Hugh Morris

A global analysis of parenchyma tissue fractions in secondary xylem of seed plants

Summary Parenchyma is an important tissue in secondary xylem of seed plants, with functions ranging from storage to defence and with effects on the physical and mechanical properties of wood. Currently, we lack a large‐scale quantitative analysis of ray parenchyma (RP) and axial parenchyma (AP) tissue fractions. Here, we use data from the literature on AP and RP fractions to investigate the potential relationships of climate and growth form with total ray and axial parenchyma fractions (RAP). We found a 29‐fold variation in RAP fraction, which was more strongly related to temperature than with precipitation. Stem succulents had the highest RAP values (mean ± SD: 70.2 ± 22.0%), followed by lianas (50.1 ± 16.3%), angiosperm trees and shrubs (26.3 ± 12.4%), and conifers (7.6 ± 2.6%). Differences in RAP fraction between temperate and tropical angiosperm trees (21.1 ± 7.9% vs 36.2 ± 13.4%, respectively) are due to differences in the AP fraction, which is typically three times higher in tropical than in temperate trees, but not in RP fraction. Our results illustrate that both temperature and growth form are important drivers of RAP fractions. These findings should help pave the way to better understand the various functions of RAP in plants.

Are needles of Pinus pinaster more vulnerable to xylem embolism than branches? New insights from X‐ray computed tomography

Plants can be highly segmented organisms with an independently redundant design of organs. In the context of plant hydraulics, leaves may be less embolism resistant than stems, allowing hydraulic failure to be restricted to distal organs that can be readily replaced. We quantified drought-induced embolism in needles and stems of Pinus pinaster using high-resolution computed tomography (HRCT). HRCT observations of needles were compared with the rehydration kinetics method to estimate the contribution of extra-xylary pathways to declining hydraulic conductance. High-resolution computed tomography images indicated that the pressure inducing 50% of embolized tracheids was similar between needle and stem xylem (P50 needle xylem  = -3.62 MPa, P50 stem xylem  = -3.88 MPa). Tracheids in both organs showed no difference in torus overlap of bordered pits. However, estimations of the pressure inducing 50% loss of hydraulic conductance at the whole needle level by the rehydration kinetics method were significantly higher (P50 needle  = -1.71 MPa) than P50 needle xylem derived from HRCT. The vulnerability segmentation hypothesis appears to be valid only when considering hydraulic failure at the entire needle level, including extra-xylary pathways. Our findings suggest that native embolism in needles is limited and highlight the importance of imaging techniques for vulnerability curves.

Anatomical features associated with water transport in imperforate tracheary elements of vessel-bearing angiosperms

BACKGROUND AND AIMS Imperforate tracheary elements (ITEs) in wood of vessel-bearing angiosperms may or may not transport water. Despite the significance of hydraulic transport for defining ITE types, the combination of cell structure with water transport visualization in planta has received little attention. This study provides a quantitative analysis of structural features associated with the conductive vs. non-conductive nature of ITEs. METHODS Visualization of water transport was studied in 15 angiosperm species by dye injection and cryo-scanning electron microscopy. Structural features of ITEs were examined using light and electron microscopy. KEY RESULTS ITEs connected to each other by pit pairs with complete pit membranes contributed to water transport, while cells showing pit membranes with perforations up to 2 µm were hydraulically not functional. A close relationship was found between pit diameter and pit density, with both characters significantly higher in conductive than in non-conductive cells. In species with both conductive and non-conductive ITEs, a larger diameter was characteristic of the conductive cells. Water transport showed no apparent relationship with the length of ITEs and vessel grouping. CONCLUSIONS The structure and density of pits between ITEs represent the main anatomical characters determining water transport. The pit membrane structure of ITEs provides a reliable, but practically challenging, criterion to determine their conductive status. It is suggested that the term tracheids should strictly be used for conductive ITEs, while fibre-tracheids and libriform fibres are non-conductive.

