Living on the edge: A continental-scale assessment of forest vulnerability to drought.

Globally, forests are facing an increasing risk of mass tree mortality events associated with extreme droughts and higher temperatures. Hydraulic dysfunction is considered a key mechanism of drought-triggered dieback. By leveraging the climate breadth of the Australian landscape and a national network of research sites (Terrestrial Ecosystem Research Network), we conducted a continental-scale study of physiological and hydraulic traits of 33 native tree species from contrasting environments to disentangle the complexities of plant response to drought across communities. We found strong relationships between key plant hydraulic traits and site aridity. Leaf turgor loss point and xylem embolism resistance were correlated with minimum water potential experienced by each species. Across the data set, there was a strong coordination between hydraulic traits, including those linked to hydraulic safety, stomatal regulation and the cost of carbon investment into woody tissue. These results illustrate that aridity has acted as a strong selective pressure, shaping hydraulic traits of tree species across the Australian landscape. Hydraulic safety margins were constrained across sites, with species from wetter sites tending to have smaller safety margin compared with species at drier sites, suggesting trees are operating close to their hydraulic thresholds and forest biomes across the spectrum may be susceptible to shifts in climate that result in the intensification of drought.

Non-invasive imaging shows no evidence of embolism repair after drought in tree species of two genera.

Drought stress can result in significant impairment of the plant hydraulic system via blockage of xylem conduits by gas emboli. Recovery after drought stress is an essential component of plant survival but is still a poorly understood process. In this study, we examined the capacity of woody species from two genera (Eucalyptus and Quercus) to refill embolized xylem vessels during a cycle of drought and recovery. Observations were made on intact plants of Eucalyptus calmudulensis, E. grandis, E. saligna and Quercus palustris using X-ray microtomography. We found no evidence of an effective xylem refilling mechanism in any of the plant species. Despite rehydration and recovery of plant water potential to near pre-drought levels, embolized vessels were not refilled up to 72 h after rewatering. In E. saligna, water droplets accumulated in previously air-filled vessels for a very small percentage of vessels. However, no instances of complete refilling that would restore embolized vessels to hydraulic function were observed. Our observations suggest that rapid refilling of embolized vessels after drought may not be a wide spread mechanism in woody plants and that embolism formed during drought represents long term cost to the plant hydraulic system.

Mitigating the open vessel artefact in centrifuge-based measurement of embolism resistance.

Centrifuge-based techniques to assess xylem vulnerability to embolism are increasingly being used, although we are yet to reach a consensus on the nature and extent of artefactual embolism observed in some angiosperm species. In particular, there is disagreement over whether these artefacts influence both the spin (Cavitron) and static versions of the centrifuge technique equally. We tested two methods for inducing embolism: bench dehydration and centrifugation. We used three methods to measure the resulting loss of conductivity: gravimetric flow measured in bench-dehydrated and centrifuged samples (static centrifuge), in situ flow measured under tension during spinning in the centrifuge (Cavitron) and direct imaging using X-ray computed microtomography (microCT) observations in stems of two species of Hakea that differ in vessel length. Both centrifuge techniques were prone to artefactual embolism in samples with maximum vessel length longer than, or similar to, the centrifuge rotor diameter. Observations with microCT indicated that this artefactual embolism occurred in the outermost portions of samples. The artefact was largely eliminated if flow was measured in an excised central part of the segment in the static centrifuge or starting measurements with the Cavitron at pressures lower than the threshold of embolism formation in open vessels. The simulations of loss of conductivity in centrifuged samples with a new model, CAVITOPEN, confirmed that the impact of open vessels on the vulnerability to embolism curve was higher when vessels were long, samples short and when embolism is formed in open vessels at less negative pressures. This model also offers a robust and quantitative tool to test and correct for artefactual embolism at low xylem tensions.

Is xylem of angiosperm leaves less resistant to embolism than branches? Insights from microCT, hydraulics, and anatomy.

According to the hydraulic vulnerability segmentation hypothesis, leaves are more vulnerable to decline of hydraulic conductivity than branches, but whether stem xylem is more embolism resistant than leaves remains unclear. Drought-induced embolism resistance of leaf xylem was investigated based on X-ray microcomputed tomography (microCT) for Betula pendula, Laurus nobilis, and Liriodendron tulipifera, excluding outside-xylem, and compared with hydraulic vulnerability curves for branch xylem. Moreover, bordered pit characters related to embolism resistance were investigated for both organs. Theoretical P50 values (i.e. the xylem pressure corresponding to 50% loss of hydraulic conductance) of leaves were generally within the same range as hydraulic P50 values of branches. P50 values of leaves were similar to branches for L. tulipifera (-2.01 versus -2.10 MPa, respectively), more negative for B. pendula (-2.87 versus -1.80 MPa), and less negative for L. nobilis (-6.4 versus -9.2 MPa). Despite more narrow conduits in leaves than branches, mean interconduit pit membrane thickness was similar in both organs, but significantly higher in leaves of B. pendula than in branches. This case study indicates that xylem shows a largely similar embolism resistance across leaves and branches, although differences both within and across organs may occur, suggesting interspecific variation with regard to the hydraulic vulnerability segmentation hypothesis.

