Evidence for within-species transition between drought response strategies in Nicotiana benthamiana.

Nicotiana benthamiana is predominantly distributed in arid habitats across northern Australia. However, none of six geographically isolated accessions shows obvious xerophytic morphological features. To investigate how these tender-looking plants withstand drought, we examined their responses to water deprivation, assessed phenotypic, physiological, and cellular responses, and analysed cuticular wax composition and wax biosynthesis gene expression profiles. Results showed that the Central Australia (CA) accession, globally known as a research tool, has evolved a drought escape strategy with early vigour, short life cycle, and weak, water loss-limiting responses. By contrast, a northern Queensland (NQ) accession responded to drought by slowing growth, inhibiting flowering, increasing leaf cuticle thickness, and altering cuticular wax composition. Under water stress, NQ increased the heat stability and water impermeability of its cuticle by extending the carbon backbone of cuticular long-chain alkanes from c. 25 to 33. This correlated with rapid upregulation of at least five wax biosynthesis genes. In CA, the alkane chain lengths (c. 25) and gene expression profiles remained largely unaltered. This study highlights complex genetic and environmental control over cuticle composition and provides evidence for divergence into at least two fundamentally different drought response strategies within the N. benthamiana species in < 1 million years.

Aridity drives clinal patterns in leaf traits and responsiveness to precipitation in a broadly distributed Australian tree species.

Aridity shapes species distributions and plant growth and function worldwide. Yet, plant traits often show complex relationships with aridity, challenging our understanding of aridity as a driver of evolutionary adaptation. We grew nine genotypes of Eucalyptus camaldulensis subsp. camaldulensis sourced from an aridity gradient together in the field for ~650 days under low and high precipitation treatments. Eucalyptus camaldulesis is considered a phreatophyte (deep-rooted species that utilizes groundwater), so we hypothesized that genotypes from more arid environments would show lower aboveground productivity, higher leaf gas-exchange rates, and greater tolerance/avoidance of dry surface soils (indicated by lower responsiveness) than genotypes from less arid environments. Aridity predicted genotype responses to precipitation, with more arid genotypes showing lower responsiveness to reduced precipitation and dry surface conditions than less arid genotypes. Under low precipitation, genotype net photosynthesis and stomatal conductance increased with home-climate aridity. Across treatments, genotype intrinsic water-use efficiency and osmotic potential declined with increasing aridity while photosynthetic capacity (Rubisco carboxylation and RuBP regeneration) increased with aridity. The observed clinal patterns indicate that E. camaldulensis genotypes from extremely arid environments possess a unique strategy defined by lower responsiveness to dry surface soils, low water-use efficiency, and high photosynthetic capacity. This strategy could be underpinned by deep rooting and could be adaptive under arid conditions where heat avoidance is critical and water demand is high.

Key hydraulic traits control the dynamics of plant dehydration in four contrasting tree species during drought.

Trees are at risk of mortality during extreme drought, yet our understanding of the traits that govern the timing of drought-induced hydraulic failure remains limited. To address this, we tested SurEau, a trait-based soil-plant-atmosphere model designed to predict the dynamics of plant dehydration as represented by the changes in water potential against those observed in potted trees of four contrasting species (Pinus halepensis Mill., Populus nigra L., Quercus ilex L. and Cedrus atlantica (Endl.) Manetti ex Carriére) exposed to drought. SurEau was parameterized with a range of plant hydraulic and allometric traits, soil and climatic variables. We found a close correspondence between the predicted and observed plant water potential (in MPa) dynamics during the early phase drought, leading to stomatal closure, as well as during the latter phase of drought, leading to hydraulic failure in all four species. A global model's sensitivity analysis revealed that, for a common plant size (leaf area) and soil volume, dehydration time from full hydration to stomatal closure (Tclose) was most strongly controlled by the leaf osmotic potential (Pi0) and its influence on stomatal closure, in all four species, while the maximum stomatal conductance (gsmax) also contributed to Tclose in Q. ilex and C. atlantica. Dehydration times from stomatal closure to hydraulic failure (Tcav) was most strongly controlled by Pi0, the branch residual conductance (gres) and Q10a sensitivity of gres in the three evergreen species, while xylem embolism resistance (P50) was most influential in the deciduous species P. nigra. Our findings point to SurEau as a highly useful model for predicting changes in plant water status during drought and suggest that adjustments made in key hydraulic traits are potentially beneficial to delaying the onset of drought-induced hydraulic failure in trees.

