The structure and hardening status of Scots pine needles at different potassium availability levels

 Scots pine (Pinus sylvestris L.) seedlings were exposed to three levels of potassium (low, medium and high) and their needle morphology, the cellular structure of the mesophyll and transfusion parenchyma, and the hardening status of the mesophyll cells were examined by light and transmission electron microscopy. The higher the potassium level the greater was the growth of the needles. The area of the mesophyll tissue increased slightly and those of the phloem, xylem and resin ducts decreased in the needles of the seedlings grown at the high K level. Cellular studies revealed that swelling of the chloroplast thylakoids, accumulation of starch in the chloroplasts, translucency of the cytoplasm and plasmolysis in the mesophyll cells were related to a low K level. The hardening status of the mesophyll cells was enhanced after 5 weeks of hardening treatment at high K as seen in changes in chloroplast shape and position and the structure of the endoplasmic reticulum, but the pines showed no major differences in the hardening status of their mesophyll cells between K levels at the end of the experiment, after 9 weeks of hardening. Frost resistance, as shown by the electrolyte leakage test, was nevertheless highest at low K, being related to the increase in the concentration of polyamine putrescine at this potassium level. Received: 23 December 1997 / Accepted: 30 March 1998     

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.     

The rapid yellowing of spruce at a mountain site in the Central Black Forest (Germany). Combined effects of Mg deficiency and ozone on biochemical, physiological and structural properties of the chloroplasts

Biochemical, physiological and ultrastructural changes of the chloroplasts were examined in the course of the rapid yellowing process of spruce (Picea abies (L.) Karst.) at a Mg-deficient and ozone polluted mountain site (Sch?llkopf mountain, Central Black Forest, Germany, 840 m a.s.l.). While at an early stage of yellowing the chlorophyll (Chl) content of the needles decreased slowly, significant changes occurred in the chloroplasts: The lability of the light-harvesting Chl a/b protein complex LHC II increased; the thylakoid cross-sectional area of chloroplasts in the outer mesophyll of the needles decreased, and their Chl fluorescence showed typical changes like the decrease of Fv/Fm and the increase of the photoinhibitory Fv quenching. Later on, the Chl content decreased rapidly, the changes in the chloroplasts continued and the needles turned yellow. Lutein and the pigments of the xanthophyll cycle were enhanced in relation to Chl a. Light and dark reactions of the xanthophyll cycle were highly active indicating efficient proton pumping and NADPH formation. The ratio of nonappressed to appressed thylakoid membranes increased with decreasing Fv/Fm suggesting that structural and fluorescence properties of the chloroplasts were related. The response of the needles to defined shading and improved Mg supply was also examined. The combined effects of strong sun light, low levels of non-Chl-bound Mg (Mg(free)) and ozone concentrations exceeding 80 microg m(-3) are shown to be necessary to induce the rapid yellowing process. For needles with Mg(free) < 0.12 mg g(-1) needle dry matter, the lability of the LHC II was correlated with the ozone concentration suggesting that the destabilization of the LHC II plays a central role in the rapid yellowing process.     

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.     

ABA诱导离体红松针叶衰老过程中叶肉细胞超微结构的变化

ABA处理离体针叶,使RNase活性明显提高,针叶变黄加剧以及叶肉细胞超微结构有序衰老,证明ABA可以加速江松针叶的衰老。作者以红松(Pinus koraiea sis sieb et Zucc)针叶叶肉细胞各细胞器对衰老的敏感程度为序,将针叶衰老过程划分衰老早期、衰老中期及衰老后期。     

盐胁迫对玉米叶片叶肉细胞生物膜超微结构的影响

研究了NaCl胁迫对玉米叶肉细胞生物膜超微结构的影响. 结果表明：NaCl胁迫破坏了玉米叶片叶肉细胞生物膜的正常结构,50 mmol·L^-1 NaCl处理胁迫下,玉米叶肉细胞核膜,线粒体膜,细胞膜,叶绿体膜,液泡膜都受到不同程度的破坏,叶绿体基粒类囊体膨胀,间质片层空间增大,片层紊乱。100 mmol·L^-1 NaCl处理胁迫下,质膜,液泡膜,线粒体,叶绿体都受到严重的破坏。细胞质膜破坏,破损的叶绿体充斥在细胞间隙中;叶绿体外膜破坏,甚至解体消失,叶肉细胞中充满膜结构,基粒排列方向改变,垛叠层数减少,基粒和基质片层界限模糊不清,有的基粒解体消失,甚至叶绿体完全解体;核膜破坏、解体,核中的染色质高度凝缩;线粒体的数量增多,线粒体膜破坏,脊的数量减少,甚至整个线粒体破损解体;液泡膜破坏;由于各种生物膜的破坏,使细胞内充满许多囊状小泡、多泡体或斑层小体;叶肉细胞发生严重的质壁分离,严重时发生细胞壁断裂;甚至整个细胞溶解。     