Monitoring of Freezing Dynamics in Trees: A Simple Phase Shift Causes Complexity.

During winter, trees have to cope with harsh conditions, including extreme freeze-thaw stress. This study focused on ice nucleation and propagation, related water shifts and xylem cavitation, as well as cell damage and was based on in situ monitoring of xylem (thermocouples) and surface temperatures (infrared imaging), ultrasonic emissions, and dendrometer analysis. Field experiments during late winter on Picea abies growing at the alpine timberline revealed three distinct freezing patterns: (1) from the top of the tree toward the base, (2) from thin branches toward the main stem's top and base, and (3) from the base toward the top. Infrared imaging showed freezing within branches from their base toward distal parts. Such complex freezing causes dynamic and heterogenous patterns in water potential and probably in cavitation. This study highlights the interaction between environmental conditions upon freezing and thawing and demonstrates the enormous complexity of freezing processes in trees. Diameter shrinkage, which indicated water fluxes within the stem, and acoustic emission analysis, which indicated cavitation events near the ice front upon freezing, were both related to minimum temperature and, upon thawing, related to vapor pressure deficit and soil temperature. These complex patterns, emphasizing the common mechanisms between frost and drought stress, shed new light on winter tree physiology.

Visualization of xylem embolism by X-ray microtomography: a direct test against hydraulic measurements.

X-ray microtomography (microCT) is becoming a valuable noninvasive tool for advancing our understanding of plant-water relations. Laboratory-based microCT systems are becoming more affordable and provide better access than synchrotron facilities. However, some systems come at the cost of comparably lower signal quality and spatial resolution than synchrotron facilities. In this study, we evaluated laboratory-based X-ray microCT imaging as a tool to nondestructively analyse hydraulic vulnerability to drought-induced embolism in a woody plant species. We analysed the vulnerability to drought-induced embolism of benchtop-dehydrated Eucalyptus camaldulensis plants using microCT and hydraulic flow measurements on the same sample material, allowing us to directly compare the two methods. Additionally, we developed a quantitative procedure to improve microCT image analysis at limited resolution and accurately measure vessel lumens. Hydraulic measurements matched closely with microCT imaging of the current-year growth ring, with similar hydraulic conductivity and loss of conductivity due to xylem embolism. Optimized thresholding of vessel lumens during image analysis, based on a physiologically meaningful parameter (theoretical conductivity), allowed us to overcome common potential constraints of some lab-based systems. Our results indicate that estimates of vulnerability to embolism provided by microCT visualization agree well with those obtained from hydraulic measurements on the same sample material.

Herb Hydraulics: Inter- and Intraspecific Variation in Three Ranunculus Species.

The requirements of the water transport system of small herbaceous species differ considerably from those of woody species. Despite their ecological importance for many biomes, knowledge regarding herb hydraulics remains very limited. We compared key hydraulic features (vulnerability to drought-induced hydraulic decline, pressure-volume relations, onset of cellular damage, in situ variation of water potential, and stomatal conductance) of three Ranunculus species differing in their soil humidity preferences and ecological amplitude. All species were very vulnerable to water stress (50% reduction in whole-leaf hydraulic conductance [kleaf] at -0.2 to -0.8 MPa). In species with narrow ecological amplitude, the drought-exposed Ranunculus bulbosus was less vulnerable to desiccation (analyzed via loss of kleaf and turgor loss point) than the humid-habitat Ranunculus lanuginosus Accordingly, water stress-exposed plants from the broad-amplitude Ranunculus acris revealed tendencies toward lower vulnerability to water stress (e.g. osmotic potential at full turgor, cell damage, and stomatal closure) than conspecific plants from the humid site. We show that small herbs can adjust to their habitat conditions on interspecific and intraspecific levels in various hydraulic parameters. The coordination of hydraulic thresholds (50% and 88% loss of kleaf, turgor loss point, and minimum in situ water potential) enabled the study species to avoid hydraulic failure and damage to living cells. Reversible recovery of hydraulic conductance, desiccation-tolerant seeds, or rhizomes may allow them to prioritize toward a more efficient but vulnerable water transport system while avoiding the severe effects that water stress poses on woody species.

Elevational trends in hydraulic efficiency and safety of Pinus cembra roots.