The amount of parenchyma and living fibers affects storage of nonstructural carbohydrates in young stems and roots of temperate trees

PREMISE OF THE STUDY Concentrations of nonstructural carbohydrates (NSCs) are used as proxies for the net carbon balance of trees and as indicators of carbon starvation resulting from environmental stress. Woody organs are the largest NSC-storing compartments in forest ecosystems; therefore, it is essential to understand the factors that affect the size of this important storage pool. In wood, NSC are predominantly deposited in ray and axial parenchyma (RAP); however, direct links between nutrient storage and RAP anatomy have not yet been established. Here, we tested whether the NSC storage capacity of wood is influenced by the amount of RAP. METHODS We measured NSC concentrations and RAP fractions in root and stem sapwood of 12 temperate species sampled at the onset of winter dormancy and in stem sapwood of four tropical trees growing in an evergreen lowland rainforest. The patterns of starch distribution were visualized by staining with Lugols solution. KEY RESULTS The concentration of NSCs in sapwood of temperate trees scales tightly with the amount of RAP and living fibers (LFs), with almost all RAP and LFs being densely packed with starch grains. In contrast, the tropical species had lower NSC concentrations despite their higher RAP and LFs fraction and had considerable interspecific differences in starch distribution. CONCLUSIONS The differences in RAP and LFs abundance affect the ability of sapwood to store NSC in temperate trees, whereas a more diverse set of functions of RAP might be pronounced in species growing in a tropical environment with little seasonality.

The Parenchyma of Secondary Xylem and Its Critical Role in Tree Defense against Fungal Decay in Relation to the CODIT Model

This review examines the roles that ray and axial parenchyma (RAP) plays against fungal pathogens in the secondary xylem of wood within the context of the CODIT model (Compartmentalization of Decay in Trees), a defense concept first conceived in the early 1970s by Alex Shigo. This model, simplistic in its design, shows how a large woody perennial is highly compartmented. Anatomical divisions in place at the time of infection or damage, (physical defense) alongside the ‘active’ response by the RAP during and after wounding work together in forming boundaries that function to restrict air or decay spread. The living parenchyma cells play a critical role in all of the four walls (differing anatomical constructs) that the model comprises. To understand how living cells in each of the walls of CODIT cooperate, we must have a clear vision of their complex interconnectivity from a three-dimensional perspective, along with knowledge of the huge variation in ray parenchyma (RP) and axial parenchyma (AP) abundance and patterns. Crucial patterns for defense encompass the symplastic continuum between both RP and AP and the dead tissues, with the latter including the vessel elements, libriform fibers, and imperforate tracheary elements (i.e., vasicentric and vascular tracheids). Also, the heartwood, a chemically altered antimicrobial non-living substance that forms the core of many trees, provides an integral part of the defense system. In the heartwood, dead RAP can play an important role in defense, depending on the genetic constitution of the species. Considering the array of functions that RAP are associated with, from capacitance, through to storage, and long-distance water transport, deciding how their role in defense fits into this suite of functions is a challenge for plant scientists, and likely depends on a range of factors. Here, we explore the important role of RAP in defense against fungal pathogens and the trade-offs involved from a viewpoint for structure-function relations, while also examining how fungi can breach the defense system using an array of enzymes in conjunction with the physically intrusive hyphae.

Vessel diameter is related to amount and spatial arrangement of axial parenchyma in woody angiosperms

Parenchyma represents a critically important living tissue in the sapwood of the secondary xylem of woody angiosperms. Considering various interactions between parenchyma and water transporting vessels, we hypothesize a structure-function relationship between both cell types. Through a generalized additive mixed model approach based on 2,332 woody angiosperm species derived from the literature, we explored the relationship between the proportion and spatial distribution of ray and axial parenchyma and vessel size, while controlling for maximum plant height and a range of climatic factors. When factoring in maximum plant height, we found that with increasing mean annual temperatures, mean vessel diameter showed a positive correlation with axial parenchyma proportion and arrangement, but not for ray parenchyma. Species with a high axial parenchyma tissue fraction tend to have wide vessels, with most of the parenchyma packed around vessels, whereas species with small diameter vessels show a reduced amount of axial parenchyma that is not directly connected to vessels. This finding provides evidence for independent functions of axial parenchyma and ray parenchyma in large vesselled species and further supports a strong role for axial parenchyma in long-distance xylem water transport.