LEAF OF SPINACIA OLERACEA (SPINACH): ULTRASTRUCTURE,AND PLASMODESMATAL DISTRIBUTION AND FREQUENCY,IN RELATION TO SIEVE-TUBE LOADING

Minor veins and contiguous tissues of the Spinacia oleracea leaf were analyzed by electron microscopy to determine the characteristics of the component cells and the structure, distribution, and frequency of plasmodesmata between the various cell types of the leaf. Mesophyll and bundle-sheath cells contain components typical of photosynthetic cells although the latter cell type contains smaller chloroplasts and fewer mitochondria and microbodies than the mesophyll cells. In addition, the mesophyll cells contain numerous invaginations of the plasmalemma bordering the chloroplasts and evaginations of the outer membrane of the opposing chloroplast envelope. In places, these membranes appear continuous with each other. The minor veins consist of tracheary elements, xylem parenchyma cells, sieve-tube members, companion and phloem parenchyma cells, and other cells simply designated vascular parenchyma cells. The companion and phloem parenchyma cells are typically larger than the sieve-tube members with the companion cells containing a much denser cytoplasm that the phloem parenchyma. Cytoplasmic connections occur along all possible routes from the mesophyll to the sieve-tube members and consist of either simple or branched plasmodesmata between parenchymatic elements or pore-plasmodesmata between the sieve-tube members and parenchyma cells. The highest frequency of plasmodesmata occurs between the sieve-tube members and companion cells, although the value is essentially the same as between the various parenchymatic elements of the phloem. Compared to several previously studied species, the frequency of plasmodesmata between cell types of the spinach leaf is low. These results are discussed in relation to apoplastic vs. symplastic solute transport and sieve-tube loading in this species.     

葛(豆科)叶的解剖学研究

利用光学显微镜和扫描电镜观察了葛(Pueraria lobata)叶的解剖学特征。结果表明,葛叶片的上、下表皮都只有一层表皮细胞,上表皮比下表皮厚。上、下表皮都有腺毛和非腺毛。气孔主要分布在下表皮,下表皮的气孔密度为(261±17)mm-2,上表皮只有(6±3)mm-2。叶肉由两层栅栏组织细胞和一层海绵组织细胞构成。叶肉细胞中有丰富的叶绿体。在栅栏组织和海绵组织之间有一层平行于叶脉的薄壁细胞。叶脉中含有大量的草酸钙晶体。葛叶的这些形态特征与其喜阳、耐旱的特点相适应。     

Structural changes in the vascular bundles of light-exposed and shaded spruce needles suffering from Mg deficiency and ozone pollution

The correlation between structural changes of the vascular bundles and needle yellowing was examined for needles of damaged spruce (Picea abies (L.) Karst.) growing at a Mg-deficient and ozone polluted mountain site in the Central Black Forest (840m a.s.l.). In the previous year's sun-exposed needles, the following sequence of events was observed: (1) rapid needle yellowing, (2) hypertrophy and anomalous divisions of cambium cells, (3) phloem collapse, and, (4) production of atypical xylem tracheids. Under defined shade (reduction of the photosynthetically active photon flux density of the ambient light by 85-90%), the needles remained green, while the phloem collapsed completely within the first 6 weeks of shading; subsequently, a reversal of the collapse was observed. Under both light conditions, the content of Mg not bound to chlorophyll (Mg(free)) was in the range of 0.1 mg g(-1) needle dry matter, and hardly changed throughout the investigation period. After Mg fertilization, the Mg(free) level of the previous year's needles increased to 0.2 mg g(-1) dry matter, the light-exposed needles remained green, and the vascular bundles developed no anomalies. The data show that the rapid needle yellowing of ozone-exposed Mg-deficient needles did not depend on the collapse of the phloem. Mg deficiency played a key role in the development of anomalous vascular bundles under light, and also appears to explain the transient changes in sieve cell structure under shade. The role of Mg deficiency, rather than ozone pollution, in the damage of the sieve cells was confirmed in a long-term ozone exposure experiment with young clonal spruce growing under defined conditions.     

