Ethylene and not embolism is required for wound-induced tylose development in stems of grapevines

The pruning of actively growing grapevines (Vitis vinifera) resulted in xylem vessel embolisms and a stimulation of tylose formation in the vessels below the pruning wound. Pruning was also followed by a 10-fold increase in the concentration of ethylene at the cut surface. When the pruning cut was made under water and maintained in water, embolisms were prevented, but there was no reduction in the formation of tyloses or the accumulation of ethylene. Treatment of the stems with inhibitors of ethylene biosynthesis (aminoethoxyvinylglycine) and/or action (silver thiosulfate) delayed and greatly reduced the formation of tyloses in xylem tissue and the size and number of those that formed in individual vessels. Our data are consistent with the hypotheses that wound ethylene production is the cause of tylose formation and that embolisms in vessels are not directly required for wound-induced tylosis in pruned grapevines. The possible role of ethylene in the formation of tyloses in response to other stresses and during development, maturation, and senescence is discussed.     

Applications of the compensating pressure theory of water transport

Some predictions of the recently proposed theory of long-distance water transport in plants (the Compensating Pressure Theory) have been verified experimentally in sunflower leaves. The xylem sap cavitates early in the day under quite small water stress, and the compensating pressure P (applied as the tissue pressure of turgid cells) pushes water into embolized vessels, refilling them during active transpiration. The water potential, as measured by the pressure chamber or psychrometer, is not a measure of the pressure in the xylem, but (as predicted by the theory) a measure of the compensating pressure P. As transpiration increases, P is increased to provide more rapid embolism repair. In many leaf petioles this increase in P is achieved by the hydrolysis of starch in the starch sheath to soluble sugars. At night P falls as starch is reformed. A hypothesis is proposed to explain these observations by pressure-driven reverse osmosis of water from the ground parenchyma of the petiole. Similar processes occur in roots and are manifested as root pressure. The theory requires a pump to transfer water from the soil into the root xylem. A mechanism is proposed by which this pump may function, in which the endodermis acts as a one-way valve and a pressure-confining barrier. Rays and xylem parenchyma of wood act like the xylem parenchyma of petioles and roots to repair embolisms in trees. The postulated root pump permits a re-appraisal of the work done by evaporation during transpiration, leading to the proposal that in tall trees there is no hydrostatic gradient to be overcome in lifting water. Some published observations are re-interpreted in terms of the theory: doubt is cast on the validity of measurements of hydraulic conductance of wood; vulnerability curves are found not to measure the cavitation threshold of water in the xylem, but the osmotic pressure of the xylem parenchyma; if measures of xylem pressure and of hydraulic conductance are both suspect, the accepted view of the hydraulic architecture of trees needs drastic revision; observations that xylem feeding insects feed faster as the water potential becomes more negative are in accord with the theory; tyloses, which have been shown to form in vessels especially vulnerable to cavitation, are seen as necessary for the maintenance of P, and to conserve the supplementary refilling water. Far from being a metastable system on the edge of disaster, the water transport system of the xylem is ultrastable: robust and self-sustaining in response to many kinds of stress.     

Development of tyloses in the xylem vessels of Meili grapevine and their effect on water transportation

Shoot pruning could cause short-term damages to vines. In response to damage, tyloses develop in shoot xylem vessels interfering free water and mineral transportation. In this study, the tylosis development at different nodes of the current-year and perennial shoots of sixty three-year-old grapevines (Vitis vinifera L.) after pruning was investigated. The results showed that tyloses at the nodes closest to the trunk developed rapidly; and tylosis development initiated at the time when the size of vessel-ray pit was greater than that of parenchyma cell-parenchyma cell pit. In current-year shoots, tyloses were formed in up to 87% of the vessels, and 40% of the vessels were completely blocked by tyloses. In wound-induced perennial shoots, 30% of the vessels were completely blocked by tyloses. When few vessels were blocked by tyloses, new vessels could differentiate, and water transportation system may be restored. However, when tyloses developed in a large number of vessels and the large number of the vessels were blocked, the original capability of water transport was decreased (the largest decrease was 21.1% in this study), resulting in dehiscence or shrinkage cracking in this area. The study proved that the tylosis formation in functional vessels limited the water transport efficiency.     