Vessel-associated cells in angiosperm xylem: Highly specialized living cells at the symplast-apoplast boundary

BACKGROUND Vessel-associated cells (VACs) are highly specialized, living parenchyma cells that are in direct contact with water-conducting, dead vessels. The contact may be sparse or in large tight groups of parenchyma that completely surrounds vessels. VACs differ from vessel distant parenchyma in physiology, anatomy, and function and have half-bordered pits at the vessel-parenchyma juncture. The distinct anatomy of VACs is related to the exchange of substances to and from the water-transport system, with the cells long thought to be involved in water transport in woody angiosperms, but where direct experimental evidence is lacking. SCOPE This review focuses on our current knowledge of VACs regarding anatomy and function, including hydraulic capacitance, storage of nonstructural carbohydrates, symplastic and apoplastic interactions, defense against pathogens and frost, osmoregulation, and the novel hypothesis of surfactant production. Based on microscopy, we visually represent how VACs vary in dimensions and general appearance between species, with special attention to the protoplast, amorphous layer, and the vessel-parenchyma pit membrane. CONCLUSIONS An understanding of the relationship between VACs and vessels is crucial to tackling questions related to how water is transported over long distances in xylem, as well as defense against pathogens. New avenues of research show how parenchyma-vessel contact is related to vessel diameter and a new hypothesis may explain how surfactants arising from VAC can allow water to travel under negative pressure. We also reinforce the message of connectivity between VAC and other cells between xylem and phloem.

On research priorities to advance understanding of the safety–efficiency tradeoff in xylem

