The Relationships between Xylem Safety and Hydraulic Efficiency in the Cupressaceae: The Evolution of Pit Membrane Form and Function

Water transport in conifers occurs through single-celled tracheids that are connected to one another via intertracheid pit membranes. These membranes have two components: the porous margo, which allows water to pass through the membrane, and the impermeable torus, which functions to isolate gas-filled tracheids. During drought, tracheids can become air filled and thus hydraulically dysfunctional, a result of air entering through the pit membrane and nucleating cavitation in the water column. What are the hydraulic tradeoffs associated with cavitation resistance at the pit level, and how do they vary within the structural components of the intertracheid pit? To address these questions, we examined pit structure in 15 species of Cupressaceae exhibiting a broad range of cavitation resistances. Across species, cavitation resistance was most closely correlated to the ratio of the torus to pit aperture diameter but did not vary systematically with margo porosity. Furthermore, our data indicate that constraints on pit hydraulic efficiency are shared: the pit aperture limits pit conductivity in more drought-resistant taxa, while increased margo resistance is more likely to control pit conductivity in species that are more vulnerable to cavitation. These results are coupled with additional data concerning pit membrane structure and function and are discussed in the context of the evolutionary biogeography of the Cupressaceae.Water transport in conifers occurs through narrow, single-celled conduits (tracheids) that are organized in overlapping, longitudinal files. This simple, homoxylous arrangement represents an ancestral vascular design that has remained remarkably consistent since its first appearance in the progymnosperms of the Mid-Devonian (Taylor et al., 2009). However, the small size of tracheids can impose a high resistance to water transport as compared with the large, hydraulically efficient vessels present in many angiosperms (Hacke et al., 2004; Sperry et al., 2006). Despite this handicap, conifer tracheids can be just as hydraulically efficient as angiosperm xylem for a given conduit diameter, a result that can be wholly attributed to the distinctive structure of the conifer intertracheid pit membrane (Pittermann et al., 2005; Sperry et al., 2006).Because pit membranes also function to limit the spread of air from one conduit to another (cavitation), the physiological consequences of the transport efficiency versus cavitation safety tradeoffs in conifer and angiosperm pit membranes have received considerable attention at the pit and xylem levels, whereby cavitation resistance in north temperate woody plants appears to come at the cost of hydraulic efficiency (Pittermann et al., 2006a, 2006b; Sperry et al., 2006; Choat et al., 2008; Domec et al., 2008; Jansen et al., 2009; Schoonmaker et al., 2010). Previous work has shown that the integrated vascular performance of plants is key to understanding species distributions (Sperry et al., 1994; Brodribb and Hill, 1999; Pockman and Sperry, 2000; Choat et al., 2007), and within this framework, pit membranes have the potential to act as the nexus of the cavitation safety versus transport efficiency compromise. Yet, despite our progress, we are just starting to learn how these tradeoffs play out at the level of the pit membrane, particularly in one as complex as that of conifers. Hence, the goals of this study were to determine whether selection has acted to optimize conifer pit membrane performance in a manner that reflects species cavitation resistance and habitat distribution as well as to examine the role, if any, of evolutionary lineage.Unlike the homogenous pit membrane of angiosperm vessels, the conifer pit membrane is composed of two distinct regions: a thickened, centrally located torus and a porous margo region that surrounds it (Fig. 1; for study species, see Hacke et al., 2004; Choat et al., 2008; Choat and Pittermann, 2009). When tracheids are water filled, the pit membrane is centrally located in the pit chamber and water moves from tracheid to tracheid through the margo. Should an air-seeding event (cavitation) occur, causing a tracheid to become air filled, (i.e. embolized), the negative xylem pressure in the water-filled tracheid will act on the air-water interface in the margo pores by deflecting the pit membrane in the direction of the functional tracheid, thereby appressing the torus against the pit aperture border (Bailey, 1913; Liese, 1965; Liese and Bauch, 1967; Petty, 1972). This valve action of the membrane can create an effective seal that prevents further spread of air in the xylem. Cavitation is thought to occur when the water potential of the water-filled tracheid becomes negative enough to dislodge the torus from its sealing position, allowing air to enter the conduit. Overall, the structure of the torus-margo pit membrane must optimize what at first glance appear to be conflicting functional requirements: on the one hand, cavitation resistance selects for a combination of large tori and small apertures, but on the other hand, hydraulic efficiency favors porous margos, large apertures, and small tori.Open in a separate windowFigure 1.SEM images of intertracheid pit membranes belonging to nine Cupressaceae species (of 15) that represent the broad range of observed cavitation pressures. The opaque torus region of the membrane (T) is held centrally by the microfibrils of the margo (M). Visually, increased cavitation resistance appears to be associated with increased margo porosity, but quantitative estimates of margo resistance made on the most intact regions of the pit membranes (Fig. 8) revealed no differences among the species surveyed.