Tyloses and ecophysiology of the early carboniferous progymnosperm tree Protopitys buchiana

Trunk woods of Early Carboniferous Protopitys buchiana show the earliest example of tylose formation and the first record for a progymnosperm. Protopitys tyloses are more densely located in inner trunk woods and near growth layer boundaries. We suggest, therefore, that an altered physiological state of living ray cells, during dormancy and/or following water stress, was necessary to make the woods vulnerable to tylose formation. Coupled with the distribution and proximity of abundant wood ray parenchyma to large xylem conducting cells, the positions of conduits filled with tyloses can be interpreted as ecophysiological responses of the plant to changes in local environment. In addition, some xylem conducting cells might have functioned as vessels. Fungal hyphae are present in some tracheary cells and in some areas with tyloses, but there is no evidence for wood trauma; we conclude, therefore, that these particular cases of tyloses are probably not induced by wound trauma. Protopitys buchiana wood thus shows structure/function similarities to modern woods with vessels, such as those of dicot angiosperms. This implies that ancient and modern plant ecophysiological responses correlate well with the physical parameters of their cellular construction.     

Defense response of resistant host Impatiens balsamina to the parasitic angiosperm Cuscuta japonica

The response of the stem of a resistant host (Impatiens baslamina) to infection by the parasitic flowering plant Cuscuta japonica was studied with light and electron microscopy. The intra- and interfascicular cambial cells in the host stem first reacted to the penetrating upper haustorium by dividing, and the differentiation of the host xylem (vascular) tissues proceeded toward interfascicular areas from vascular bundles. When the host vascular tissue was invaded by the endophyte (haustorial portion in the host stem), the host xylem was displaced, and host vessels became occluded with parenchyma cells, resulting in tyloses. As the parasitism progressed, areas of the host stem penetrated by the endophyte became swollen via secondary growth and cell division in the parenchymatous cortex, pith, and interfascicular areas. During this intrusion by the endophyte, darkly stained necrotic reactions were detected at the interface between the host tissue and the invading endophyte. The results suggested that in the host tissues penetrated by the parasite, the formation of secondary tissue and swellings caused by active cell division of ground tissue and host vessel occlusion by tyloses constitute the host structural defense against the parasite.     

Pruning-induced tylose development in stems of current-year shoots of Vitis vinifera (Vitaceae)

Tyloses form in xylem vessels in response to various environmental stimuli, but little is known of the kinetics or regulation of their development. Preliminary investigations indicated that wounds seal quickly with tyloses after pruning of grapevine shoots. In this study, tylose development was analyzed qualitatively and quantitatively at different depths and times from pruning cuts along current-year shoots of grapevines at basal, middle, and apical stem regions. Tyloses developed simultaneously within a single vessel but much separated in time among vessels. Pruning caused prodigious tylosis in vessels of grape stems, extending to approximately 1 cm deep and to 7 d after wounding, but about half of the vessels did not become completely occluded. The fraction of vessels forming tyloses was greatest in basal (85%) and least in apical (50%) regions. The depth of maximum density of tyloses was 4 mm from the cut in the basal region and 2 mm from the cut in the middle and apical regions. Tylose development was faster in the basal and middle than in the apical region. The pattern of tylose development is discussed in the context of wound repair and pathogen movement in grapevines.     