Cold temperature and aridity shape the evolution of drought tolerance traits in Tasmanian species of Eucalyptus.

Perennial plant species from water-limiting environments (including climates of extreme drought, heat and freezing temperatures) have evolved traits that allow them to tolerate these conditions. As such, traits that are associated with water stress may show evidence of adaptation to climate when compared among closely related species inhabiting contrasting climatic conditions. In this study, we tested whether key hydraulic traits linked to drought stress, including the vulnerability of leaves to embolism (P50 leaf) and the minimum diffusive conductance of shoots (gmin), were associated with climatic characteristics of 14 Tasmanian eucalypt species from sites that vary in precipitation and temperature. Across species, greater cavitation resistance (more negative P50 leaf) was associated with increasing aridity and decreasing minimum temperature. By contrast, gmin showed strong associations with aridity only. Among these Tasmanian eucalypts, evidence suggests that trait variation is influenced by both cold and dry conditions, highlighting the need to consider both aspects when exploring adaptive trait-climate relationships.

Plant hydraulic modelling of leaf and canopy fuel moisture content reveals increasing vulnerability of a Mediterranean forest to wildfires under extreme drought.

Fuel moisture content (FMC) is a crucial driver of forest fires in many regions world-wide. Yet, the dynamics of FMC in forest canopies as well as their physiological and environmental determinants remain poorly understood, especially under extreme drought. We embedded a FMC module in the trait-based, plant-hydraulic SurEau-Ecos model to provide innovative process-based predictions of leaf live fuel moisture content (LFMC) and canopy fuel moisture content (CFMC) based on leaf water potential ( ψ Leaf ). SurEau-Ecos-FMC relies on pressure-volume (p-v) curves to simulate LFMC and vulnerability curves to cavitation to simulate foliage mortality. SurEau-Ecos-FMC accurately reproduced ψ Leaf and LFMC dynamics as well as the occurrence of foliage mortality in a Mediterranean Quercus ilex forest. Several traits related to water use (leaf area index, available soil water, and transpiration regulation), vulnerability to cavitation, and p-v curves (full turgor osmotic potential) had the greatest influence on LFMC and CFMC dynamics. As the climate gets drier, our results showed that drought-induced foliage mortality is expected to increase, thereby significantly decreasing CFMC. Our results represent an important advance in our capacity to understand and predict the sensitivity of forests to wildfires.

AusTraits, a curated plant trait database for the Australian flora.

We introduce the AusTraits database - a compilation of values of plant traits for taxa in the Australian flora (hereafter AusTraits). AusTraits synthesises data on 448 traits across 28,640 taxa from field campaigns, published literature, taxonomic monographs, and individual taxon descriptions. Traits vary in scope from physiological measures of performance (e.g. photosynthetic gas exchange, water-use efficiency) to morphological attributes (e.g. leaf area, seed mass, plant height) which link to aspects of ecological variation. AusTraits contains curated and harmonised individual- and species-level measurements coupled to, where available, contextual information on site properties and experimental conditions. This article provides information on version 3.0.2 of AusTraits which contains data for 997,808 trait-by-taxon combinations. We envision AusTraits as an ongoing collaborative initiative for easily archiving and sharing trait data, which also provides a template for other national or regional initiatives globally to fill persistent gaps in trait knowledge.

Acclimation of hydraulic and morphological traits to water deficit delays hydraulic failure during simulated drought in poplar.