Table I.

Study species, figure abbreviations (Fig. Abbrevs.), locations (SFBG, San Francisco Botanical Garden, San Francisco; UCBG, University of California Botanical Garden, Berkeley, CA; UCSC, University of California, Santa Cruz, Arboretum, Santa Cruz, CA), and species natural history (Farjon, 2005)

Bordered pit structure and function determine spatial patterns of air-seeding thresholds in xylem of Douglas-fir (Pseudotsuga menziesii; Pinaceae) trees

The air-seeding hypothesis predicts that xylem embolism resistance is linked directly to bordered pit functioning. We tested this prediction in trunks, roots, and branches at different vertical and radial locations in young and old trees of Pseudotsuga menziesii. Dimensions of bordered pits were measured from light and scanning electron micrographs, and physiological data were from published values. Consistent with observations, calculations showed that earlywood tracheids were more resistant to embolism than latewood tracheids, mainly from earlywood having stretchier pit membranes that can distend and cover the pit aperture. Air seeding that occurs in earlywood appears to happen through gaps between the torus edge and pit border, as shown by the similar calculated pressures required to stretch the membrane over the pit aperture and to cause embolism. Although bordered pit functioning was correlated with tracheid hydraulic diameter, pit pore size and above all pit aperture constrained conductivity the most. From roots to branches and from the trunk base to higher on the trunk, hydraulic resistance of the earlywood pit membrane increased significantly because of a decrease in the size of the pit aperture and size and number of margo pores. Moreover, overall wood conductivity decreased, in part due to lower pit conductivity and a decrease in size and frequency of pits. Structural and functional constraints leading to the trade-off of efficiency against safety of water transport were also demonstrated at the individual pit level, with a positive correlation between pit membrane resistance on an area basis and the pressure differential required to cause membrane stretching, a characteristic that is essential for pit aspiration.     

Plasmodesmatal pores in the torus of bordered pit membranes affect cavitation resistance of conifer xylem

The pit membrane in bordered pits of conifer tracheids is characterized by a porous margo and central thickening (torus), which is traditionally considered to function as an impermeable safety valve against air-seeding. However, electron microscopy based on 33 conifer species, including five families and 19 genera, reveals that pores occur in the torus of 13 of the species studied. The pores have a plasmodesmatal origin with an average diameter of 51 nm and grouped arrangement. Evidence for embolism spreading via pores in tori is supported by the pore sizes, which correspond relatively well with the pressure inducing cavitation. Predictions based on earlier correlations between pit structure and cavitation resistance were only weakly supported for species with punctured tori. Moreover, species with punctured tori are significantly less resistant to cavitation than species with non-punctured tori. Nevertheless, absolute pore diameters must be treated with caution and correlations between theoretical and measured air-seeding pressures are weak. Because most pores appear not to traverse the torus but are limited to one torus pad, only complete pores would trigger air-seeding. Embolism spreading through a leaky torus is not universal across gymnosperms and unlikely to represent the only air-seeding mechanism.     

Computational fluid dynamics models of conifer bordered pits show how pit structure affects flow

? The flow of xylem sap through conifer bordered pits, particularly through the pores in the pit membrane, is not well understood, but is critical for an understanding of water transport through trees. ? Models solving the Navier-Stokes equation governing fluid flow were based on the geometry of bordered pits in black spruce (Picea mariana) and scanning electron microscopy images showing details of the pores in the margo of the pit membrane. ? Solutions showed that the pit canals contributed a relatively small fraction of resistance to flow, whereas the torus and margo pores formed a large fraction, which depended on the structure of the individual pit. The flow through individual pores in the margo was strongly dependent on pore area, but also on the radial location of the pore with respect to the edge of the torus. ? Model results suggest that only a few per cent of the pores in the margo account for nearly half of the flow and these pores tend to be located in the inner region of the margo where their contribution will be maximized. A high density of strands in outer portions of the margo (hence narrower pores) may be more significant for mechanical support of the torus.     

Force–displacement measurements of earlywood bordered pits using a mesomechanical tester

The elastic properties of pit membranes are reported to have important implications in understanding air‐seeding phenomena in gymnosperms, and pit aspiration plays a large role in wood technological applications such as wood drying and preservative treatment. Here we present force–displacement measurements for pit membranes of circular bordered pits, collected on a mesomechanical testing system. The system consists of a quartz microprobe attached to a microforce sensor that is positioned and advanced with a micromanipulator mounted on an inverted microscope. Membrane displacement is measured from digital image analysis. Unaspirated pits from earlywood of never‐dried wood of Larix and Pinus and aspirated pits from earlywood of dried wood of Larix were tested to generate force–displacement curves up to the point of membrane failure. Two failure modes were observed: rupture or tearing of the pit membrane by the microprobe tip, and the stretching of the pit membrane until the torus was forced out of the pit chamber through the pit aperture without rupture, a condition we refer to as torus prolapse.     