Suberized tyloses in trees: An ultrastructural and cytochemical study

The nature of the wall layers observed in suberized tyloses was studied in Populus basalmifera L., Ulmus americana L. and Quercus rubra L. As the suberin layers were present only in tyloses that had completed their expansion, most of the results concern mature tyloses. The cyto- and immunocytochemical tests were conducted, respectively, with an exoglucanase having a binding affinity for β(1→4)-D-glucans, the subunits of cellulose, and with two monoclonal antibodies specific for un-esterified and esterified pectic molecules. In the three species, labelling for pectic compounds was intense over the external layer of tyloses but usually more dispersed or nearly absent over the layer corresponding to a primary wall that was, however, intensely labelled for β(1→4)-D-glucans. The outer wall layer, comparable to a middle lamella in mature tyloses, was continuous with similar material that appeared to be secreted by the tylosis. This material was particularly abundant in pit chambers, in void spaces between the tylosis and the vessel wall, particularly at the junction of the vessel and two adjacent cells, and close to the rim of vessel perforation plates. In P. balsamifera, a single suberized layer or occasionally a succession of suberized and cellulose-containing layers was observed internal to the tylosis primary wall. In U. americana, the wall of tylosis was similar to that of P. balsamifera except that, at times, a secondary-wall-like layer was formed and only a single suberized layer was observed. In Q. rubra, the suberized layer was always observed internal to the tylosis secondary wall. Simple pits were also constantly noted in Q. rubra tyloses. The occasional occurrence of a cellulosic layer internally to the suberized layer was observed in the three species. Histochemical tests revealed that lignin was also an important component of the tylosis wall. The tyloses frequently contained phenolic compounds in close association with the suberized layers. The significance of the formation of suberized tyloses in trees is discussed.     

树木树液上升机理研究进展

水分在植物体内的运输一直是很多植物生理生态学家所关注的一个重要问题。介绍了内聚力学说的基本假设和其存在争议,总结了近年来这一研究领域的几个热点问题,主要包括：（1）木质部栓塞及其恢复机理;（2）木质部压力探针和压力室法测定的木质部张力值不一致的现象及其可能原因;（3）补偿压学说;（4）不同界面层张力以及输水管道的毛细作用力、薄壁细胞膨压和木质部渗透压、逆向蒸腾等在树木汁液上升中的贡献;（5）最近发现的存在于木质部导管伴胞和韧皮部薄壁细胞等质膜中的水孔蛋白在植物水分运输中的调控作用等。这些方面在解释树木的树液上升中都起着重要的作用。     

Induction of air embolism in xylem conduits of pre-defined diameter.

A new method is presented that enables the induction of embolisms in a fraction of all xylem vessels, based on diameter, at one cut end of a stem segment. The method is based on the different capillary characteristic of xylem vessels of different cross-sectional size. To verify the method, air embolisms were induced in cut xylem vessels of chrysanthemum (Dendranthemaxgrandiflorum Tzvelev cv. Cassa) stem segments at different xylem tensions and compared with the distribution of gas-filled vessels as visualized by cryo-scanning electron microscopy (Cryo-SEM). At -6 kPa xylem pressure, air-entrance was only induced in large diameter vessels (>30 microm), while at -24 kPa embolisms were induced in almost all xylem vessels (>10 microm). Although the principle of the embolization method worked well, smaller diameter vessels were observed to be embolized than was expected according to the calculations. The role of cross-sectional shape and contact angle between xylem sap and vessel wall at the menisci are discussed. After correction for the observed (diameter independent) deviation from circularity of the cross-sectional vessel shape the contact angle was calculated to be approximately 55 degrees. Hydraulic resistance (Rh) measurements before and after embolization showed that the effect of embolizing only large diameter cut xylem vessels had only a small influence on overall Rh of a stem segment. Embolizing all cut xylem vessels at one cut end almost trebled overall Rh. The difference was discussed in the light of the networking capacity of the xylem system.     

Cu2+ inhibition of gel secretion in the xylem and its potential implications for water uptake of cut Acacia holosericea stems

