leaf segment C. Only traces of [¹⁴C]linolenic acid were ever found in phosphatidylcholine and it is concluded that this phospholipid cannot serve as a substrate for linoleic acid desaturation in rye grass. The synthesis of fatty acids was sensitive to arsenite, fluoride and the herbicide EPTC. The latter was only inhibitory towards those leaf segments which made very long chain fatty acids. Formation of fatty acids from [1-¹⁴C]acetate was also studied in chloroplasts prepared from successive leaf sections of rye grass. Chloroplasts isolated from the middle leaf sections had the highest activity. Palmitic and oleic acids were the main fatty acid products in all chloroplast preparations. Linolenic acid synthesis was highest in chlorplasts isolated from the distal leaf sections of rye grass.     

Morphological and quantitative data of plastids and mitochondria within drought-stressed spinach leaves

Summary. Selected cell organelles were investigated at a high level of resolution with the transmission electron microscope, using ultrathin serial sections to create three-dimensional reconstructions. On the basis of these reconstructions, morphological data of chloroplast fine structures, mitochondria, and peroxisomes from control and drought-stressed spinach leaves were evaluated and compared. Mesophyll cell chloroplasts of control plants contained 60% stroma, 23% thylakoids, and 16% starch. In drought-stressed plants, the volume of both the stroma and the thylakoids increased to 68% and 32%, respectively. The amount of plastoglobuli was about 0.3% in both samples. Chloroplasts of stressed plants differed from control plants not only in the thylakoid and stroma values but also in the lack of starch grains. Mitochondria occurred in variable forms in control and stressed samples. In stressed plants, mitochondria had only 65% of the volume compared with control plants. Peroxisomes were inconspicuous.     

Combined use of 13C- and 19F-NMR to analyse the mode of action and the metabolism of the herbicide isoxaflutole

¹³C-Nuclear magnetic resonance was used to characterize the effects of the trifluoromethyl herbicide isoxaflutole on plant metabolism. A specific accumulation of tyrosine and phenylalanine in pea leaves was observed after treatment with this herbicide, consistent with an inhibition of 4-hydroxyphenylpyruvate dioxygenase (HPPD), an enzyme involved in plastoquinone and tocopherol synthesis and the known target of isoxaflutole. ¹⁹F-NMR permitted the analysis of the uptake and degradation of isoxaflutole, with detection limits around 1 μM in perchloric acid extracts. Isoxaflutole was incorporated and transformed to an active diketonitrile derivative, which further underwent degradation to an inactive benzoic acid derivative.     

Changes of Anatomical Features, Photosynthesis and Ribulose Bisphosphate Carboxylase-Oxygenase Content of Mango Leaves

Changes in anatomical and physiological features, includingchanges in amount per unit area of anthocyanin and chlorophyll,in leaves of seedling mango (Mangifera indica L. cv. Irwin)trees were determined to understand what controls the rate ofphotosynthesis (Pn) at various stages of development. The youngleaves of seedling trees contained high concentrations of anthocyanin.During enlargement of leaves, the disappearance of anthocyaninand the accumulation of chlorophyll occurred concomitantly;the anthocyanin content began to decrease markedly once theleaf area had reached a maximum. During the early period ofleaf development, the thickness of mesophyll tissue decreasedtemporarily, but when the length of the leaf reached half thatof a mature leaf, the mesophyll began to thicken again. Smallstarch grains appeared in the chloroplasts of the young leavesand chloroplast nucleoids (ct-nuclei) were distributed throughoutthe chloroplasts. When leaves matured, ct-nuclei were displacedto the periphery of chloroplasts because of the accumulationof large starch grains. Compared with young leaves, green andmature leaves contained greater concentrations of ribulose bisphosphatecarboxylase-oxygenase (RuBisCO) protein. The results of immunocytochemicalexamination of RuBisCO under the light microscope reflectedthe results of electrophoresis measurements of RuBisCO. Pn waslow during the chocolate-coloured stage of early leaf development.In green and mature leaves Pn was higher; the average Pn was7·6 mg CO₂ dm^-2 h^-1 under light at intensities above500 µmol m^-2 s^-1.Copyright 1995, 1999 Academic Press Mangifera indica L., mango leaf, chloroplast nucleoids, chloroplast ultrastructure, starch accumulation, anthocyanin, chlorophyll, DAPI staining, SDS-PAGE, immunocytochemical technique