Cytological Observations of the Infection Process by Phomopsis helianthi (Munt.-Cvet) in Leaves of Sunflower

The infection process of Phomopsis helianthi and the specific degradation of infected tissue were studied in detail using light and transmission electron microscopy. In comparison with other vascular pathogens, the infection and degradation process was in some aspects different. The favourite tissue for the pathogen to grow in was the phloem. Parenchymatic cells in and around vascular bundles were extremely sensitive to infection long before hyphae arrived, probably due to a toxin. In the parenchymatic cells the first changes were visible at the chloroplasts where electron-dense material accumulated in the thylakoid space. The chloroplast stroma changed contrast and later the whole cytoplasm also appeared electron dense. In the vascular bundles, first the phloem was destroyed and then hyphae invaded the adjacent mesophyll, the cambium, and finally the vessel elements. In particular, the compact mesophyll of the midvein was severely affected. Vessel elements were lined with electron-dense material and some were filled with flocculent material. Severe wall destruction indicated the action of a complete set of cell wall-degrading enzymes before hyphae entered the tissue; it always started at the innermost wall layer. Wall degradation in vascular tissue and adjacent parenchyma with intercellular spaces was different. Before the degradation of the protoplasts started, the cell walls were completely metabolized and only the secondary walls of the vessels resisted for longer. There were no host–cell reactions visible that could be interpreted as a defence reaction.     

Two different modes of early development and nitrogen assimilation in gymnosperm seedlings†

Light-independent chloroplast development and expression of genes encoding chloroplast proteins occur in many but not all species of gymnosperms. Early development in maritime pine (Pinus pinaster) seedlings was strongly light-independent, whereas Ginkgo biloba seedlings exhibited a typical angiosperm-like morphogenesis with differentiated patterns in light and dark. In pine, chloroplast polypeptides were undetectable in the seed embryo and accumulated in cotyledons of both light- and dark-grown plants in good correlation with light-independent chlorophyll synthesis. In contrast, chlorophyll and chloroplast proteins were only detected in light-grown ginkgo. Pine cytosolic glutamine synthetase (GS) and ferredoxin glutamate synthase (Fd-GOGAT) were present at low levels in the seeds and accumulated at comparable amounts in light- and dark-grown seedlings. Fd-GOGAT was also barely detectable in the seeds of ginkgo and only accumulated in green plants with mature chloroplasts. In G. biloba seeds and etiolated plants only cytosolic GS was identified, while in light-grown seedlings this molecular form was present at low abundance and choroplastic GS was the predominant isoenzyme. The above results have been confirmed by immunolocalization of GS protein in pine and ginkgo plantlets. In pine, GS was present in the peripheral cytoplasm of mesophyll cells and also in the phloem region of the vascular bundle. Immunocytochemical analysis showed that the labelling of mesophyll and phloem cells was only cytoplasmic. In developing ginkgo, GS antigens were present in the chloroplasts of mesophyll parenchyma cells of leaflets and green cotyledons. In contrast, a weak labelling of GS was observed in the parenchyma and phloem cells of non-green cotyledons enclosed in the seed coat. Taking all this into account, our data indicate the existence of two different modes of GS and GOGAT regulation in gymnosperms in close correlation with the differential response of plants to light. Furthermore, the results suggest that glutamine and glutamate biosynthesis is confined to the chloroplast of mesophyll cells in species with light-dependent chloroplast, development whereas compartmentation would be required in species with light-independent plastid development.     

Ageing-related Anatomical and Ultrastructural Changes in Leaves of Birch (Betula pendula Roth.) Clones as Affected by Low Ozone Exposure

Effects of ozone on the leaf anatomy and ultrastructure of fivebirch (Betula pendula Roth.) clones were studied during onegrowing season in open-field conditions. Cumulative ozone exposurewas 1·5 times higher than ambient. Ozone exposure decreasedtotal leaf thickness in one, ozone sensitive, clone. The effecton palisade spongy mesophyll thickness was clone-specific, whilethe amount of palisade intercellular space was reduced in allclones. A second effect was a change in the relative amountsof adaxial and abaxial epidermis. In palisade and spongy parenchymacells of all clones, ozone increased the number of irregularand spherical shaped chloroplasts, the electron density of chloroplaststroma, swelling and curling of thylakoids, translucency ofthe mitochondrial matrix and also the amount of cytoplasmiclipids. In the sensitive clone shorter chloroplasts and reducedamount of starch were observed in ozone-exposed plants, whilst,in the tolerant clone, the size of chloroplasts and the amountof starch were unaffected. Ozone effects on number, size andelectron density of plastoglobuli and vacuolar tannin were clone-dependent.At the ultrastructural level, the normal leaf ageing processprogressed at different rates in the birch clones. Ozone acceleratedsenescence-related structural changes, in accordance with earlierobservations of deciduous species.Copyright 1995, 1999 AcademicPress Betula pendula Roth., birch, clones, ageing, ozone, leaf anatomy, ultrastructure     