Maintaining a high rate of water uptake is crucial for maximum longevity of cut stems. Physiological gel/tylosis formation decreases water transport efficiency in the xylem. The primary mechanism of action for post‐harvest Cu²⁺ treatments in improving cut flower and foliage longevity has been elusive. The effect of Cu²⁺ on wound‐induced xylem vessel occlusion was investigated for Acacia holosericea A. Cunn. ex G. Don. Experiments were conducted using a Cu²⁺ pulse (5 h, 2.2 mM) and a Cu²⁺ vase solution (0.5 mM) vs a deionized water (DIW) control. Development of xylem blockage in the stem‐end region 10 mm proximal to the wounded stem surface was examined over 21 days by light and transmission electron microscopy. Xylem vessels of stems stood into DIW were occluded with gels secreted into vessel lumens via pits from surrounding axial parenchyma cells. Gel secretion was initiated within 1–2 days post‐wounding and gels were detected in the xylem from day 3. In contrast, Cu²⁺ treatments disrupted the surrounding parenchyma cells, thereby inhibiting gel secretion and maintaining the vessel lumens devoid of occlusions. The Cu²⁺ treatments significantly improved water uptake by the cut stems as compared to the control.     

Water relations in silver birch during springtime: How is sap pressurised?

• Positive sap pressures are produced in the xylem of birch trees in boreal conditions during the time between the thawing of the soil and bud break. During this period, xylem embolisms accumulated during wintertime are refilled with water. The mechanism for xylem sap pressurization and its environmental drivers are not well known.
• We measured xylem sap flow, xylem sap pressure, xylem sap osmotic concentration, xylem and whole stem diameter changes, and stem and root non‐structural carbohydrate concentrations, along with meteorological conditions at two sites in Finland during and after the sap pressurisation period.
• The diurnal dynamics of xylem sap pressure and sap flow during the sap pressurisation period varied, but were more often opposite to the diurnal pattern after bud burst, i.e. sap pressure increased and sap flow rate mostly decreased when temperature increased. Net conversion of soluble sugars to starch in the stem and roots occurred during the sap pressurisation period. Xylem sap osmotic pressure was small in comparison to total sap pressure, and it did not follow changes in environmental conditions or tree water relations.
• Based on these findings, we suggest that xylem sap pressurisation and embolism refilling occur gradually over a few weeks through water transfer from parenchyma cells to xylem vessels during daytime, and then the parenchyma are refilled mostly during nighttime by water uptake from soil. Possible drivers for water transfer from parenchyma cells to vessels are discussed. Also the functioning of thermal dissipation probes in conditions of changing stem water content is discussed.

     

Changes in leaf organisation, photosynthetic performance and wood formation during ex vitro acclimatisation of black mulberry (Morus nigra L.)

Changes in anatomical organisation of the leaf, photosynthetic performance and wood formation were examined to evaluate the temporal and spatial patterns of acclimatisation of micropropagated slow-growing black mulberry ( Morus nigra L.) plantlets to the ex vitro environment. Leaf structure differentiation, the rates of net photosynthesis (P_n), transpiration (E) and stomatal conductance (g_s), and secondary xylem growth were determined in the course of a 56-day acclimatisation. Differentiation of palisade parenchyma was observed 7 days after transfer. At this stage, the rates of P_n, E and g_s reached maximum values, after which the rates of all three gas exchange parameters gradually decreased. The highest proportion of woody area occupied by vessels was also observed 7 days after transfer. An important feature of developing woody tissue is the difference in patterns of vessel distribution from the characteristic differentiation patterns of earlywood and latewood vessels in mature wood of ring-porous trees. Vessels with lumen areas over 3000 μm² were only differentiated in acclimatised plantlets, whereas vessels in stems sampled on days 0 and 7 had very small lumen areas of up to 560 μm². Full acclimatisation, observed 56 days after transfer to the ex vitro environment, was associated with the rapid growth of new in vivo formed leaves, very low rates of E and g_s, and much increased secondary xylem tissue within the stem area.     