The capacity of trees to tolerate and survive increasing drought conditions in situ will depend in part on their ability to acclimate (via phenotypic plasticity) key hydraulic and morphological traits that increase drought tolerance and delay the onset of drought-induced hydraulic failure. However, the effect of water-deficit acclimation in key traits that determine time to hydraulic failure (THF) during extreme drought remains largely untested. We measured key hydraulic and morphological traits in saplings of a hybrid poplar grown under well-watered and water-limited conditions. The time for plants to dry-down to critical levels of water stress (90% loss of stem hydraulic conductance), as well as the relative contribution of drought acclimation in each trait to THF, was simulated using a soil-plant hydraulic model (SurEau). Compared with controls, water-limited plants exhibited significantly lower stem hydraulic vulnerability (P50stem), stomatal conductance and total canopy leaf area (LA). Taken together, adjustments in these and other traits resulted in longer modelled THF in water-limited (~160 h) compared with well-watered plants (~50 h), representing an increase of more than 200%. Sensitivity analysis revealed that adjustment in P50stem and LA contributed the most to longer THF in water-limited plants. We observed a high degree of trait plasticity in poplar saplings in response to water-deficit growth conditions, with decreases in stem hydraulic vulnerability and leaf area playing a key role in delaying the onset of hydraulic failure during a simulated drought event. These findings suggest that understanding the capacity of plants to acclimate to antecedent growth conditions will enable better predictions of plant survivorship during future drought.

Identifying areas at risk of drought-induced tree mortality across South-Eastern Australia.

South-East Australia has recently been subjected to two of the worst droughts in the historical record (Millennium Drought, 2000-2009 and Big Dry, 2017-2019). Unfortunately, a lack of forest monitoring has made it difficult to determine whether widespread tree mortality has resulted from these droughts. Anecdotal observations suggest the Big Dry may have led to more significant tree mortality than the Millennium drought. Critically, to be able to robustly project future expected climate change effects on Australian vegetation, we need to assess the vulnerability of Australian trees to drought. Here we implemented a model of plant hydraulics into the Community Atmosphere Biosphere Land Exchange (CABLE) land surface model. We parameterized the drought response behaviour of five broad vegetation types, based on a common garden dry-down experiment with species originating across a rainfall gradient (188-1,125 mm/year) across South-East Australia. The new hydraulics model significantly improved (~35%-45% reduction in root mean square error) CABLE's previous predictions of latent heat fluxes during periods of water stress at two eddy covariance sites in Australia. Landscape-scale predictions of the greatest percentage loss of hydraulic conductivity (PLC) of about 40%-60%, were broadly consistent with satellite estimates of regions of the greatest change in both droughts. In neither drought did CABLE predict that trees would have reached critical PLC in widespread areas (i.e. it projected a low mortality risk), although the model highlighted critical levels near the desert regions of South-East Australia where few trees live. Overall, our experimentally constrained model results imply significant resilience to drought conferred by hydraulic function, but also highlight critical data and scientific gaps. Our approach presents a promising avenue to integrate experimental data and make regional-scale predictions of potential drought-induced hydraulic failure.

The DroughtBox: A new tool for phenotyping residual branch conductance and its temperature dependence during drought.

Xylem hydraulic failure is a major driver of tree death during drought. However, to better understand mortality risk in trees, especially during hot-drought events, more information is required on both rates of residual water-loss from small branches (gres ) after stomatal closure, as well as the phase transition temperature (Tp ), beyond which gres significantly increases. Here, we describe and test a novel low-cost tool, the DroughtBox, for phenotyping gres and Tp across species. The system consists of a programmable climatically controlled chamber in which branches dehydrate and changes in the mass recorded. Test measurements show that the DroughtBox maintains stable temperature and relative humidity across a range of set points, a prerequisite for getting accurate gres and Tp values. Among a study group of four conifer and one angiosperm species, we observed a range of gres (0.44-1.64 mmol H2 O m-2 s-1 ) and Tp (39.4-43.8°C) values. Furthermore, the measured time to hydraulic failure varied between two conifers species and was shortened in both species following a heatwave event. The DroughtBox is a reliable and customizable tool for phenotyping gres and Tp , as well as for testing models of time to hydraulic failure that will improve our ability to assess climate change impacts on plants.

Desiccation time during drought is highly predictable across species of Eucalyptus from contrasting climates.

Catastrophic failure of the water transport pathway in trees is a principal mechanism of mortality during extreme drought. To be able to predict the probability of mortality at an individual and landscape scale we need knowledge of the time for plants to reach critical levels of hydraulic failure. We grew plants of eight species of Eucalyptus originating from contrasting climates before allowing a subset to dehydrate. We tested whether a trait-based model of time to plant desiccation tcrit , from stomatal closure gs90 to a critical level of hydraulic dysfunction Ψcrit is consistent with observed dry-down times. Plant desiccation time varied among species, ranging from 96.2 to 332 h at a vapour-pressure deficit of 1 kPa, and was highly predictable using the tcrit model in conjunction with a leaf shedding function. Plant desiccation time was longest in species with high cavitation resistance, strong vulnerability segmentation, wide stomatal-hydraulic safety, and a high ratio of total plant water content to leaf area. Knowledge of tcrit in combination with water-use traits that influence stomatal closure could significantly increase our ability to predict the timing of drought-induced mortality at tree and forest scales.