Hydraulic acclimation to shading in boreal conifers of varying shade tolerance

The purpose of this study was to determine how shading affects the hydraulic and wood‐anatomical characteristics of four boreal conifers (Pinus banksiana, Pinus contorta, Picea glauca and Picea mariana) that differ in shade tolerance. Plants were grown in an open field and under a deciduous‐dominated overstory for 6 years. Sapwood‐ and leaf‐area specific conductivity, vulnerability curves, and anatomical measurements (light and scanning electron microscopy) were made on leading shoots from six to nine trees of each treatment combination. There was no difference in sapwood‐area specific conductivity between open‐grown and understory conifers, although two of four species had larger tracheid diameters in the open. Shaded conifers appeared to compensate for small diameter tracheids by changes in pit membrane structure. Scanning electron microscopy revealed that understory conifers had thinner margo strands, greater maximum pore size in the margo, and more torus extensions. All of these trends may contribute to inadequate sealing of the torus. This is supported by the fact that all species showed increased vulnerability to cavitation when grown in the understory. Although evaporative demand in an understory environment is low, a rapid change into fully exposed conditions could be detrimental for shaded conifers.     

Drought‐induced xylem pit membrane damage in aspen and balsam poplar

Drought induces an increase in a tree's vulnerability to a loss of its hydraulic conductivity in many tree species, including two common in western Canada, trembling aspen (Populus tremuloides) and balsam poplar (Populus balsamifera). Termed ‘cavitation fatigue’ or ‘air‐seeding fatigue’, the mechanism of this phenomenon is not well understood, but hypothesized to be a result of damage to xylem pit membranes. To examine the validity of this hypothesis, the effect of drought on the porosity of pit membranes in aspen and balsam poplar was investigated. Controlled drought and bench dehydration treatments were used to induce fatigue and scanning electron microscopy (SEM) was used to image pit membranes for relative porosity evaluations from air‐dried samples after ethanol dehydration. A significant increase in the diameter of the largest pore was found in the drought and dehydration treatments of aspen, while an increase in the percentage of porous pit membranes was found in the dehydration treatments of both species. Additionally, the location of the largest pore per pit membrane was observed to tend toward the periphery of the membrane.     

Water-stress-induced xylem embolism in three species of conifers

Abstract. The mechanism of water-stress-induced xylem embolism was studied in three species of conifers: Abies balsamea (L.) Mill., Picca rubens Sarg, and Juniperus virginiana L. Each species showed a characteristic relationship between xylem tension and the loss of hydraulic conductivity by air embolism. Abics balsamea and Picca rubens began to embolize at tensions between 2 and 3 MPa and were completely non-conducting between 3 and 4 MPa. Juniperus virginiana was least vulnerable, beginning to embolize at 4 and still retaining approximately 10% conductivity at 10 MPa. As with a previous study of the vessel-bearing Accr saccharum Marsh., a brief perfusion of branch segments with an oxalic acid and calcium solution (10 and 0.1 mol m⁻³. respectively) increased the vulnerability of the xylem to embolism; this was especially pronounced in Abies balsamea . In order to test whether embolism was caused by aspiration of air into functional tracheids from neighbouring embolized, ones (the 'air-seeding'hypothesis), hydrated branch segments were injected with air at various pressures and measured for embolism. Results supported the air-seeding hypothesis because the relationship between injection pressure and embolism for both native and oxalic-calcium-treated segments was essentially the same as for embolism induced by xylem tension. Structural and experimental evidence suggested the air seeding occurred through inter-tracheid pit membranes when the thickened torus region of the membrane became displaced from its normal sealing position over the pit aperture. Thus, the embolism-inducing tension may be a function of pit membrane flexibility. This tension is of ecological significance because it reflects to some extent the range of xylem tensions to which a species is adapted.     

Functional and ecological xylem anatomy

Cohesion-tension transport of water is an energetically efficient way to carry large amounts of water from the roots up to the leaves. However, the cohesion-tension mechanism places the xylem water under negative hydrostatic pressure (P_x), rendering it susceptible to cavitation. There are conflicts among the structural requirements for minimizing cavitation on the one hand vs maximizing efficiency of transport and construction on the other. Cavitation by freeze-thaw events is triggered by in situ air bubble formation and is much more likely to occur as conduit diameter increases, creating a direct conflict between conducting efficiency and sensitivity to freezing induced xylem failure. Temperate ring-porous trees and vines with wide diameter conduits tend to have a shorter growing season than conifers and diffuse-porous trees with narrow conduits. Cavitation by water stress occurs by air seeding at interconduit pit membranes. Pit membrane structure is at least partially uncoupled from conduit size, leading to a much less pronounced trade-off between conducting efficiency and cavitation by drought than by freezing. Although wider conduits are generally more susceptible to drought-induced cavitation within an organ, across organs or species this trend is very weak. Different trade-offs become apparent at the level of the pit membranes that interconnect neighbouring conduits. Increasing porosity of pit membranes should enhance conductance but also make conduits more susceptible to air seeding. Increasing the size or number of pit membranes would also enhance conductance, but may weaken the strength of the conduit wall against implosion. The need to avoid conduit collapse under negative pressure creates a significant trade-off between cavitation resistance and xylem construction cost, as revealed by relationships between conduit wall strength, wood density and cavitation pressure. Trade-offs involving cavitation resistance may explain the correlations between wood anatomy, cavitation resistance, and the physiological range of negative pressure experienced by species in their native habitats.     