We appreciate Bittencourt et al.’s (2016) constructive contributions following our paper, Gleason et al. (2016), on the proposed tradeoff between hydraulic safety and efficiency. To continue this dialog we would like to comment on which of the research directions proposed by Bittencourt et al. seem most promising to us. We agree that various xylem tissue fractions could potentially modify the safety–efficiency relationship. In principle, any tissue fraction could trade off with any other tissue fraction. However, as a matter of observation, parenchyma fraction is negatively correlated with fiber fraction, whereas parenchyma and fiber fractions are not strongly correlated with vessel lumen fraction (Ziemi! nska et al., 2015; Morris et al., 2016). As such, vessel lumen fraction, vessel diameter, and vessel frequency are largely uncoupled from nonvessel tissue fractions across self-supporting angiosperm species (Zanne et al., 2010), and are therefore unlikely to trade off with mechanical safety and hydraulic efficiency (or safety). Furthermore, vessel lumen fraction itself does not vary markedly across angiosperms, ranging from c. 5% to 20% (mean! 15%) (Zanne et al., 2010; Morris et al., 2016), although larger fractions are not uncommon in ring-porous and climbing species. Contrasts between climbing (e.g. lianas) and freestanding growth forms are more likely to show differences in allocation to vessel vs nonvessel space, and the climbing habit therefore may offer a more appropriate system for evaluating this idea (Gartner, 1991). Gymnosperm xylem differs from angiosperm xylem in that it generally lacks axial parenchyma, and conduits are both conductive and load-bearing. Greater mechanical safety may be negatively correlated with both hydraulic safety and efficiency across gymnosperm species (Mayr & Cochard, 2003; Mayr et al., 2003). Considering angiosperms, it is likely that nonvessel tissue fractions influence the safety–efficiency tradeoff indirectly (e.g. via their contribution to xylem capacitance, or whole-plant growth), rather than being forced by limited xylem space. Bittencourt et al. suggest that expressing conductivity as a ratio with mass, rather than cross-sectional area, might better characterize the energetic costs associated with xylem. We agree with this suggestion and did consider the influence of specific gravity on the safety–efficiency tradeoff in our paper (Table 2 and Figs 3d, 4d in Gleason et al., 2016). Here, we formulate these results by expressing the y axis explicitly as xylem-specific conductivity/ specific gravity (Fig. 1), as suggested by Bittencourt et al. Specific gravity, safety and efficiency values were generally obtained from the same published reports. Similar to the results we report in Gleason et al. (2016), including specific gravity in the analyses increases the tradeoff r 2 in both angiosperms (0.11–0.14) and gymnosperms (0.10–0.15) when safety is defined as P50. A similar increase in r 2 is achieved when defining safety as P88 and there is no change when defining safety as P12. Although including xylem density does increase the amount of variation explained by the models, they still fall far short of explaining why many species exhibit both low safety and low efficiency. Despite the analysis provided in Fig. 1, we feel that the clearest approach to analyzing these inter-correlated variables will be to consider all known sources of variation (e.g. structural equation models) rather than expressing them as a ratio with conductivity (e.g. conductivity/parenchyma fraction). Such ratios build an assumption of proportionality between the two elements of the ratio, which are not necessarily what we should expect. It remains a possibility that xylem safety, expressed as the xylem water potential at which a fraction of maximal conductance is lost, may not be an accurate approximation for all species in all situations. Although we agree in principle that safety (e.g. P50) may not correlate with mortality similarly across species, there is good evidence to suggest that it does for many angiosperm and gymnosperm species (Pratt et al., 2008; Brodribb & Cochard, 2009; Brodribb et al., 2010). This suggests that there may be an intrinsic property of xylem to resist desiccation (angiosperm Ψleaf > P88; gymnosperm Ψleaf > P50), beyond which the probability of mortality increases precipitously. Measurements of hydraulic safety, as well as conductivity during drought, should serve as more appropriate predictors of mortality than other measurements of water status (e.g. turgor loss point in leaves or stomatal response) because percentage loss of conductance is a meaningful representation of xylem desiccation. However, it is also clear that there are mechanisms that delay the time to reach a desiccation–mortality threshold. As suggested by Bittencourt et al. and Brodersen (2016), these would include deciduousness, deep rooting, reduced stomatal ‘leakiness’, reduced cuticular conductance, CAM and C4 metabolism, and capacitance. However, considering tradeoffs with either safety or efficiency, in isolation of one another (e.g. safety–capacitance), does not inform our efforts to understand the proposed link between safety and efficiency. For example, if greater capacitance reduces the requirement for safety, natural selection should still be free to improve efficiency, which would provide benefit via greater

On research priorities to advance understanding of the safety-efficiency tradeoff in xylem: A response to Bittencourt et al.'s (2016) comment 'On xylem hydraulic efficiencies, wood space-use and the safety-efficiency tradeoff': in this issue of New Phytologist, pp. 1152-1155