Drought tolerance traits do not vary across sites differing in water availability in Banksia serrata (Proteaceae).

Interspecific variation in plant hydraulic traits plays a major role in shaping species distributions across climates, yet variation within species is poorly understood. Here we report on intraspecific variation of hydraulic traits in Banksia serrata (L.f.) sampled from three sites characterised by contrasting climates (warm-wet, warm-dry and cool-wet). Hydraulic characteristics including vulnerability to embolism, hydraulic conductance, pressure-volume traits and key morphological traits were measured. Vulnerability to embolism in leaf and stem, defined by the water potential inducing 50 and 88% loss of hydraulic conductivity (P50 and P88 respectively), did not differ across sites. However, plants from the warm-dry environment exhibited higher stem conductivity (Ks) than the cool-wet environment. Leaf turgor loss point (TLP) did not vary among sites, but warm-dry site plants showed lower leaf capacitance (C*FT) and higher modulus of elasticity (ε) than the other two sites. Plants from the cool-wet site had lower specific leaf area (SLA) and plants from the warm-dry site had lower sapwood density (WD). Overall, key hydraulic traits were generally conserved across populations despite differences in mean site water availability, and the safety-efficiency trade-off was absent in this species. These results suggest that B. serrata has limited ability to adjust hydraulic architecture in response to environmental change and thus may be susceptible to climate change-type drought stress.

Drought response strategies and hydraulic traits contribute to mechanistic understanding of plant dry-down to hydraulic failure.

Drought-induced tree mortality alters forest structure and function, yet our ability to predict when and how different species die during drought remains limited. Here, we explore how stomatal control and drought tolerance traits influence the duration of drought stress leading to critical levels of hydraulic failure. We examined the growth and physiological responses of four woody plant species (three angiosperms and one conifer) representing a range of water-use and drought tolerance traits over the course of two controlled drought-recovery cycles followed by an extended dry-down. At the end of the final dry-down phase, we measured changes in biomass ratios and leaf carbohydrates. During the first and second drought phases, plants of all species closed their stomata in response to decreasing water potential, but only the conifer species avoided water potentials associated with xylem embolism as a result of early stomatal closure relative to thresholds of hydraulic dysfunction. The time it took plants to reach critical levels of water stress during the final dry-down was similar among the angiosperms (ranging from 39 to 57 days to stemP88) and longer in the conifer (156 days to stemP50). Plant dry-down time was influenced by a number of factors including species stomatal-hydraulic safety margin (gsP90 - stemP50), as well as leaf succulence and minimum stomatal conductance. Leaf carbohydrate reserves (starch) were not depleted at the end of the final dry-down in any species, irrespective of the duration of drought. These findings highlight the need to consider multiple structural and functional traits when predicting the timing of hydraulic failure in plants.

Coordination between leaf, stem, and root hydraulics and gas exchange in three arid-zone angiosperms during severe drought and recovery.

The ability to resist hydraulic dysfunction in leaves, stems, and roots strongly influences whether plants survive and recover from drought. However, the coordination of hydraulic function among different organs within species and their links to gas exchange during drought and recovery remains understudied. Here, we examine the interaction between gas exchange and hydraulic function in the leaves, stems, and roots of three semiarid evergreen species exposed to a cycle of severe water stress (associated with substantial cavitation) and recovery. In all species, stomatal closure occurred at water potentials well before 50% loss of stem hydraulic conductance, while in two species, leaves and/or roots were more vulnerable than stems. Following soil rewetting, leaf-level photosynthesis (Anet ) returned to prestress levels within 2-4 weeks, whereas stomatal conductance and canopy transpiration were slower to recover. The recovery of Anet was decoupled from the recovery of leaf, stem, and root hydraulics, which remained impaired throughout the recovery period. Our results suggest that in addition to high embolism resistance, early stomatal closure and hydraulic vulnerability segmentation confers drought tolerance in these arid zone species. The lack of substantial embolism refilling within all major organs suggests that vulnerability of the vascular system to drought-induced dysfunction is a defining trait for predicting postdrought recovery.

