Low acclimation potential compromises the performance of water-stressed pine saplings under Mediterranean xeric conditions.

Predicted hotter and drier climatic conditions in the Mediterranean Basin will probably hamper current afforestations and reforestations by negatively influencing tree performance. Understanding how saplings can adjust their physiology to shortages in water availability is essential to predict early-stage success of forest ecological restoration. Pines are common target species used in afforestations and reforestations; however, the capacity of their saplings for physiological plasticity to promote drought tolerance remains largely unexplored. In this study, we evaluated the demographical and resource-use consequences of short-term irrigation among four pine species (Pinus halepensis, Pinus pinea, Pinus nigra and Pinus sylvestris) growing under water-limiting conditions in a common garden experiment. Summer irrigation increased the survival rate of those pines that were suffering from hydric stress under the xeric conditions of the common garden (i.e. P. pinea, P. nigra and P. sylvestris). Moreover, short-term water supplementation slightly enhanced aboveground biomass production across species. However, leaf isotopic composition and nutrient concentrations did not change after summer irrigation. Independently of water supplementation, P. halepensis was the best adapted species to water scarcity and showed the best physiological and growth performance. By contrast, P. pinea, P. nigra and P. sylvestris saplings exhibited drought-induced reductions in stomatal conductance and low water-use efficiency, nutrient deficiency, and severe N:P and N:K stoichiometric imbalances, leading to impaired growth. We conclude that the lack of physiological plasticity of water-stressed pine saplings to withstand the impacts of climate aridification will likely cause severe impairment of their nutrient status, growth and survival, with dire implications for the successful establishment of Mediterranean afforestation and reforestation programs.

Squandering water in drylands: the water-use strategy of the phreatophyte Ziziphus lotus in a groundwater-dependent ecosystem.

PREMISE: Water is the most limiting factor in dryland ecosystems, and plants are adapted to cope with this constraint. Particularly vulnerable are phreatophytic plants from groundwater-dependent ecosystems (GDEs) in regions that have to face water regime alterations due to the impacts of climate and land-use changes. METHODS: We investigated two aspects related to the water-use strategy of a keystone species that dominates one of the few terrestrial GDEs in European drylands (Ziziphus lotus): where it obtains water and how it regulates its use. We (1) evaluated plants' water sources and use patterns using a multiple-isotope approach (δ2 H, δ18 O, and Δ13 C); (2) assessed the regulation of plant water potential by characterizing the species on an isohydric-anisohydric continuum; and (3) evaluated plants' response to increasing water stress along a depth-to-groundwater (DTGW) gradient by measuring foliar gas exchange and nutrient concentrations. RESULTS: Ziziphus lotus behaves as a facultative or partial phreatophyte with extreme anisohydric stomatal regulation. However, as DTGW increased, Z. lotus (1) reduced the use of groundwater, (2) reduced total water uptake, and (3) limited transpiration water loss while increasing water-use efficiency. We also found a physiological threshold at 14 m depth to groundwater, which could indicate maximum rooting length beyond which optimal plant function could not be sustained. CONCLUSIONS: Species such as Z. lotus survive by squandering water in drylands because of a substantial groundwater uptake. However, the identification of DTGW thresholds indicates that drawdowns in groundwater level would jeopardize the functioning of the GDE.

Water uptake depth is coordinated with leaf water potential, water-use efficiency and drought vulnerability in karst vegetation.

Root access to bedrock water storage or groundwater is an important trait allowing plant survival in seasonally dry environments. However, the degree of coordination between water uptake depth, leaf-level water-use efficiency (WUEi) and water potential in drought-prone plant communities is not well understood. We conducted a 135-d rainfall exclusion experiment in a subtropical karst ecosystem with thin skeletal soils to evaluate the responses of 11 co-occurring woody species of contrasting life forms and leaf habits to a severe drought during the wet growing season. Marked differences in xylem water isotopic composition during drought revealed distinct ecohydrological niche separation among species. The contrasting behaviour of leaf water potential in coexisting species during drought was largely explained by differences in root access to deeper, temporally stable water sources. Smaller-diameter species with shallower water uptake, more negative water potentials and lower WUEi showed extensive drought-induced canopy defoliation and/or mortality. By contrast, larger-diameter species with deeper water uptake, higher leaf-level WUEi and more isohydric behaviour survived drought with only moderate canopy defoliation. Severe water limitation imposes strong environmental filtering and/or selective pressures resulting in tight coordination between tree diameter, water uptake depth, iso/anisohydric behaviour, WUEi and drought vulnerability in karst plant communities.