We appreciate Bittencourt et al.’s (2016) constructive contributions following our paper, Gleason et al. (2016), on the proposed tradeoff between hydraulic safety and efficiency. To continue this dialog we would like to comment on which of the research directions proposed by Bittencourt et al. seem most promising to us. We agree that various xylem tissue fractions could potentially modify the safety–efficiency relationship. In principle, any tissue fraction could trade off with any other tissue fraction. However, as a matter of observation, parenchyma fraction is negatively correlated with fiber fraction, whereas parenchyma and fiber fractions are not strongly correlated with vessel lumen fraction (Ziemi! nska et al., 2015; Morris et al., 2016). As such, vessel lumen fraction, vessel diameter, and vessel frequency are largely uncoupled from nonvessel tissue fractions across self-supporting angiosperm species (Zanne et al., 2010), and are therefore unlikely to trade off with mechanical safety and hydraulic efficiency (or safety). Furthermore, vessel lumen fraction itself does not vary markedly across angiosperms, ranging from c. 5% to 20% (mean! 15%) (Zanne et al., 2010; Morris et al., 2016), although larger fractions are not uncommon in ring-porous and climbing species. Contrasts between climbing (e.g. lianas) and freestanding growth forms are more likely to show differences in allocation to vessel vs nonvessel space, and the climbing habit therefore may offer a more appropriate system for evaluating this idea (Gartner, 1991). Gymnosperm xylem differs from angiosperm xylem in that it generally lacks axial parenchyma, and conduits are both conductive and load-bearing. Greater mechanical safety may be negatively correlated with both hydraulic safety and efficiency across gymnosperm species (Mayr & Cochard, 2003; Mayr et al., 2003). Considering angiosperms, it is likely that nonvessel tissue fractions influence the safety–efficiency tradeoff indirectly (e.g. via their contribution to xylem capacitance, or whole-plant growth), rather than being forced by limited xylem space. Bittencourt et al. suggest that expressing conductivity as a ratio with mass, rather than cross-sectional area, might better characterize the energetic costs associated with xylem. We agree with this suggestion and did consider the influence of specific gravity on the safety–efficiency tradeoff in our paper (Table 2 and Figs 3d, 4d in Gleason et al., 2016). Here, we formulate these results by expressing the y axis explicitly as xylem-specific conductivity/ specific gravity (Fig. 1), as suggested by Bittencourt et al. Specific gravity, safety and efficiency values were generally obtained from the same published reports. Similar to the results we report in Gleason et al. (2016), including specific gravity in the analyses increases the tradeoff r 2 in both angiosperms (0.11–0.14) and gymnosperms (0.10–0.15) when safety is defined as P50. A similar increase in r 2 is achieved when defining safety as P88 and there is no change when defining safety as P12. Although including xylem density does increase the amount of variation explained by the models, they still fall far short of explaining why many species exhibit both low safety and low efficiency. Despite the analysis provided in Fig. 1, we feel that the clearest approach to analyzing these inter-correlated variables will be to consider all known sources of variation (e.g. structural equation models) rather than expressing them as a ratio with conductivity (e.g. conductivity/parenchyma fraction). Such ratios build an assumption of proportionality between the two elements of the ratio, which are not necessarily what we should expect. It remains a possibility that xylem safety, expressed as the xylem water potential at which a fraction of maximal conductance is lost, may not be an accurate approximation for all species in all situations. Although we agree in principle that safety (e.g. P50) may not correlate with mortality similarly across species, there is good evidence to suggest that it does for many angiosperm and gymnosperm species (Pratt et al., 2008; Brodribb & Cochard, 2009; Brodribb et al., 2010). This suggests that there may be an intrinsic property of xylem to resist desiccation (angiosperm Ψleaf > P88; gymnosperm Ψleaf > P50), beyond which the probability of mortality increases precipitously. Measurements of hydraulic safety, as well as conductivity during drought, should serve as more appropriate predictors of mortality than other measurements of water status (e.g. turgor loss point in leaves or stomatal response) because percentage loss of conductance is a meaningful representation of xylem desiccation. However, it is also clear that there are mechanisms that delay the time to reach a desiccation–mortality threshold. As suggested by Bittencourt et al. and Brodersen (2016), these would include deciduousness, deep rooting, reduced stomatal ‘leakiness’, reduced cuticular conductance, CAM and C4 metabolism, and capacitance. However, considering tradeoffs with either safety or efficiency, in isolation of one another (e.g. safety–capacitance), does not inform our efforts to understand the proposed link between safety and efficiency. For example, if greater capacitance reduces the requirement for safety, natural selection should still be free to improve efficiency, which would provide benefit via greater

On research priorities to advance understanding of the safety-efficiency tradeoff in xylem: A response to Bittencourtet al.'s (2016) comment ‘On xylem hydraulic efficiencies, wood space-use and the safety-efficiency tradeoff’