Photosynthesis and carbon allocation are both important predictors of genotype productivity responses to elevated CO2 in Eucalyptus camaldulensis.

Intraspecific variation in biomass production responses to elevated atmospheric carbon dioxide (eCO2) could influence tree species' ecological and evolutionary responses to climate change. However, the physiological mechanisms underlying genotypic variation in responsiveness to eCO2 remain poorly understood. In this study, we grew 17 Eucalyptus camaldulensis Dehnh. subsp. camaldulensis genotypes (representing provenances from four different climates) under ambient atmospheric CO2 and eCO2. We tested whether genotype leaf-scale photosynthetic and whole-tree carbon (C) allocation responses to eCO2 were predictive of genotype biomass production responses to eCO2. Averaged across genotypes, growth at eCO2 increased in situ leaf net photosynthesis (Anet) (29%) and leaf starch concentrations (37%). Growth at eCO2 reduced the maximum carboxylation capacity of Rubisco (-4%) and leaf nitrogen per unit area (Narea, -6%), but Narea calculated on a total non-structural carbohydrate-free basis was similar between treatments. Growth at eCO2 also increased biomass production and altered C allocation by reducing leaf area ratio (-11%) and stem mass fraction (SMF, -9%), and increasing leaf mass area (18%) and leaf mass fraction (5%). Overall, we found few significant CO2 × provenance or CO2 × genotype (within provenance) interactions. However, genotypes that showed the largest increases in total dry mass at eCO2 had larger increases in root mass fraction (with larger decreases in SMF) and photosynthetic nitrogen-use efficiency (PNUE) with CO2 enrichment. These results indicate that genetic differences in PNUE and carbon sink utilization (in roots) are both important predictors of tree productivity responsiveness to eCO2.

Xylem embolism measured retrospectively is linked to canopy dieback in natural populations of Eucalyptus piperita following drought.

Manipulative experiments have suggested that embolism-induced hydraulic impairment underpins widespread tree mortality during extreme drought, yet in situ evidence is rare. One month after drought-induced leaf and branch dieback was observed in field populations of Eucalyptus piperita Sm. in the Blue Mountains (Australia), we measured the level of native stem embolism and characterized the extent of leaf death in co-occurring dieback and healthy (non-dieback) trees. We found that canopy dieback-affected trees showed significantly higher levels of native embolism (26%) in tertiary order branchlets than healthy trees (11%). Furthermore, there was a significant positive correlation (R2 = 0.51) between the level of leaf death and the level of native embolism recorded in branchlets from dieback-affected trees. This retrospective study suggests that hydraulic failure was the primary mechanism of leaf and branch dieback in response to a natural drought event in the field. It also suggests that post-drought embolism refilling is minimal or absent in this species of eucalypt.

The links between leaf hydraulic vulnerability to drought and key aspects of leaf venation and xylem anatomy among 26 Australian woody angiosperms from contrasting climates.

Background and Aims: The structural properties of leaf venation and xylem anatomy strongly influence leaf hydraulics, including the ability of leaves to maintain hydraulic function during drought. Here we examined the strength of the links between different leaf venation traits and leaf hydraulic vulnerability to drought (expressed as P50leaf by rehydration kinetics) in a diverse group of 26 woody angiosperm species, representing a wide range of leaf vulnerabilities, from four low-nutrient sites with contrasting rainfall across eastern Australia. Methods: For each species we measured key aspects of leaf venation design, xylem anatomy and leaf morphology. We also assessed for the first time the scaling relationships between hydraulically weighted vessel wall thickness (th) and lumen breadth (bh) across vein orders and habitats. Key Results: Across species, variation in P50leaf was strongly correlated with the ratio of vessel wall thickness (th) to lumen breadth (bh) [(t/b)h; an index of conduit reinforcement] at each leaf vein order. Concomitantly, the scaling relationship between th and bh was similar across vein orders, with a log-log slope less than 1 indicating greater xylem reinforcement in smaller vessels. In contrast, P50leaf was not related to th and bh individually, to major vein density (Dvmajor) or to leaf size. Principal components analysis revealed two largely orthogonal trait groupings linked to variation in leaf size and drought tolerance. Conclusions: Our results indicate that xylem conduit reinforcement occurs throughout leaf venation, and remains closely linked to leaf drought tolerance irrespective of leaf size.