Simulated climate change decreases nutrient resorption from senescing leaves.

Nutrient resorption is the process whereby plants recover nutrients from senescing leaves and reallocate them to storage structures or newer tissues. Elemental resorption of foliar N and P has been shown to respond to temperature and precipitation, but we know remarkably little about the influence of warming and drought on the resorption of these and other essential plant macro- and micronutrients, which could alter the ability of species to recycle their nutrients. We conducted a 5 year manipulative field study to simulate predicted climate change conditions and studied the effects of warming (W), rainfall reduction (RR), and their combination (W+RR) on nutrient resorption efficiency in five coexisting shrub species in a semiarid shrubland. Both mature and senesced leaves showed significant reductions in their nutrient contents and an altered stoichiometry in response to climate change conditions. Warming (W, W+RR) reduced mature leaf N, K, Ca, S, Fe, and Zn and senesced leaf N, Ca, Mg, S, Fe, and Zn contents relative to ambient temperature conditions. Warming increased mature leaf C/N ratios and decreased N/P and C/P ratios and increased senesced leaf C/N and C/P ratios. Furthermore, W and W+RR reduced nutrient resorption efficiencies for N (6.3%), K (19.8%), S (70.9%) and increased Ca and Fe accumulation in senesced leaves (440% and 35.7%, respectively) relative to the control treatment. Rainfall reduction decreased the resorption efficiencies of N (6.7%), S (51%), and Zn (46%). Reductions in nutrient resorption efficiencies with warming and/or rainfall reduction were rather uniform and consistent across species. The negative impacts of warming and rainfall reduction on foliar nutrient resorption efficiency will likely cause an impairment of plant nutrient budgets and fitness across coexisting native shrubs in this nutrient-poor habitat, with probable implications for key ecosystem functions such as reductions in nutrient retention in vegetation, litter decomposition, and nutrient cycling rates.

Long-term nutrient imbalances linked to drought-triggered forest dieback.

Drought-induced forest dieback is causing reductions in productivity, increasing tree mortality and impairing terrestrial carbon uptake worldwide. However, the role played by long-term nutrient imbalances during drought-induced dieback is still unknown. To improve our knowledge on the relationships between dieback and nutrient imbalances, we analysed wood anatomical traits (tree-ring width and wood density), soil properties and long-term chemical information in tree-ring wood (1900-2010) by non-destructive Micro X-ray fluorescence (µXRF) and destructive (ICP-OES) techniques. We studied two major European conifers with ongoing drought-induced dieback in mesic (Abies alba, silver fir) and xeric (Pinus sylvestris, Scots pine) sites. In each site we compared coexisting declining (D) and non-declining (ND) trees. We used dendrochronology and generalized additive and linear mixed models to analyse trends in tree-ring nutrients and their relationships with wood traits. The D trees presented lower growth and higher minimum wood density than ND trees, corresponding to a smaller lumen area of earlywood tracheids and thus a lower theoretical hydraulic conductivity. These differences in growth and wood-anatomy were more marked in silver fir than in Scots pine. Moreover, most of the chemical elements showed higher concentrations in D than in ND trees during the last two-five decades (e.g., Mn, K and Mg), while Ca and Na increased in the sapwood of ND trees. The Mn concentrations, and related ratios (Ca:Mn, Mn:Al and P:Mn) showed the highest differences between D and ND trees for both tree species. These findings suggest that a reduced hydraulic conductivity, consistent with hydraulic impairment, is affecting the use of P in D trees, making them more prone to drought-induced damage. The retrospective quantifications of Mn ratios may be used as early-warning signals of impending dieback.