We appreciate Bittencourt et al.’s (2016) constructive contributions following our paper, Gleason et al. (2016), on the proposed tradeoff between hydraulic safety and efficiency. To continue this dialog we would like to comment on which of the research directions proposed by Bittencourt et al. seem most promising to us. We agree that various xylem tissue fractions could potentially modify the safety–efficiency relationship. In principle, any tissue fraction could trade off with any other tissue fraction. However, as a matter of observation, parenchyma fraction is negatively correlated with fiber fraction, whereas parenchyma and fiber fractions are not strongly correlated with vessel lumen fraction (Ziemi! nska et al., 2015; Morris et al., 2016). As such, vessel lumen fraction, vessel diameter, and vessel frequency are largely uncoupled from nonvessel tissue fractions across self-supporting angiosperm species (Zanne et al., 2010), and are therefore unlikely to trade off with mechanical safety and hydraulic efficiency (or safety). Furthermore, vessel lumen fraction itself does not vary markedly across angiosperms, ranging from c. 5% to 20% (mean! 15%) (Zanne et al., 2010; Morris et al., 2016), although larger fractions are not uncommon in ring-porous and climbing species. Contrasts between climbing (e.g. lianas) and freestanding growth forms are more likely to show differences in allocation to vessel vs nonvessel space, and the climbing habit therefore may offer a more appropriate system for evaluating this idea (Gartner, 1991). Gymnosperm xylem differs from angiosperm xylem in that it generally lacks axial parenchyma, and conduits are both conductive and load-bearing. Greater mechanical safety may be negatively correlated with both hydraulic safety and efficiency across gymnosperm species (Mayr & Cochard, 2003; Mayr et al., 2003). Considering angiosperms, it is likely that nonvessel tissue fractions influence the safety–efficiency tradeoff indirectly (e.g. via their contribution to xylem capacitance, or whole-plant growth), rather than being forced by limited xylem space. Bittencourt et al. suggest that expressing conductivity as a ratio with mass, rather than cross-sectional area, might better characterize the energetic costs associated with xylem. We agree with this suggestion and did consider the influence of specific gravity on the safety–efficiency tradeoff in our paper (Table 2 and Figs 3d, 4d in Gleason et al., 2016). Here, we formulate these results by expressing the y axis explicitly as xylem-specific conductivity/ specific gravity (Fig. 1), as suggested by Bittencourt et al. Specific gravity, safety and efficiency values were generally obtained from the same published reports. Similar to the results we report in Gleason et al. (2016), including specific gravity in the analyses increases the tradeoff r 2 in both angiosperms (0.11–0.14) and gymnosperms (0.10–0.15) when safety is defined as P50. A similar increase in r 2 is achieved when defining safety as P88 and there is no change when defining safety as P12. Although including xylem density does increase the amount of variation explained by the models, they still fall far short of explaining why many species exhibit both low safety and low efficiency. Despite the analysis provided in Fig. 1, we feel that the clearest approach to analyzing these inter-correlated variables will be to consider all known sources of variation (e.g. structural equation models) rather than expressing them as a ratio with conductivity (e.g. conductivity/parenchyma fraction). Such ratios build an assumption of proportionality between the two elements of the ratio, which are not necessarily what we should expect. It remains a possibility that xylem safety, expressed as the xylem water potential at which a fraction of maximal conductance is lost, may not be an accurate approximation for all species in all situations. Although we agree in principle that safety (e.g. P50) may not correlate with mortality similarly across species, there is good evidence to suggest that it does for many angiosperm and gymnosperm species (Pratt et al., 2008; Brodribb & Cochard, 2009; Brodribb et al., 2010). This suggests that there may be an intrinsic property of xylem to resist desiccation (angiosperm Ψleaf > P88; gymnosperm Ψleaf > P50), beyond which the probability of mortality increases precipitously. Measurements of hydraulic safety, as well as conductivity during drought, should serve as more appropriate predictors of mortality than other measurements of water status (e.g. turgor loss point in leaves or stomatal response) because percentage loss of conductance is a meaningful representation of xylem desiccation. However, it is also clear that there are mechanisms that delay the time to reach a desiccation–mortality threshold. As suggested by Bittencourt et al. and Brodersen (2016), these would include deciduousness, deep rooting, reduced stomatal ‘leakiness’, reduced cuticular conductance, CAM and C4 metabolism, and capacitance. However, considering tradeoffs with either safety or efficiency, in isolation of one another (e.g. safety–capacitance), does not inform our efforts to understand the proposed link between safety and efficiency. For example, if greater capacitance reduces the requirement for safety, natural selection should still be free to improve efficiency, which would provide benefit via greater