Vessel scaling in evergreen angiosperm leaves conforms with Murray's law and area-filling assumptions: implications for plant size, leaf size and cold tolerance.

Water transport in leaf vasculature is a fundamental process affecting plant growth, ecological interactions and ecosystem productivity, yet the architecture of leaf vascular networks is poorly understood. Although Murray's law and the West-Brown-Enquist (WBE) theories predict convergent scaling of conduit width and number, it is not known how conduit scaling is affected by habitat aridity or temperature. We measured the scaling of leaf size, conduit width and conduit number within the leaves of 36 evergreen Angiosperms spanning a large range in aridity and temperature in eastern Australia. Scaling of conduit width and number in midribs and 2° veins did not differ across species and habitats (P > 0.786), and did not differ from that predicted by Murray's law (P = 0.151). Leaf size was strongly correlated with the hydraulic radius of petiole conduits (r2  = 0.83, P < 0.001) and did not differ among habitats (P > 0.064), nor did the scaling exponent differ significantly from that predicted by hydraulic theory (P = 0.086). The maximum radius of conduits in petioles was positively correlated with the temperature of the coldest quarter (r2  = 0.67; P < 0.001), suggesting that habitat temperature restricts the occurrence of wide-conduit species in cold habitats.

CO2 availability influences hydraulic function of C3 and C4 grass leaves.

Atmospheric CO2 (ca) has increased since the last glacial period, increasing photosynthetic water use efficiency and improving plant productivity. Evolution of C4 photosynthesis at low ca led to decreased stomatal conductance (gs), which provided an advantage over C3 plants that may be reduced by rising ca. Using controlled environments, we determined how increasing ca affects C4 water use relative to C3 plants. Leaf gas exchange and mass per area (LMA) were measured for four C3 and four C4 annual, crop-related grasses at glacial (200 µmol mol-1), ambient (400 µmol mol-1), and super-ambient (640 µmol mol-1) ca. C4 plants had lower gs, which resulted in a water use efficiency advantage at all ca and was broadly consistent with slower stomatal responses to shade, indicating less pressure on leaf water status. At glacial ca, net CO2 assimilation and LMA were lower for C3 than for C4 leaves, and C3 and C4 grasses decreased leaf hydraulic conductance (Kleaf) similarly, but only C4 leaves decreased osmotic potential at turgor loss. Greater carbon availability in C4 leaves at glacial ca generated a different hydraulic adjustment relative to C3 plants. At current and future ca, C4 grasses have advantages over C3 grasses due to lower gs, lower stomatal sensitivity, and higher absolute water use efficiency.

Trees tolerate an extreme heatwave via sustained transpirational cooling and increased leaf thermal tolerance.

Heatwaves are likely to increase in frequency and intensity with climate change, which may impair tree function and forest C uptake. However, we have little information regarding the impact of extreme heatwaves on the physiological performance of large trees in the field. Here, we grew Eucalyptus parramattensis trees for 1 year with experimental warming (+3°C) in a field setting, until they were greater than 6 m tall. We withheld irrigation for 1 month to dry the surface soils and then implemented an extreme heatwave treatment of 4 consecutive days with air temperatures exceeding 43°C, while monitoring whole-canopy exchange of CO2 and H2 O, leaf temperatures, leaf thermal tolerance, and leaf and branch hydraulic status. The heatwave reduced midday canopy photosynthesis to near zero but transpiration persisted, maintaining canopy cooling. A standard photosynthetic model was unable to capture the observed decoupling between photosynthesis and transpiration at high temperatures, suggesting that climate models may underestimate a moderating feedback of vegetation on heatwave intensity. The heatwave also triggered a rapid increase in leaf thermal tolerance, such that leaf temperatures observed during the heatwave were maintained within the thermal limits of leaf function. All responses were equivalent for trees with a prior history of ambient and warmed (+3°C) temperatures, indicating that climate warming conferred no added tolerance of heatwaves expected in the future. This coordinated physiological response utilizing latent cooling and adjustment of thermal thresholds has implications for tree tolerance of future climate extremes as well as model predictions of future heatwave intensity at landscape and global scales.