The "isohydric trap": A proposed feedback between water shortage, stomatal regulation, and nutrient acquisition drives differential growth and survival of European pines under climatic dryness.

Climatic dryness imposes limitations on vascular plant growth by reducing stomatal conductance, thereby decreasing CO2 uptake and transpiration. Given that transpiration-driven water flow is required for nutrient uptake, climatic stress-induced nutrient deficit could be a key mechanism for decreased plant performance under prolonged drought. We propose the existence of an "isohydric trap," a dryness-induced detrimental feedback leading to nutrient deficit and stoichiometry imbalance in strict isohydric species. We tested this framework in a common garden experiment with 840 individuals of four ecologically contrasting European pines (Pinus halepensis, P. nigra, P. sylvestris, and P. uncinata) at a site with high temperature and low soil water availability. We measured growth, survival, photochemical efficiency, stem water potentials, leaf isotopic composition (δ13 C, δ18 O), and nutrient concentrations (C, N, P, K, Zn, Cu). After 2 years, the Mediterranean species Pinus halepensis showed lower δ18 O and higher δ13 C values than the other species, indicating higher time-integrated transpiration and water-use efficiency (WUE), along with lower predawn and midday water potentials, higher photochemical efficiency, higher leaf P, and K concentrations, more balanced N:P and N:K ratios, and much greater dry-biomass (up to 63-fold) and survival (100%). Conversely, the more mesic mountain pine species showed higher leaf δ18 O and lower δ13 C, indicating lower transpiration and WUE, higher water potentials, severe P and K deficiencies and N:P and N:K imbalances, and poorer photochemical efficiency, growth, and survival. These results support our hypothesis that vascular plant species with tight stomatal regulation of transpiration can become trapped in a feedback cycle of nutrient deficit and imbalance that exacerbates the detrimental impacts of climatic dryness on performance. This overlooked feedback mechanism may hamper the ability of isohydric species to respond to ongoing global change, by aggravating the interactive impacts of stoichiometric imbalance and water stress caused by anthropogenic N deposition and hotter droughts, respectively.

Plant δ15 N reflects the high landscape-scale heterogeneity of soil fertility and vegetation productivity in a Mediterranean semiarid ecosystem.

We investigated the magnitude and drivers of spatial variability in soil and plant δ15 N across the landscape in a topographically complex semiarid ecosystem. We hypothesized that large spatial heterogeneity in water availability, soil fertility and vegetation cover would be positively linked to high local-scale variability in δ15 N. We measured foliar δ15 N in three dominant plant species representing contrasting plant functional types (tree, shrub, grass) and mycorrhizal association types (ectomycorrhizal or arbuscular mycorrhizal). This allowed us to investigate whether δ15 N responds to landscape-scale environmental heterogeneity in a consistent way across species. Leaf δ15 N varied greatly within species across the landscape and was strongly spatially correlated among co-occurring individuals of the three species. Plant δ15 N correlated tightly with soil δ15 N and key measures of soil fertility, water availability and vegetation productivity, including soil nitrogen (N), organic carbon (C), plant-available phosphorus (P), water-holding capacity, topographic moisture indices and normalized difference vegetation index. Multiple regression models accounted for 62-83% of within-species variation in δ15 N across the landscape. The tight spatial coupling and interdependence of the water, N and C cycles in drylands may allow the use of leaf δ15 N as an integrative measure of variations in moisture availability, biogeochemical activity, soil fertility and vegetation productivity (or 'site quality') across the landscape.

Photosynthesis and growth reduction with warming are driven by nonstomatal limitations in a Mediterranean semi-arid shrub.

