A method for measuring hydraulic conductivity and embolism in xylem

Abstract Hydraulic conductivity of the xylem is computed as the quotient of mass flow rate and pressure gradient. Measurements on excised plant stems can be difficult to interpret because of time-dependent reductions in flow rate, and because of variable degrees of embolism. Using Acer saccharum Marsh. stems, we found that certain perfusing solutions including dilute fixatives (e.g. 0.05% formaldehyde) and acids with pH below 3 (e.g. 10 mol m^?3 oxalic) prevent long-term decline in conductivity. Xylem embolism can be quantified by expressing the initial conductivity as a percentage of the maximum obtained after flow-impeding air emboli have been removed by repeated high-pressure (175 kPa) flushes. Correlation between microbial contamination and declining conductivity suggests that long-term (> 4h) declines are caused by microbial growth within the vessels. Unpredictable trends in short-term (< 4h) measurements may be caused by movements of air emboli in vessels and/or participate matter.     

8种木本植物木质部栓塞变化与生理生态指标关系的研究Ⅰ.与植物木质部水势的关系

以自然状况下生长良好的耐旱树种刺槐(Robinia pseudoacacia L.)、元宝枫(Acer truncatum Bge.)、沙棘(Hippophae rhamnoides L.)、白榆(Ulmus pumila L.)、油松(Pinus tabulaeformis Carr.)、白皮松(Pinus bungeana Zucc.ex Endl.)及中生树种女贞(Ligustrum lucidum Ait.)、柳树(Salix matsudana Koidz. f. pendula Schneid.)为研究对象,用压力室法测定木质部水势,用冲洗法测定木质部栓塞程度,研究不同生长季节木质部栓塞与水势间的关系.结果表明针叶树油松、白皮松在各季节水势均较高,水势变化幅度较小,木质部不易发生栓塞,这与其木质部由管胞构成,对木质部栓塞不敏感,在干旱时采用高水势延迟脱水的耐旱策略有关.阔叶树刺槐、元宝枫、沙棘、白榆、女贞和柳树的木质部栓塞现象是其在每天正常生长过程中不可避免的 "平常事件",是它们适应干旱的一种方式.它们的木质部栓塞程度与水势表现出了相反的变化趋势,即同一树种在同一季节内水势值越低,木质部栓塞程度越大,但在不同树种及同一树种的不同季节不存在这种关系.由此可见,植物木质部栓塞对水势的敏感程度(即木质部栓塞脆弱性)主要由树种的木质部结构决定,同时受到树种特性、树木生长发育时期、外界环境因子的影响,木质部栓塞的脆弱性也具有季节变化特征.     

The spatial pattern of air seeding thresholds in mature sugar maple trees

Air seeding threshold (P_a) of xylem vessels from current year growth rings were measured along the vertical axis of mature sugar maple trees (Acer saccharum Marsh.), with sampling points in primary leaf veins, petioles, 1-, 3-, and 7-year-old branches, large branches, the trunk and roots. The air seeding threshold was taken as the pressure required to force nitrogen gas through intervessel pit membranes. Although all measurements were made on wood produced in the same year, P_a varied between different regions of A. saccharum, with distal organs such as leaves and petioles having lower P_a than basal regions. Mean (SE) P_a ranged from 1.0 (± 0.1) MPa in primary leaf veins to 4.8 (± 0.1) MPa in the main trunk. Roots exhibited a P_a of 2.8 (± 0.2) MPa, lower than all other regions of the tree except leaf veins and petioles. Mean xylem vessel diameter increased basipetally, with the widest vessels occurring in the trunk and roots. Within the shoot, wider vessels had greater air seeding thresholds, contrasting with trends previously reported. However, further experimentation revealed that differences in P_a between regions of the stem were driven by the presence of primary xylem conduits, rather than differences in vessel diameter. In 1-year-old branches, P_a was significantly lower in primary xylem vessels than in adjacent secondary xylem vessels. This explained the lower values of P_a measured in petioles and leaf veins, which possessed a greater ratio of primary xylem to secondary xylem than other regions. The difference in P_a between primary and secondary xylem was attributed to the greater area of primary cell wall (pit membrane) exposed in primary xylem conduits with helical or annular thickening.     