The ultrastructural organization of spruce needles of different ages exposed to radiation]

The ultrastructure of mesophyll cells of spruce needle of different age within the region of Chernobyl catastrophe has been investigated. The quantitative characteristics of chloroplasts are shown to be a function of the absorbed dose rate. The effect of ionizing radiation on the processes of needle tissue ageing is discussed.     

SECONDARY PULVINUS OF ROBINIA PSEUDOACACIA (LEGUMINOSAE): STRUCTURAL AND ULTRASTRUCTURAL FEATURES

The structure of the secondary pulvinus of Robinia pseudoacacia has been examined together with ultrastructural features of motor cells both in open and closed pulvini, to identify ultrastructural changes associated with leaflet movement. Pulvini have a central vascular core bordered by thick-walled collenchyma cells, which in turn are surrounded by several layers of cortical parenchyma cells. Cortical motor cells exhibit ultrastructural features similar to those reported in homologous cells of other pulvini. The vacuolar compartment contains two kinds of vacuoles: nontannin vacuoles, which change both in number and size during leaflet movement, and tannin vacuoles, which may act as an ion reservoir. No differences in wall thickness were found between flexor and extensor motor cells. Thick walls of collenchyma cells show numerous pits with plasmodesmata through which the phloem parenchyma cells and the inner cortical motor cells are connected. Tannin vacuoles and calcium oxalate crystals are common inclusions of phloem parenchyma cells. The tissue arrangement and the occurrence of pits with plasmodesmata in the central cylinder cells provide evidence of symplastic continuity through the central cylinder between the extensor and flexor regions of the motor organs. The greater amplitude of Robinia leaflet movements may be related to the extension of motor regions, the scarcity of lignification in the central vascular core, and the thin flexor walls.     

Leaf phloem alterations in two provenances ofAbies alba Miller (silver fir) during ageing

The phenomenon of premature needle senescence in two Italian provenances of silver fir was studied as part of a more general research project on decliningAbies alba. The two provenances showed a different degree of sensitivity to atmospheric pollutants. Comparative observations on 1-, 3-, 5-, 7- and 9-year-old needles from both provenances highlighted a faster loss of function in the phloem of needles from the Vallombrosa provenance. This fact appears to be related more to a loss of function in the cambium, than to a different rate of degeneration of sieve cells. This phenomenon may help explain why needles of the Vallombrosa provenance tend to be shed sooner than those of the Serra San Bruno provenance.     

Conifer decline in the north-east of France: Characteristic changes in chloroplast protein pattern and absence of anti-oxidative defense capability point to an involvement of ozone

A method yielding intact and biologically active chloroplasts from needles of mature spruce ( Picea abies L. cv. Karsten) or fir ( Abies alba Mill.) growing in the field is presented. Using this method, chloroplasts were prepared from yellow and green needles of declining spruce and fir from two forest areas of the Vosges mountains (north-east of France). Both quantitative and qualitative changes could be shown in the two-dimensional chloroplast protein pattern of yellowing needles, as compared to that of apparently healthy needles. We observed that an accumulation of photosystem II D1-protein in the thylakoids was associated with needle decline. Furthermore, a lack of resistance capability against free radical attack was observed in yellow needles. We have attempted to correlate our observations with pollutant levels and we propose that, in the forest areas studied, O₃ is one of the main factors involved in conifer decline. It also appears from our results that decline and senescence are distinct phenomena.     