In alpine regions, elevational gradients in environmental parameters are reflected by structural and functional changes in plant traits. Elevational changes in plant water relations have also been demonstrated, but comparable information on root hydraulics is generally lacking. We analyzed the hydraulic efficiency (specific hydraulic conductivity k s, entire root system conductance K R) and vulnerability to drought-induced embolism (water potential at 50 % loss of conductivity Ψ 50) of the roots of Pinus cembra trees growing along an elevational transect of 600 m. Hydraulic parameters of the roots were compared with those of the stem and related to anatomical traits {mean conduit diameter (d), wall reinforcement [(t/b)(2)]}. We hypothesized that temperature-related restrictions in root function would cause a progressive limitation of hydraulic efficiency and safety with increasing elevation. We found that both root k s and K R decreased from low (1600 m a.s.l.: k s 5.6 ± 0.7 kg m(-1) s(-1) MPa(-1), K R 0.049 ± 0.005 kg m(-2) s (-1) MPa(-1)) to high elevation (2100 m a.s.l.: k s 4.2 ± 0.6 kg m(-1) s(-1) MPa(-1), K R 0.035 ± 0.006 kg m(-2) s(-1) MPa(-1)), with small trees showing higher K R than large trees. k s was higher in roots than in stems (0.5 ± 0.05 kg m(-1)s(-1)MPa(-1)). Ψ 50 values were similar across elevations and overall less negative in roots (Ψ 50 -3.6 ± 0.1 MPa) than in stems (Ψ 50 -3.9 ± 0.1 MPa). In roots, large-diameter tracheids were lacking at high elevation and (t/b)(2) increased, while d did not change. The elevational decrease in root hydraulic efficiency reflects a limitation in timberline tree hydraulics. In contrast, hydraulic safety was similar across elevations, indicating that avoidance of hydraulic failure is important for timberline trees. As hydraulic patterns can only partly be explained by the anatomical parameters studied, limitations and/or adaptations at the pit level are likely.

Stem and leaf hydraulic properties are finely coordinated in three tropical rain forest tree species.

Coordination of stem and leaf hydraulic traits allows terrestrial plants to maintain safe water status under limited water supply. Tropical rain forests, one of the world's most productive biomes, are vulnerable to drought and potentially threatened by increased aridity due to global climate change. However, the relationship of stem and leaf traits within the plant hydraulic continuum remains understudied, particularly in tropical species. We studied within-plant hydraulic coordination between stems and leaves in three tropical lowland rain forest tree species by analyses of hydraulic vulnerability [hydraulic methods and ultrasonic emission (UE) analysis], pressure-volume relations and in situ pre-dawn and midday water potentials (Ψ). We found finely coordinated stem and leaf hydraulic features, with a strategy of sacrificing leaves in favour of stems. Fifty percent of hydraulic conductivity (P50 ) was lost at -2.1 to -3.1 MPa in stems and at -1.7 to -2.2 MPa in leaves. UE analysis corresponded to hydraulic measurements. Safety margins (leaf P50 - stem P50 ) were very narrow at -0.4 to -1.4 MPa. Pressure-volume analysis and in situ Ψ indicated safe water status in stems but risk of hydraulic failure in leaves. Our study shows that stem and leaf hydraulics were finely tuned to avoid embolism formation in the xylem.

Xylem cavitation resistance can be estimated based on time-dependent rate of acoustic emissions.

Acoustic emission (AE) analysis allows nondestructive monitoring of embolism formation in plant xylem, but signal interpretation and agreement of acoustically measured hydraulic vulnerability with reference hydraulic techniques remain under debate. We compared the hydraulic vulnerability of 16 species and three crop tree cultivars using hydraulic flow measurements and acoustic emission monitoring, proposing the use of time-dependent AE rates as a novel parameter for AE analysis. There was a linear correlation between the water potential (Ψ) at 50% loss of hydraulic conductivity (P50 ) and the Ψ at maximum AE activity (Pmaxrate ), where species with lower P50 also had lower Pmaxrate (P < 0.001, R(2)  = 0.76). Using AE rates instead of cumulative counts for AE analysis allows more efficient estimation of P50 , while excluding problematic AE at late stages of dehydration.

Physiological acclimation to drought stress in Solidago canadensis.

Solidago canadensis is an invasive species from North America that is spreading across Europe, Australia and temperate Asia. We hypothesized that the species' wide ecological amplitude is also based on its potential in hydraulic acclimation, and analyzed hydraulic and anatomical properties along a transect with decreasing soil humidity. Stem hydraulic conductivity, vulnerability to drought-induced embolism, stomatal closure during dehydration and xylem-anatomical parameters were quantified at three sites. At the humid site, specific hydraulic conductivity of stems (1.0 ± 0.2 kg m(-1) MPa(-1) s(-1)) was about twofold higher, and leaf-specific conductivity about 1.5 times higher (3.1 ± 0.5 kg m(-1) MPa(-1) s(-1)) than at the dry site. Water potential (Ψ) at 50% loss of conductivity was -3.7 ± 0.1 MPa at the dry site and -3.1 ± 0.2 MPa at the humid site (September). Vulnerability to drought-induced embolism decreased along the transect and over the vegetation period. At drier sites, stomata started closing at lower Ψ while complete stomatal closure was reached at less negative Ψ (12% of maximum stomatal conductance: -2.5 ± 0.0 and -3.0 ± 0.2 MPa at the dry and humid site). The safety margin between stomatal closure and 50% loss of conductivity was 1.2 and 0.2 MPa at the dry and humid sites. The observed variability indicated an efficient acclimation in hydraulic conductivity and safety: plants at dry sites exhibited lower specific hydraulic conductivity, higher embolism resistance and broader safety margins, signifying a trade-off between the hydraulic safety and efficiency. The observed intraspecific plasticity in hydraulic and anatomical traits may help to explain the invasive potential of this species.