8种木本植物木质部栓塞变化与生理生态指标关系的研究 I.与植物木质部水势的关系

以自然状况下生长良好的耐旱树种刺槐（Robinia pseudoacacia L.）、元宝枫（Acer truncatum Bge）、沙棘（Hippophae rhamnoides L.）、白榆（Ulmus pumila L.）、油松（Pinus tabulaeformis Carr.）、白皮松（Pinus bungeana Zucc.ex Endl.）及中生树种女贞（Ligustrum lucidum Ait.）、柳树（Salix matsudana Koidz.f.pendula Schneid.）为研究对象,用压力室法测定木质部水势,用冲洗法测定木质部栓塞程度,研究不同生长季节木质部栓塞与水势间的火系。结果表明：针叶树油松、白皮松在各个季节水势均较高,水势变化幅度较小,木质部不易发生栓塞,这与其木质部由管胞构成,对木质部栓塞不敏感,在干旱时采用高水势延迟脱水的耐旱策略有关。阔叶树刺槐、元宝枫、沙棘、白榆、女贞和柳树的木质部栓塞现象是其在每天正常生长过程中不可避免的“平常事件”,是它们适应干旱的一种方式。它们的木质部栓塞程度与水势表现出了相反的变化趋势,即同一树种在同一季节内水势值越低,木质部栓塞程度越大,但在不同树种及同一树种的不同季节不存在这种关系。由此可见,植物木质部栓塞对水势的敏感程度（即木质部栓塞脆弱性）主要由树种的木质部结构决定,同时受到树种特性、树木生长发育时期、外界环境因子的影响,木质部栓塞的脆弱性也具有季节变化特征。     

植物体内水分长距离运输的生理生态学机制

植物体内长距离水分运输是植物生理生态学研究中的一个重要问题,长期为植物生理学家和生理生态学家所关注。木质部探针技术的问世,掀起了近年来植物生理学界最为激烈的一场争论。提出了已经有100多年,风行40年的内聚力-张力(Cohesion-Tension, C-T)学说受到质疑。随后维护派和质疑派围绕木质部探针技术、压力室技术(C-T理论的主要支撑实验技术)的可靠性展开辩论。进一步从物理学原理和各种实验上就C-T理论的3个支柱(木质部导管或管胞中巨大的张力、沿树高的压力梯度、连续水柱)进行争论。这场争论似暂告一段落,C-T理论没有被推翻,但仍留有问题期待以后的研究。     

Translocation of nitrogen in a vegetative wheat plant (Triticum aestivum)

The translocation of nitrogen was studied in vegetative wheat plants ( Triticum aestivum L. cv. SUN 9E) grown with a limited supply of nitrogen. The concentration of nitrogen in xylem sap exuding from the excised roots was the same as the nitrogen concentration in the transpiration stream. Translocation of nitrogen to the shoot was, therefore, calculated as the product of the transpiration rate and the concentration of nitrogen in xylem exudates. On the 22nd day from sowing more nitrogen was translo-cated to the shoot than it incorporated, and 56% of the nitrogen translocated to the shoot was retranslocated to the roots. The nitrogen retranslocated to the roots was more than adequate to supply the requirements of the roots for growth, and the balance of the retranslocated nitrogen was reloaded into the xylem stream. Expressed as a proportion of the total increment of nitrogen in the plant on day 22, between 79 and 100% of the nitrogen absorbed by the plant was "cycled' in the plant (root → shoot → root → shoot). It is suggested that the size of this mobile reserve of nitrogen may vary depending on the growth requirement of the plant, its nitrogen-uptake capacity and the contribution of nitrogen from mobilisation of leaf protein during senescence.     

New evidence for large negative xylem pressures and their measurement by the pressure chamber method

Pressure probe measurements have been interpreted as showing that xylem pressures below c. –0.4 MPa do not exist and that pressure chamber measurements of lower negative pressures are invalid. We present new evidence supporting the pressure chamber technique and the existence of xylem pressures well below –0.4 MPa. We deduced xylem pressures in water-stressed stem xylem from the following experiment: (1) loss of hydraulic conductivity in hydrated stem xylem (xylem pressure = atmospheric pressure) was induced by forcing compressed air into intact xylem conduits; (2) loss of hydraulic conductivity from cavitation and embolism in dehydrating stems was measured, and (3) the xylem pressure in dehydrated stems was deduced as being equal and opposite to the air pressure causing the same loss of hydraulic conductivity in hydrated stems. Pressures determined in this way are only valid if cavitation was caused by air entering the xylem conduits (air-seeding). Deduced xylem pressure showed a one-to-one correspondence with pressure chamber measurements for 12 species (woody angiosperms and gymnosperms); data extended to c. –10 MPa. The same correspondence was obtained under field conditions in Betula occidentalis Hook., where pressure differences between air- and water-filled conduits were induced by a combination of in situ xylem water pressure and applied positive air pressure. It is difficult to explain these results if xylem pressures were above –0.4 MPa, if the pressure chamber was inaccurate, and if cavitation occurred by some mechanism other than air-seeding. A probable reason why the pressure probe does not register large negative pressures is that, just as cavitation within the probe limits its calibration to pressures above c. –0.5 MPa, cavitation limits its measurement range in situ.     