Whereas warming enhances plant nutrient status and photosynthesis in most terrestrial ecosystems, dryland vegetation is vulnerable to the likely increases in evapotranspiration and reductions in soil moisture caused by elevated temperatures. Any warming-induced declines in plant primary production and cover in drylands would increase erosion, land degradation, and desertification. We conducted a four-year manipulative experiment in a semi-arid Mediterranean ecosystem to evaluate the impacts of a ~2°C warming on the photosynthesis, transpiration, leaf nutrient status, chlorophyll content, isotopic composition, biomass growth, and postsummer survival of the native shrub Helianthemum squamatum. We predicted that warmed plants would show reduced photosynthetic activity and growth, primarily due to the greater stomatal limitation imposed by faster and more severe soil drying under warming. On average, warming reduced net photosynthetic rates by 36% across the study period. Despite this strong response, warming did not affect stomatal conductance and transpiration. The reduction of peak photosynthetic rates with warming was more pronounced in a drought year than in years with near-average rainfall (75% and 25-40% reductions relative to controls, respectively), with no indications of photosynthetic acclimation to warming through time. Warmed plants had lower leaf N and P contents, δ (13)C, and sparser and smaller leaves than control plants. Warming reduced shoot dry mass production by 31%. However, warmed plants were able to cope with large reductions in net photosynthesis, leaf area, and shoot biomass production without changes in postsummer survival rates. Our findings highlight the key role of nonstomatal factors (biochemical and/or nutritional) in reducing net carbon assimilation rates and growth under warming, which has important implications for projections of plant carbon balance under the warmer and drier climatic scenario predicted for drylands worldwide. Projected climate warming over the coming decades could reduce net primary production by about one-third in semi-arid gypsum shrublands dominated by H. squamatum.

Growth and stable isotope signals associated with drought-related mortality in saplings of two coexisting pine species.

Drought-induced events of massive tree mortality appear to be increasing worldwide. Species-specific vulnerability to drought mortality may alter patterns of species diversity and affect future forest composition. We have explored the consequences of the extreme drought of 2005, which caused high sapling mortality (approx. 50 %) among 10-year-old saplings of two coexisting pine species in the Mediterranean mountains of Sierra Nevada (Spain): boreo-alpine Pinus sylvestris and Mediterranean P. nigra. Sapling height growth, leaf δ(13)C and δ(18)O, and foliar nitrogen concentration in the four most recent leaf cohorts were measured in dead and surviving saplings. The foliar isotopic composition of dead saplings (which reflects time-integrated leaf gas-exchange until mortality) displayed sharp increases in both δ(13)C and δ(18)O during the extreme drought of 2005, suggesting an important role of stomatal conductance (g(s)) reduction and diffusional limitations to photosynthesis in mortality. While P. nigra showed decreased growth in 2005 compared to the previous wetter year, P. sylvestris maintained similar growth levels in both years. Decreased growth, coupled with a sharper increase in foliar δ(18)O during extreme drought in dead saplings, indicate a more conservative water use strategy for P. nigra. The different physiological behavior of the two pine species in response to drought (further supported by data from surviving saplings) may have influenced 2005 mortality rates, which contributed to 2.4-fold greater survival for P. nigra over the lifespan of the saplings. This species-specific vulnerability to extreme drought could lead to changes in dominance and distribution of pine species in Mediterranean mountain forests.

Leaf δ18O of remaining trees is affected by thinning intensity in a semiarid pine forest.

Silvicultural thinning usually improves the water status of remaining trees in water-limited forests. We evaluated the usefulness of a dual stable isotope approach (δ¹³C, δ¹8O) for comparing the physiological performance of remaining trees between forest stands subjected to two different thinning intensities (moderate versus heavy) in a 60-year-old Pinus halepensis Mill. plantation in semiarid southeastern Spain. We measured bulk leaf δ¹³C and δ¹8O, foliar elemental concentrations, stem water content, stem water δ¹8O (δ¹8O(stem water)), tree ring widths and leaf gas exchange rates to assess the influence of forest stand density on tree performance. Remaining trees in low-density stands (heavily thinned) showed lower leaf δ¹8O, and higher stomatal conductance (g(s)), photosynthetic rate and radial growth than those in moderate-density stands (moderately thinned). By contrast, leaf δ¹³C, intrinsic water-use efficiency, foliar elemental concentrations and δ¹8O(stem water) were unaffected by stand density. Lower foliar δ¹8O in heavily thinned stands reflected higher g(s) of remaining trees due to decreased inter-tree competition for water, whereas higher photosynthetic rate was largely attributable to reduced stomatal limitation to CO2 uptake. The dual isotope approach provided insight into the early (12 months) effects of stand density manipulation on the physiological performance of remaining trees.