黄芩根中木间木栓的形成与发育特性研究

应用光学显微镜和电子显微镜对3年生黄芩(Scutellaria baicalensis Georgi)根中木间木栓的形成及其发育过程进行了细致观察,以揭示黄芩木间木栓形成的特征以及发育过程中导管及周围薄壁细胞的变化,旨在为中药枯芩的形成提供理论依据和实验证据。结果表明：(1)黄芩根的次生结构由周皮、皮层和次生维管组织组成;木间木栓发生在木质部异常导管的周围,先是导管内出现侵填体或其他物质,接着这些导管邻近的部分薄壁细胞内也出现一些物质,随后导管外围的正常薄壁细胞恢复分生能力,产生木栓形成层,进而形成木间木栓。(2)木间木栓将健康组织与这些异常的导管和薄壁细胞隔离,隔离在内的组织会逐渐成为死亡组织。(3)横切面观,小木间木栓可同时或先后发生在木质部的不同位置,由内向外逐渐扩大,与周围的木间木栓相连,最终形成大的木间木栓。(4)纵切面观,木间木栓以管状形式由根头部向下延伸,部分根中距根头部以下3 cm内有木间木栓存在。     

Possible Mechanisms Limiting Movement of Ralstonia solanacearum in Resistant Tomato Tissues

The distribution and appearance of Ralstonia solanacearum in the upper hypocotyl tissues of root‐inoculated tomato seedlings of resistant rootstock cultivar LS‐89 (a selection from Hawaii 7998) and susceptible cultivar Ponderosa were compared to clarify the mechanism that limits the movement of the bacterial pathogen in resistant tomato tissues. In stems of wilted Ponderosa plants, bacteria colonized both the primary and the secondary xylem tissues. Bacteria were abundant in vessels, of which the pit membranes were often degenerated. All parenchyma cells adjacent to vessels with bacteria were necrotic and some of them were colonized with bacteria. In stems of LS‐89 plants showing no discernible wilting symptoms, bacteria were observed in the primary xylem tissues but not in the secondary xylem tissues. Necrosis of parenchyma cells adjacent to vessels with bacteria was observed occasionally. The pit membranes were often thicker with high electron density. The inner electron‐dense layer of cell wall of parenchyma cells and vessels was thicker and more conspicuous in xylem tissues of infected LS‐89 than in xylem of infected Ponderosa or mock‐inoculated plants. Electron‐dense materials accumulated in or around pit cavities in parenchyma cells next to vessels with bacteria, and in vessels with bacteria. Many bacterial cells appeared normal in vessels, except for those in contact with the pit membranes. These results indicate that R. solanacearum moves from vessel to vessel in infected tissues through degenerated pit membranes and that restricted movement in xylem tissues was the characteristic feature in LS‐89. The limitation in bacterial movement may be related to the thickening of the pit membranes and/or the accumulations of electron‐dense materials in vessels and parenchyma cells.     

Grapevine xylem response to fungi involved in trunk diseases

Grapevine trunk diseases (GTD), caused by a wide range of different fungi, are responsible for decline and productivity losses in vines at all growth stages. Grapevine responses to fungal attack include morphological and physiochemical defence mechanisms in the vascular system to reduce fungal infections. However, the extent to which these responses could control further spread by GTD‐fungi in the xylem vessels is poorly known. This study shows the formation of tyloses inside xylem vessels of diseased grapevines, as well as extracellular ligninolytic activities [lignin peroxidase, manganese peroxidase (MnP) and/or laccase] exhibited by some GTD‐fungi isolated here from symptomatic grapevines. In particular, Botryosphaeriaceae spp. and Phaeoacremonium minimum showed all three lignin‐degrading enzymatic activities. We also examined whether selected vine phenolic compounds, often located in the vascular system in response to fungal infection, could affect the lignin‐degrading activity from those GTD‐fungi as well as fungal colonisation. We found that phenolic compounds appeared to inhibit MnP activity, in addition to reducing fungal growth by causing anomalies in the hyphae morphology. Our results support that affected grapevines can initiate the tylosis formation in order to constrain fungi in the xylem vessels, while highlight the complementary action of the phenolic compounds to inhibit the fungi growth and colonisation. Phenolic compounds are therefore likely to have important role in alternative strategies for preventing trunk diseases.     