The importance of xylem constraints in the distribution of conifer species

Vulnerability of stem xylem to cavitation was measured in 10 species of conifers using high pressure air to induce xylem embolism. Mean values of air pressure required to induce a 50% loss in hydraulic conductivity (φ₅₀) varied enormously between species, ranging from a maximum of 14.2±0.6 MPa (corresponding to a xylem water potential of −14.2 MPa) in the semi-arid species Actinostrobus acuminatus to a minimum of 2.3±0.2 MPa in the rainforest species Dacrycarpus dacrydioides . Mean φ₅₀ was significantly correlated with the mean rainfall of the driest quarter within the distribution of each species. The value of φ₅₀ was also compared with leaf drought tolerance data for these species in order to determine whether xylem dysfunction during drought dictated drought response at the leaf level. Previous data describing the maximum depletion of internal CO₂ concentration (c_i) in the leaves of these species during artificial drought was strongly correlated with φ₅₀ suggesting a primary role of xylem in effecting leaf drought response. The possibility of a trade-off between xylem conductivity and xylem vulnerability was tested in a sub-sample of four species, but no evidence of an inverse relationship between φ₅₀ and either stem-area specific (K_a) or leaf-area specific conductivity (K₁) was found.     

Limits to xylem refilling under negative pressure in Laurus nobilis and Acer negundo

The ability of juvenile Laurus nobilis and Acer negundo plants to refill embolized xylem vessels was tested under conditions of soil drought when xylem sap pressure was substantially negative, thus violating the expected condition that pressure must rise to near atmospheric for refilling. Intact potted plants were dried to a stem water potential (Σ_W) corresponding with approximately 80% loss of hydraulic conductivity (PLC) in shoots. Then plants were re‐watered and kept at a less negative target Ψ_W for 1–48 h. The Ψ_W was measured continuously with stem psychrometers. Rewatered L. nobilis held at the target Ψ_W for 1 h showed no evidence for refilling unless Ψ_W was within a few tenths of a MPa of zero. In contrast, re‐watered L. nobilis held for 24 and 48 h at water potentials well below zero showed a significant reduction in PLC. The recovery was highly variable, being complete in some stem segments, and scarcely evident in others. Embolism repair was accompanied by a significant but moderate decrease in the osmotic potential (Ψ) of the bulk xylem sap (Ψ = ?67 kPa in recovering plants versus ?31 kPa in controls). In contrast, embolized and re‐watered A. negundo plants held for 24 h at target Ψ_W of ?0·9 and ?0·3 MPa showed no embolism reversal. The mechanism allowing L. nobilis plants to refill under negative pressure is unknown, but does not appear to operate in A. negundo, and is slower to act for drought‐induced embolism than when embolism was artificially induced by air injection as previously shown for L. nobilis.     

Rooting depth,water relations and non‐structural carbohydrate dynamics in three woody angiosperms differentially affected by an extreme summer drought

In 2012, an extreme summer drought induced species‐specific die‐back in woody species in Northeastern Italy. Quercus pubescens and Ostrya carpinifolia were heavily impacted, while Prunus mahaleb was largely unaffected. By comparing seasonal changes in isotopic composition of xylem sap, rainfall and deep soil samples, we show that P. mahaleb has a deeper root system than the other two species. This morphological trait allowed P mahaleb to maintain higher water potential (Ψ), gas exchange rates and non‐structural carbohydrates content (NSC) throughout the summer, when compared with the other species. More favourable water and carbon states allowed relatively stable maintenance of stem hydraulic conductivity (k) throughout the growing season. In contrast, in Quercus pubescens and Ostrya carpinifolia, decreasing Ψ and NSC were associated with significant hydraulic failure, with spring‐to‐summer k loss averaging 60%. Our data support the hypothesis that drought‐induced tree decline is a complex phenomenon that cannot be modelled on the basis of single predictors of tree status like hydraulic efficiency, vulnerability and carbohydrate content. Our data highlight the role of rooting depth in seasonal progression of water status, gas exchange and NSC, with possible consequences for energy‐demanding mechanisms involved in the maintenance of vascular integrity.     