The effects of intervessel pit characteristics on xylem hydraulic efficiency and photosynthesis in hemiepiphytic and non‐hemiepiphytic Ficus species

Xylem vulnerability to cavitation and hydraulic efficiency are directly linked to fine‐scale bordered pit features in water‐conducting cells of vascular plants. However, it is unclear how pit characteristics influence water transport and carbon economy in tropical species. The primary aim of this study was to evaluate functional implications of changes in pit characteristics for water relations and photosynthetic traits in tropical Ficus species with different growth forms (i.e. hemiepiphytic and non‐hemiepiphytic) grown under common conditions. Intervessel pit characteristics were measured using scanning electron microscopy in five hemiepiphytic and five non‐hemiepiphytic Ficus species to determine whether these traits were related to hydraulics, leaf photosynthesis, stomatal conductance and wood density. Ficus species varied greatly in intervessel pit structure, hydraulic conductivity and leaf physiology, and clear differences were observed between the two growth forms. The area and diameter of pit aperture were negatively correlated with sapwood‐specific hydraulic conductivity, mass‐based net assimilation rate, stomatal conductance (g_s), intercellular CO₂ concentration (C_i) and the petiole vessel lumen diameters (D_v), but positively correlated with wood density. Pit morphology was only negatively correlated with sapwood‐ and leaf‐specific hydraulic conductivity and D_v. Pit density was positively correlated with g_s, C_i and D_v, but negatively with intrinsic leaf water‐use efficiency. Pit and pit aperture shape were not significantly correlated with any of the physiological traits. These findings indicate a significant role of pit characteristics in xylem water transport, carbon assimilation and ecophysiological adaptation of Ficus species in tropical rain forests.     

Inter-vessel pitting and cavitation in woody Rosaceae and other vesselled plants: a basis for a safety versus efficiency trade-off in xylem transport

The hypothesis that greater safety from cavitation by air-seeding through inter-vessel pits comes at the cost of less porous pit membranes with greater flow resistance was tested . Sixteen vessel-bearing species were compared: 11 from the Rosaceae, four from other angiosperm families, and one fern. Unexpectedly, there was no relationship between pit resistance (and hence the prevailing membrane porosity) and cavitation pressure. There was, however, an inverse relationship between pit area per vessel and vulnerability to cavitation (r² = 0.75). This suggests that cavitation is caused by the rare largest membrane pore per vessel, the average size of which increases with total pit area per vessel. If safety from cavitation constrains pit membrane surface area, it also limits vessel surface area and the minimum vessel resistivity. This trade-off was consistent with an approximately three-fold increase in vessel resistivity with cavitation pressure dropping from −0.8 to −6.6 MPa. The trade-off was compensated for by a reduction in the percentage of vessel wall pitted: from 10–16% in vulnerable species to 2–4% in resistant species. Across species, end-wall pitting accounted for 53 ± 3% of the total xylem resistivity. This corresponded to vessels achieving on average 94 ± 2% of their maximum possible conductivity if vessel surface area is constrained.     

Cavitation Resistance in Seedless Vascular Plants: The Structure and Function of Interconduit Pit Membranes