SECONDARY GROWTH IN NEEDLE LEAVES OF PINUS LONGAEVA (BRISTLECONE PINE) AND OTHER CONIFERS: QUANTITATIVE DATA

Needle leaves of Pinus longaeva can be accurately dated and remain alive on branches for 30 or more yrs, making this species ideal to study secondary growth in leaves. Field observations and regression analysis of needle age versus mean needle length both indicate primary (elongation) growth of needles is completed in the first year. Statistical analysis of cell counts for one- to 33-yr-old needles indicate production along the entire length of needles of 1.0-1.7 cell layers per year of secondary phloem, but no secondary xylem. Microscopic measurements and cell counts reveal that with advancing needle age phloem increases radially and transfusion tissue buckles, but the number of endodermal cells and xylem width do not change. Living phloem remains constant in amount (ca. 9 layers) with advancing needle age, indicating yearly replacement of old by new phloem. For comparison to P. longaeva, needle leaves were also studied for ten other conifer taxa with maximum needle longevities ranging from 3 to 19 yrs. In needles of each taxon no secondary xylem is produced, but secondary phloem production occurs throughout the post-elongation lifespan of the needles regardless of maximum needle longevity.     

Vegetative anatomy of Pachycortnus (Anacardiaceae)

Pachycormus discolor , an arborescent desert perennial endemic to Baja California, has small, pinnately compound, hypostomatic, bifacial leaves produced on short shoots and photosynthetic stem phelloderm covered by exfoliating translucent phellem. Tightly packed laminal palisade cells are filled with tannins and lack chloroplasts. Spongy mesophyll is the major photosynthetic tissue. Leaves possess unicellular trichomes with secondary walls and uniseriate trichomes with glandular heads. Schizogenous resin ducts occur in primary phloem of stems, leaves and roots as well as all living tissues of the bark. Developmental studies reveal that initiation and differentiation of foliar primordia resembles that of other dicotyledons except that tannin cells and secretory ducts arise precociously. Primary vasculature is an open sympodial system with three principal traces diverging toward each foliar primordium. The wood is highly specialized and comprises mostly unlignified cells packed with starch grains. Thick bark is mainly produced as annual layers of secondary phloem marked by a ring of secretory ducts each surrounded by tannin cells. The possible adaptive significance of these unusual anatomical features is discussed.     

VARIATIONS IN ANATOMY,ASSOCIATIONS, AND ORIGINS OF KRANZ TISSUE

Although the unique tissue required for C₄ photosynthesis in nonsucculent plants is often described as being modified leaf parenchyma sheath, which is positioned meaningfully between the mesophyll externally and the vascular tissues internally, the actual range of locations and known associations make that concept untenable. In origin the Kranz tissue develops from procambium as well as ground parenchyma. It is found in stems as well as leaves. In position Kranz tissue can lie in the parenchyma sheath, in the mestome sheath, isolated in the mesophyll, peripherally in some thick leaves, or within the veins. It can be associated with mesophyll only, mesophyll and colorless parenchyma, mesophyll and sclerenchyma, other Kranz tissue and vascular tissues, mesophyll and mestome sheath, mesophyll and phloem, mesophyll and xylem, epidermis, and, finally, mestome sheath and xylem and phloem. The use of the term Kranz is expounded.     

THE ANATOMIC RESPONSES OF DAUCUS CAROTA TO THE ASTER YELLOWS VIRUS

Struckmeyer, B. Esther. (U. Wisconsin, Madison.) The anatomic responses of Daucus carota to the aster yellows virus. Amer. Jour. Bot. 50(9): 959–963. Ilus. 1963.—The leaves, petioles, and roots of carrots (Daucus carota) displaying aster yellows virus in the field and those infected with the aster yellows virus inoculated by the 6-spotted leafhopper were examined anatomically. Compared to the uninoculated, the young infected leaves displayed fewer layers of palisade cells and larger spongy parenchyma cells with a more compact arrangement. Mature leaves of infected plants sometimes were undulated and had few chloroplasts, many of which appeared fragmented. Hypertrophy and hyperplasia in the phloem tissue were associated with some necrosis and obliteration of cells. Long, needle-shaped crystalline inclusion bodies were present in the phloem in the leaves and roots. Most of the vascular bundles of the petiole were abnormal. Malformations included proliferating phloem cells, which in some instances almost encircled the bundle, hyperplasia of the phloem, hypertrophy of the parenchyma, and considerable necrosis and obliteration of these cells. Other responses included the division into 3 or 4 rows of the large outer phloem parenchyma by parallel walls so that a cambium-like layer was simulated. The tissue enclosed by this layer divided and underwent considerable necrosis and gummosis. Lacunae were found between the phloem bundle cap and the older phloem. Some of the cells in the phloem tissue differentiated into bundles with xylem. Numerous, short, lateral roots were conspicuous on the main root. The most noticeable response of the root tissue to this malady was hyperplasia and hypertrophy of the phloem followed by cellular disorganization, necrosis, and obliteration of cells. Some sieve tubes, companion cells, and parenchyma contained a gumlike deposit.