Hydraulic design of pine needles: one-dimensional optimization for single-vein leaves

Single-vein leaves have the simplest hydraulic design possible, yet even this linear water delivery system can be modulated to improve physiological performance. We determined the optimal distribution of transport capacity that minimizes pressure drop per given investment in xylem permeability along the needle for a given length without a change in total water delivery, or maximizes needle length for a given pressure difference between petiole and needle tip. This theory was tested by comparative analysis of the hydraulic design of three pine species that differ in the length of their needles [Pinus palustris (Engl.) Miller, approximately 50 cm; Pinus ponderosa Lawson & Lawson, approximately 20 cm and Pinus rigida Miller, approximately 5 cm]. In all three species, the distribution of hydraulic permeability was similar to that predicted by the optimum solution. The needles of P. palustris showed an almost perfect match between predicted and actual hydraulic optimum solution, providing evidence that vein design is a significant factor in the hydraulic design of pine leaves.     

A Simple Clear Technique in Observing Vascular Development of Grape Ovary

The vascular system of the grapevine (Vitis vinifera L.) flower is a channel for transporting water and nutrients to the ovary. It plays an important role in the development of the ovary and fertilization through pollination. However, the vascular bundles in the flower are so tiny that they are difficult to sample and observe by traditional slicing techniques. In this study, ‘Summer Black’ grape flowers were selected as the test materials, and the tissue samples were treated by the optical clearing technique. After simple compaction, the structure and development of the vasculature were observed by common microscopy, fluorescence microscopy and laser confocal microscopy. The results showed that the transparency effects of 3% NaOH and a saturated trichloroacetaldehyde composite agreed well with the observations of the vascular structure and the developmental process of the flower in different periods. Moreover, the samples after optical clearing could be reconstructed in 3D, which helped us know more about its development and function. According to these observations, the vasculature of the ‘Summer Black’ flower can be divided into ovule vascular bundles, peripheral vascular bundles and central vascular bundles. The peripheral vascular bundles were composed of the first-order vascular bundles and the inferior vascular bundles which branched from the superior vascular bundles. These bundles branched in different directions with no discernible pattern. The two different branching methods were as follows. First, the inferior vascular bundle was directly connected to a superior vascular bundle. Secondly, some of the superior vascular bundles bent in different ways, forming the inferior vascular bundle connecting the superior vascular bundles by a metamorphosed vessel with a triangular shape. In a comparison of the developmental changes in various periods, the growth of vascular bundles at each period was directly proportional to the growth of the flower. Laser confocal scanning was used to explore the three-dimensional morphology of the peripheral vascular bundle and showed that the peripheral vascular bundle of grapes was not completely parallel to the flower’s epidermal cells. As a result, the optical clearing technique was convenient and authentic compared with the traditional slicing operation for tiny flower organs. With these advantages according to the observations, this study provides a feasible technique and useful information for the study of vascular bundle development in grape flower organs.     

Abscisic acid content in the root hemiparasiteMelampyrum arvense L. before and after attachment to the host plant

The content of abscisic acid (ABA) in abaxial leaf epidermis of the host (Capsella bursa pastoris) and the unattached hemiparasiteMelampyrum arvense showed diurnal changes. ABA content increased during the light period and declined rapidly upon the darkening of leaves. In an attached hemiparasite the content of ABA in the epidermis was maintained at an almost constant level irrespective of the diurnal cycle. As compared with the maximum level in the host, at the end of the light phase the content of ABA in abaxial epidermis constituted about 70 % and 164 % in the unattached and attached hemiparasite, respectively. No significant changes in ABA content were recorded in adaxial epidermis. In all the samples abaxial/adaxial epidermis ABA content ratio was about 3.6:1 in light phase. In darkness this ratio decreased to about 1.1:1 in the host and the unattached hemiparasite and did not show significant change after attachment. ABA content ratio in mesophyll was 1:0.7:1.5 for the host, the unattached, and attached hemiparasite, respectively. In comparison with the host the concentration of ABA in xylem sap of the hemiparasite constituted about 31 % and 152 % for the unattached and attachedM. arvense, respectively.     