Plant water transport occurs through interconnected xylem conduits that are separated by partially digested regions in the cell wall known as pit membranes. These structures have a dual function. Their porous construction facilitates water movement between conduits while limiting the spread of air that may enter the conduits and render them dysfunctional during a drought. Pit membranes have been well studied in woody plants, but very little is known about their function in more ancient lineages such as seedless vascular plants. Here, we examine the relationships between conduit air seeding, pit hydraulic resistance, and pit anatomy in 10 species of ferns (pteridophytes) and two lycophytes. Air seeding pressures ranged from 0.8 ± 0.15 MPa (mean ± sd) in the hydric fern Athyrium filix-femina to 4.9 ± 0.94 MPa in Psilotum nudum, an epiphytic species. Notably, a positive correlation was found between conduit pit area and vulnerability to air seeding, suggesting that the rare-pit hypothesis explains air seeding in early-diverging lineages much as it does in many angiosperms. Pit area resistance was variable but averaged 54.6 MPa s m⁻¹ across all surveyed pteridophytes. End walls contributed 52% to the overall transport resistance, similar to the 56% in angiosperm vessels and 64% in conifer tracheids. Taken together, our data imply that, irrespective of phylogenetic placement, selection acted on transport efficiency in seedless vascular plants and woody plants in equal measure by compensating for shorter conduits in tracheid-bearing plants with more permeable pit membranes.Water transport in plants occurs under tension, which renders the xylem susceptible to air entry. This air seeding may lead to the rupture of water columns (cavitation) such that the air expands within conduits to create air-vapor embolisms that block further transport. (Zimmermann and Tyree, 2002). Excessive embolism such as that which occurs during a drought may jeopardize leaf hydration and lead to stomatal closure, overheating, wilting, and possibly death of the plant (Hubbard et al., 2001; Choat et al., 2012; Schymanski et al., 2013). Consequently, strong selection pressure resulted in compartmentalized and redundant plant vascular networks that are adapted to a species habitat water availability by way of life history strategy (i.e. phenology) or resistance to air seeding (Tyree et al., 1994; Mencuccini et al., 2010; Brodersen et al., 2012). The spread of drought-induced embolism is limited primarily by pit membranes, which are permeable, mesh-like regions in the primary cell wall that connect two adjacent conduits. The construction of the pit membrane is such that water easily moves across the membrane between conduits, but because of the small membrane pore size and the presence of a surface coating on the membrane (Pesacreta et al., 2005; Lee et al., 2012), the spread of air and gas bubbles is restricted up to a certain pressure threshold known as the air-seeding pressure (ASP). When xylem sap tension exceeds the air-seeding threshold, air can be aspirated from an air-filled conduit into a functional water-filled conduit through perhaps a large, preexisting pore or one that is created by tension-induced membrane stress (Rockwell et al., 2014). Air seeding leads to cavitation and embolism formation, with emboli potentially propagating throughout the xylem network (Tyree and Sperry, 1988; Brodersen et al., 2013). So, on the one hand, pit membranes are critical to controlling the spread of air throughout the vascular network, while on the other hand, they must facilitate the efficient flow of water between conduits (Choat et al., 2008; Domec et al., 2008; Pittermann et al., 2010; Schulte, 2012). Much is known about such hydraulic tradeoffs in the pit membranes of woody plants, but comparatively little data exist on seedless vascular plants such as ferns and lycophytes. Given that seedless vascular plants may bridge the evolutionary transition from bryophytes to woody plants, the lack of functional data on pit membrane structure in early-derived tracheophytes is a major gap in our understanding of the evolution of plant water transport.In woody plants, pit membranes fall into one of two categories: the torus-margo type found in most gymnosperms and the homogenous pit membrane characteristic of angiosperms (Choat et al., 2008; Choat and Pittermann, 2009). In conifers, water moves from one tracheid to another through the margo region of the membrane, with the torus sealing the pit aperture should one conduit become embolized. Air seeding occurs when water potential in the functional conduit drops low enough to dislodge the torus from its sealing position, letting air pass through the pit aperture into the water-filled tracheid (Domec et al., 2006; Delzon et al., 2010; Pittermann et al., 2010; Schulte, 2012; but see Jansen et al., 2012). Across north-temperate conifer species, larger pit apertures correlate with lower pit resistance to water flow (r_pit; MPa s m⁻¹), but it is the ratio of torus-aperture overlap that sets a species cavitation resistance (Pittermann et al., 2006, 2010; Domec et al., 2008; Hacke and Jansen, 