拐枣果序梗的发育解剖学研究

拐枣肉质膨大果序梗的发育过程可划分为前、中、后、末４个时期,前期为初生生长时期,内部结构类似一般双子叶植物茎的初生构造;中期为维管形成层活动时期,产生了不同其茎的次生木质部,由成片木质化的厚壁纤维细胞、一定量的木射线及星散在其中极少数的导管组成;后期为异常分生组织活动时期,初生木质部木薄壁组织及邻近少量髓细胞及邻近少量髓细胞转化为异常分生组织,向外产生切向排列的薄壁细胞,经扩大的切向伸长,使原导管     

Vascular defense responses in rice: peroxidase accumulation in xylem parenchyma cells and xylem wall thickening.

The rice bacterial blight pathogen Xanthomonas oryzae pv. oryzae is a vascular pathogen that elicits a defensive response through interaction with metabolically active rice cells. In leaves of 12-day-old rice seedlings, the exposed pit membrane separating the xylem lumen from the associated parenchyma cells allows contact with bacterial cells. During resistant responses, the xylem secondary walls thicken within 48 h and the pit diameter decreases, effectively reducing the area of pit membrane exposed for access by bacteria. In susceptible interactions and mock-inoculated controls, the xylem walls do not thicken within 48 h. Xylem secondary wall thickening is developmental and, in untreated 65-day-old rice plants, the size of the pit also is reduced. Activity and accumulation of a secreted cationic peroxidase, PO-C1, were previously shown to increase in xylem vessel walls and lumen. Peptide-specific antibodies and immunogold-labeling were used to demonstrate that PO-C1 is produced in the xylem parenchyma and secreted to the xylem lumen and walls. The timing of the accumulation is consistent with vessel secondary wall thickening. The PO-C1 gene is distinct but shares a high level of similarity with previously cloned pathogen-induced peroxidases in rice. PO-C1 gene expression was induced as early as 12 h during resistant interactions and peaked between 18 and 24 h after inoculation. Expression during susceptible interactions was lower than that observed in resistant interactions and was undetectable after infiltration with water, after mechanical wounding, or in mature leaves. These data are consistent with a role for vessel secondary wall thickening and peroxidase PO-C1 accumulation in the defense response in rice to X. oryzae pv. oryzae.     

DoesAcetobacter diazotrophicusLive and Move in the Xylem of Sugarcane Stems? Anatomical and Physiological Data

We have previously shown that the nitrogen-fixing endophyteof sugarcane,Acetobacter diazotrophicuslives in the sugar solutionin the intercellular-space apoplast of the stem cortex. Variousauthors have claimed that it inhabits the xylem apoplast. Thispossibility has been investigated in the clone Ja 60-5 and shownto be most unlikely for the following reasons: (1) an adequatecarbon source is lacking in the xylem sap, and cannot be suppliedfrom the intercellular-space apoplast of the cortex. Diffusionof solutes into and out of the vascular bundles is preventedby complete lignification and suberization of the bundle sheathcell walls except at the nodes. (2) Longitudinal movement ofparticles as large as bacteria is severely limited at the nodes.Vessel end walls were found in 90% of vessels at each node,and only 1% of open vessels extended through two nodes. Noneextended as far as three nodes. In addition to vessel end walls,vessel continuity at nodes was interrupted by living cells.Dye solution in the transpiration stream in metaxylem vesselsdid not pass through these living cells, but accumulated incrystals (sump formation) in the vessels below the node. Onlyin some protoxylem vessels and cavities did dye solution movethrough many nodes. It is likely that selection of sugarcaneclones such as Ja 60-5 for resistance to bacterial wilt diseaseshave selected for clones that have limited vessel continuity.(3) When culturedA. diazotrophicuswas introduced into the transpirationstream, the xylem parenchyma reacted by secreting a bright redpolymer which killed the bacteria and blocked the movement ofwater. We conclude that the xylem flow-apoplast of this cloneof sugarcane is an unsuitable habitat forA. diazotrophicusandthat additional habitats to those of the intercellular-spaceapoplast should be sought elsewhere. Acetobacter diazotrophicus; endophytic bacteria; nitrogen fixation; sugarcane; vessel end walls; xylem apoplast; xylem bacteria; xylem segmentation