Does sample length influence the shape of xylem embolism vulnerability curves? A test with the Cavitron spinning technique

The Cavitron spinning technique is used to construct xylem embolism vulnerability curves (VCs), but its reliability has been questioned for species with long vessels. This technique generates two types of VC: sigmoid ‘s’‐shaped and exponential, levelling‐off ‘r’‐shaped curves. We tested the hypothesis that ‘r’‐shaped VCs were anomalous and caused by the presence of vessels cut open during sample preparation. A Cavitron apparatus was used to construct VCs from samples of different lengths in species with contrasting vessel lengths. The results were compared with VCs obtained using other independent techniques. When vessel length exceeded sample length, VCs were ‘r’‐shaped and anomalous. Filling vessels cut open at both ends with air before measurement produced more typical ‘s’‐shaped VCs. We also found that exposing segments of 11 woody species in a Cavitron at the pressure measured in planta before sampling considerably increased the degree of embolism above the native state level for species with long vessels. We concluded that open vessels were abnormally more vulnerable to cavitation than intact vessels. We recommend restricting this technique to species with short conduits. The relevance of our conclusions for other spinning techniques is discussed.     

Genetic determinism of anatomical and hydraulic traits within an apple progeny

The apple tree is known to have an isohydric behaviour, maintaining rather constant leaf water potential in soil with low water status and/or under high evaporative demand. However, little is known on the xylem water transport from roots to leaves from the two perspectives of efficiency and safety, and on its genetic variability. We analysed 16 traits related to hydraulic efficiency and safety, and anatomical traits in apple stems, and the relationships between them. Most variables were found heritable, and we investigated the determinism underlying their genetic control through a quantitative trait loci (QTL) analysis on 90 genotypes from the same progeny. Principal component analysis (PCA) revealed that all traits related to efficiency, whether hydraulic conductivity, vessel number and area or wood area, were included in the first PC, whereas the second PC included the safety variables, thus confirming the absence of trade-off between these two sets of traits. Our results demonstrated that clustered variables were characterized by common genomic regions. Together with previous results on the same progeny, our study substantiated that hydraulic efficiency traits co-localized with traits identified for tree growth and fruit production.     

Vulnerability curves by centrifugation: is there an open vessel artefact,and are ‘r’ shaped curves necessarily invalid?

Vulnerability curves using the 'Cavitron' centrifuge rotor yield anomalous results when vessels extend from the end of the stem segment to the centre ('open-to-centre' vessels). Curves showing a decline in conductivity at modest xylem pressures ('r' shaped) have been attributed to this artefact. We determined whether the original centrifugal method with its different rotor is influenced by open-to-centre vessels. Increasing the proportion of open-to-centre vessels by shortening stems had no substantial effect in four species. Nor was there more embolism at the segment end versus centre as seen in the Cavitron. The dehydration method yielded an 'r' shaped curve in Quercus gambelii that was similar to centrifuged stems with 86% open-to-centre vessels. Both 'r' and 's' (sigmoidal) curves from Cercocarpus intricatus were consistent with each other, differing only in whether native embolism had been removed. An 'r' shaped centrifuge curve in Olea europaea was indistinguishable from the loss of conductivity caused by forcing air directly across vessel end-walls. We conclude that centrifuge curves on long-vesselled material are not always prone to the open vessel artefact when the original rotor design is used, and 'r' shaped curves are not necessarily artefacts. Nevertheless, confirming curves with native embolism and dehydration data is recommended.     

植物木质部水力学研究进展

植物为适应陆地环境进化出木质部维管系统,通过水力学机制高效安全的向光合器官长距离运输水分,木质部水分运输对蒸腾、气孔运动、光合碳同化等生理过程有调控和协调作用,被称为植物生理学的支柱。植物水力学作为木质部水分运输的研究内容和手段,已成为整合植物与生态系统功能的中心枢纽。该文首先概述了植物水分运输的水力学机制、运输系统的局限性,以及木质部结构与功能之间的关系;其次,阐述了木质部栓塞的形成机制并详细介绍了栓塞的诱导方法和测试技术,分析了水分运输系统安全与效率之间的权衡关系,总结了植物对环境的响应和干旱致死的预测模型,讨论了测试技术问题及其引发的当前木质部逆压力修复和指数型木质部栓塞脆弱性曲线有效性的争议;最后,总结了目前植物木质部水力学研究的成果,提出了尚待解决的主要问题,探讨了研究机会与方向。