2009). A similar though mechanistically different tradeoff exists in angiosperm pit membranes. Here, air seeding reflects a probabilistic relationship between membrane porosity and the total area of pit membranes present in the vessel walls. Specifically, the likelihood of air aspirating into a functional conduit is determined by the combination of xylem water potential and the diameter of the largest pore and/or the weakest zone in the cellulose matrix in the vessel’s array of pit membranes (Wheeler et al., 2005; Hacke et al., 2006; Christman et al., 2009; Rockwell et al., 2014). As it has come to be known, the rare-pit hypothesis suggests that the infrequent, large-diameter leaky pore giving rise to that rare pit reflects some combination of pit membrane traits such as variation in conduit membrane area (large or small), membrane properties (tight or porous), and hydrogel membrane chemistry (Hargrave et al., 1994; Choat et al., 2003; Wheeler et al., 2005; Hacke et al., 2006; Christman et al., 2009; Lee et al., 2012; Plavcová et al., 2013; Rockwell et al., 2014). The maximum pore size is critical because, per the Young-Laplace law, the larger the radius of curvature, the lower the air-water pressure difference under which the contained meniscus will fail (Jarbeau et al., 1995; Choat et al., 2003; Jansen et al., 2009). Consequently, angiosperms adapted to drier habitats may exhibit thicker, denser, smaller, and less abundant pit membranes than plants occupying regions with higher water availability (Wheeler et al., 2005; Hacke et al., 2007; Jansen et al., 2009; Lens et al., 2011; Scholz et al., 2013). However, despite these qualitative observations, there is no evidence that increased cavitation resistance arrives at the cost of higher r_pit. Indeed, the bulk of the data suggest that prevailing pit membrane porosity is decoupled from the presence of the single largest pore that allows air seeding to occur (Choat et al., 2003; Wheeler et al., 2005 Hacke et al., 2006, 2007).As water moves from one conduit to another, pit membranes offer considerable hydraulic resistance throughout the xylem network. On average, r_pit contributes 64% and 56% to transport resistance in conifers and angiosperms, respectively (Wheeler et al., 2005; Pittermann et al., 2006; Sperry et al., 2006). In conifers, the average r_pit is estimated at 6 ± 1 MPa s m⁻¹, almost 60 times lower than the 336 ± 81 MPa s m⁻¹ computed for angiosperms (Wheeler et al., 2005; Hacke et al., 2006; Sperry et al., 2006). Presumably, the high porosity of conifer pits compensates for the higher transport resistance offered by a vascular system composed of narrow, short, single-celled conduits (Pittermann et al., 2005; Sperry et al., 2006).Transport in seedless vascular plants presents an interesting conundrum because, with the exception of a handful of species, their primary xylem is composed of tracheids, the walls of which are occupied by homogenous pit membranes (Gibson et al., 1985; Carlquist and Schneider, 2001, 2007; but see Morrow and Dute, 1998, for torus-margo membranes in Botrychium spp.). At first pass, this combination of traits appears hydraulically maladaptive, but several studies have shown that ferns can exhibit transport capacities that are on par with more recently evolved plants (Wheeler et al., 2005; Watkins et al., 2010; Pittermann et al., 2011, 2013; Brodersen et al., 2012). Certainly, several taxa possess large-diameter, highly overlapping conduits, some even have vessels such as Pteridium aquilinum and many species have high conduit density, all of which could contribute to increased hydraulic efficiency (Wheeler et al., 2005; Pittermann et al., 2011, 2013). But how do the pit membranes of seedless vascular plants compare? Scanning electron micrographs of fern and lycopod xylem conduits suggest that they are thin, diaphanous, and susceptible to damage during specimen preparation (Carlquist and Schneider 2001, 2007). Consistent with such observations, two estimates of r_pit imply that r_pit in ferns may be significantly lower than in angiosperms; Wheeler et al. (2005) calculated r_pit in the fern Pteridium aquilinum at 31 MPa s m⁻¹, while Schulte et al. (1987) estimated r_pit at 1.99 MPa s m⁻¹ in the basal fern Psilotum nudum. The closest structural analogy to seedless vascular plant tracheids can be found in the secondary xylem of the early-derived vesselless angiosperms, in which tracheids possess homogenous pit membranes with r_pit values that at 16 MPa s m⁻¹ are marginally higher than those of conifers (Hacke et al., 2007). Given that xylem in seedless vascular plants is functionally similar to that in vesselless angiosperms, we expected convergent r_pit values in these two groups despite their phylogenetic distance. We tested this hypothesis, as well as the intrinsic cavitation resistance of conduits in seedless vascular plants, by scrutinizing the pit membranes of ferns and fern allies using the anatomical and experimental approaches applied previously to woody taxa. In particular, we focused on the relationship between pit membrane traits and cavitation resistance at the level of the individual conduit.     

Direct observation and modelling of embolism spread between xylem conduits: a case study in Scots pine

Xylem embolism is one of the main processes involved in drought‐related plant mortality. Although its consequences for plant physiology are already well described, embolism formation and spread are poorly evaluated and modelled, especially for tracheid‐based species. The aim of this study was to assess the embolism formation and spread in Pinus sylvestris as a case study using X‐ray microtomography and hydraulics methods. We also evaluated the potential effects of cavitation fatigue on vulnerability to embolism and the micro‐morphology of the bordered pits using scanning electron microscopy (SEM) to test for possible links between xylem anatomy and embolism spread. Finally, a novel model was developed to simulate the spread of embolism in a 2D anisotropic cellular structure. Results showed a large variability in the formation and spread of embolism within a ring despite no differences being observed in intertracheid pit membrane anatomical traits. Simulations from the model showed a highly anisotropic tracheid‐to‐tracheid embolism spreading pattern, which confirms the major role of tracheid‐to‐tracheid air seeding to explain how embolism spreads in Scots pine. The results also showed that prior embolism removal from the samples reduced the resistance to embolism of the xylem and could result in overestimates of vulnerability to embolism.     

Structural determinants of increased susceptibility to dehydration‐induced cavitation in post‐fire resprouting chaparral shrubs

It is well established that transpiration and photosynthetic rates generally increase in resprouting shoots after fire in chaparral shrublands. By contrast, little is known about how plant hydraulic function varies during this same recovery period. We hypothesized that vascular traits, both functional and structural, would also shift in order to support this heightened level of gas exchange and growth. We examined stem xylem‐specific hydraulic conductivity (K_s) and resistance to cavitation (P₅₀) for eight chaparral shrub species as well as several potential xylem structural determinants of hydraulic function and compared established unburned plants and co‐occurring post‐fire resprouting plants. Unburned plants were generally more resistant to cavitation than resprouting plants, but the two groups did not differ in K_s. Resprouting plants had altered vessel structure compared with unburned plants, with resprouting plants having both wider diameter vessels and higher inter‐vessel pit density. For biomechanics, unburned plants had both stronger and denser stem xylem tissue than resprouting plants. Shifts in hydraulic structure and function resulted in resprouting plants being more vulnerable to dehydration. The interaction between time since disturbance (i.e. resprouting versus established stands) and drought may complicate attempts to predict mortality risk of resprouting plants.     

Measurement of vulnerability to water stress‐induced cavitation in grapevine: a comparison of four techniques applied to a long‐vesseled species

Among woody plants, grapevines are often described as highly vulnerable to water‐stress induced cavitation with emboli forming at slight tensions. However, we found native embolism never exceeded 30% despite low xylem water potentials (Ψ_x) for stems of field grown vines. The discrepancy between native embolism measurements and those of previous reports led us to assess vulnerability curve generation using four separate methods and alterations (i.e. segment length and with/without flushing to remove embolism prior to measurement) of each. Centrifuge, dehydration and air‐injection methods, which rely on measurement of percentage loss of hydraulic conductivity (PLC) in detached stems, were compared against non‐invasive monitoring of xylem cavitation with nuclear magnetic resonance (NMR) imaging. Short segment air‐injection and flushed centrifuge stems reached >90 PLC at Ψ_x of‐0.5 and ?1.5 MPa, respectively, whereas dehydration and long‐segment air‐injection measurements indicated no significant embolism at Ψ_x > ?2.0 MPa. Observations from NMR agreed with the dehydration and long segment air‐injection methods, showing the majority of vessels were still water‐filled at Ψ_x > ?1.5 MPa. Our findings show V. vinifera stems are far less vulnerable to water stress‐induced cavitation than previously reported, and dehydration and long segment air‐injection techniques are more appropriate for long‐vesseled species and organs.     

Poplar vulnerability to xylem cavitation acclimates to drier soil conditions

Xylem vulnerability to cavitation differs between tree species according to their drought resistance, more xerophilous species being more resistant to xylem cavitation. Variability in xylem vulnerability to cavitation is also found within species, especially between in situ populations. The origin of this variability has not been clearly identified. Here we analyzed the response of xylem hydraulic traits of Populus tremula×Populus alba trees to three different soil water regimes. Stem xylem vulnerability was scored as the xylem water potential causing 12, 50 and 88% loss of conductivity (P₁₂, P₅₀ and P₈₈). Vulnerability to cavitation was found to acclimate to growing conditions under different levels of soil water content, with P₅₀ values of ?1.82, ?2.03 and ?2.45 MPa in well‐watered, moderately water‐stressed and severely water‐stressed poplars, respectively. The value of P₁₂, the xylem tension at which cavitation begins, was correlated with the lowest value of midday leaf water potential (ψm) experienced by each plant, the difference between the two parameters being approximately 0.5 MPa, consistent with the absence of any difference in embolism level between the different water treatments. These results support the hypothesis that vulnerability to cavitation is a critical trait for resistance to drought. The decrease in vulnerability to cavitation under growing conditions of soil drought was correlated with decreased vessel diameter, increased vessel wall thickness and a stronger bordered pit field (t/b)². The links between these parameters are discussed.     

Vulnerability to drought‐induced cavitation in poplars: synthesis and future opportunities

Vulnerability to drought‐induced cavitation is a key trait of plant water relations. Here, we summarize the available literature on vulnerability to drought‐induced cavitation in poplars (Populus spp.), a genus of agronomic, ecological and scientific importance. Vulnerability curves and vulnerability parameters (including the water potential inducing 50% loss in hydraulic conductivity, P₅₀) were collected from 37 studies published between 1991 and 2014, covering a range of 10 species and 12 interspecific hybrid crosses. Results of our meta‐analysis confirm that poplars are among the most vulnerable woody species to drought‐induced cavitation (mean P₅₀ = ?1.44 and ?1.55 MPa across pure species and hybrids, respectively). Yet, significant variation occurs among species (P₅₀ range: 1.43 MPa) and among hybrid crosses (P₅₀ range: 1.12 MPa), within species and hybrid crosses (max. P₅₀ range reported: 0.8 MPa) as well as in response to environmental factors including nitrogen fertilization, irradiance, temperature and drought (max. P₅₀ range reported: 0.75 MPa). Potential implications and gaps in knowledge are discussed in the context of poplar cultivation, species adaptation and climate modifications. We suggest that poplars represent a valuable model for studies on drought‐induced cavitation, especially to elucidate the genetic and molecular basis of cavitation resistance in Angiosperms.     

Modelling the mechanical behaviour of pit membranes in bordered pits with respect to cavitation resistance in angiosperms

Background and Aims

Various correlations have been identified between anatomical features of bordered pits in angiosperm xylem and vulnerability to cavitation, suggesting that the mechanical behaviour of the pits may play a role. Theoretical modelling of the membrane behaviour has been undertaken, but it requires input of parameters at the nanoscale level. However, to date, no experimental data have indicated clearly that pit membranes experience strain at high levels during cavitation events.

Methods

Transmission electron microscopy (TEM) was used in order to quantify the pit micromorphology of four tree species that show contrasting differences in vulnerability to cavitation, namely Sorbus aria, Carpinus betulus, Fagus sylvatica and Populus tremula. This allowed anatomical characters to be included in a mechanical model that was based on the Kirchhoff–Love thin plate theory. A mechanistic model was developed that included the geometric features of the pits that could be measured, with the purpose of evaluating the pit membrane strain that results from a pressure difference being applied across the membrane. This approach allowed an assessment to be made of the impact of the geometry of a pit on its mechanical behaviour, and provided an estimate of the impact on air-seeding resistance.

Key Results

The TEM observations showed evidence of residual strains on the pit membranes, thus demonstrating that this membrane may experience a large degree of strain during cavitation. The mechanical modelling revealed the interspecific variability of the strains experienced by the pit membrane, which varied according to the pit geometry and the pressure experienced. The modelling output combined with the TEM observations suggests that cavitation occurs after the pit membrane has been deflected against the pit border. Interspecific variability of the strains experienced was correlated with vulnerability to cavitation. Assuming that air-seeding occurs at a given pit membrane strain, the pressure predicted by the model to achieve this mechanical state corresponds to experimental values of cavitation sensitivity (P₅₀).

Conclusions

The results provide a functional understanding of the importance of pit geometry and pit membrane structure in air-seeding, and thus in vulnerability to cavitation.     

What causes the differences in cavitation resistance of two shrubs? Wood anatomical explanations and reliability testing of vulnerability curves

Relationships between xylem anatomical traits and cavitation resistance have always been a major content of plant hydraulics. To know how plants cope with drought, it is extremely important to acquire detailed knowledge about xylem anatomical traits and assess the cavitation resistance accurately. This study aims to increase our knowledge in the methods determining cavitation resistance and xylem anatomical traits. We selected a semi-ring-porous species, Hippophae rhamnoides L., and a diffuse-porous species, Corylus heterophylla F., to clarify the reasons for the difference in cavitation resistance based on detailed xylem anatomical traits and reliable vulnerability curves (VCs). Both Cavitron and bench dehydration (BD) were used to construct VCs. Xylem anatomical traits, including pit membrane ultrastructure of these two species, were determined. The VCs obtained by the two different techniques were of different types for H. rhamnoides, its Cavitron VCs might be unreliable because of open-vessel artifacts. On the basis of BD VCs, H. rhamnoides showed higher cavitation resistance than C. heterophylla, and this is attributed to its low vessel connectivity as well as non-porous and thicker pit membranes.     

Moving beyond the cambium necrosis hypothesis of post-fire tree mortality: cavitation and deformation of xylem in forest fires

? It is widely assumed that post-fire tree mortality results from necrosis of phloem and vascular cambium in stems, despite strong evidence that reduced xylem conductivity also plays an important role. ? In this study, experiments with Populus balsamifera were used to demonstrate two mechanisms by which heat reduces the hydraulic conductivity of xylem: air seed cavitation and conduit wall deformation. Heat effects on air seed cavitation were quantified using air injection experiments that isolate potential temperature-dependent changes in sap surface tension and pit membrane pore diameters. Heat effects on conduit wall structure were demonstrated using air conductivity measurements and light microscopy. ? Heating increased vulnerability to cavitation because sap surface tension varies inversely with temperature. Heating did not affect cavitation via changes in pit membrane pore diameters, but did cause significant reductions in xylem air conductivity that were associated with deformation of conduit walls (probably resulting from thermal softening of viscoelastic cell wall polymers). ? Additional work is required to understand the relative roles of cavitation and deformation in the reduction of xylem conductivity, and how reduced xylem conductivity in roots, stems, and branches correlates and interacts with foliage and root necroses to cause tree mortality. Future research should also examine how heat necrosis of ray parenchyma cells affects refilling of embolisms that occur during and after the fire event.