Ecosystem CO2 fluxes of arbuscular and ectomycorrhizal dominated vegetation types are differentially influenced by precipitation and temperature.

Here, we explore how interannual variations in environmental factors (i.e. temperature, precipitation and light) influence CO(2) fluxes (gross primary production and ecosystem respiration) in terrestrial ecosystems classified by vegetation type and the mycorrhizal type of dominant plants (arbuscular mycorrhizal (AM) or ectomycorrhizal (EM)). We combined 236 site-year measurements of terrestrial ecosystem CO(2) fluxes and environmental factors from 50 eddy-covariance flux tower sites with information about climate, vegetation type and dominant plant species. Across large geographical distances, interannual variations in ecosystem CO(2) fluxes for EM-dominated sites were primarily controlled by interannual variations in mean annual temperature. By contrast, interannual variations in ecosystem CO(2) fluxes at AM-dominated sites were primarily controlled by interannual variations in precipitation. This study represents the first large-scale assessment of terrestrial CO(2) fluxes in multiple vegetation types classified according to dominant mycorrhizal association. Our results support and complement the hypothesis that bioclimatic conditions influence the distribution of AM and EM systems across large geographical distances, which leads to important differences in the major climatic factors controlling ecosystem CO(2) fluxes.

Stable isotope views on ecosystem function: challenging or challenged?

Stable isotopes and their potential for detecting various and complex ecosystem processes are attracting an increasing number of scientists. Progress is challenging, particularly under global change scenarios, but some established views have been challenged. The IX meeting of the Spanish Association of Terrestrial Ecology (AAET, Ubeda, 18-22 October 2009) hosted a symposium on the ecology of stable isotopes where the linear mixing model approach of partitioning sinks and sources of carbon and water fluxes within an ecosystem was challenged, and new applications of stable isotopes for the study of plant interactions were evaluated. Discussion was also centred on the need for networks that monitor ecological processes using stable isotopes and key ideas for fostering future research with isotopes.

Bulk leaf delta(18)O and delta(13)C reflect the intensity of intraspecific competition for water in a semi-arid tussock grassland.

We investigated the extent to which plant water and nutrient status are affected by intraspecific competition intensity and microsite quality in a monodominant tussock grassland. Leaf gas exchange and stable isotope measurements were used to assess the water relations of Stipa tenacissima tussocks growing along a gradient of plant cover and soil depth in a semi-arid catchment of Southeast Spain. Stomatal conductance and photosynthetic rate decreased with increasing intensity of competition during the wet growing season, leading to foliar delta(18)O and delta(13)C enrichment. A high potential for runoff interception by upslope neighbours exerted strong detrimental effects on the water and phosphorus status of downslope S. tenacissima tussocks. Foliar delta(15)N values became more enriched with increasing soil depth. Multiple stepwise regression showed that competition potential and/or rhizosphere soil depth accounted for large proportions of variance in foliar delta(13)C, delta(18)O and delta(15)N among target tussocks (57, 37 and 64%, respectively). The results presented here highlight the key role that spatial redistribution of resources (water and nutrients) by runoff plays in semi-arid ecosystems. It is concluded that combined measurement of delta(13)C, delta(18)O and nutrient concentrations in bulk leaf tissue can provide insight into the intensity of competitive interactions occurring in natural plant communities.

Topographic position modulates the mycorrhizal response of oak trees to interannual rainfall variability.

California coast live oak (Quercus agrifolia) forms tripartite symbiotic associations with arbuscular (AMF) and ectomycorrhizal (EMF) fungi. We selected oak individuals differing in topographic position and depth to groundwater (mesic valley vs. xeric hill sites) to investigate changes of tree mycorrhizal status in response to interannual rainfall variability. EMF root colonization, as well as hyphal abundance and viability in upper rhizosphere soil (0-30 cm), were negatively affected by severe multi-year drought, although not to the same extent in each topographic location. Oak trees growing in hill sites showed EMF colonization levels <1% in upper roots during drought. By contrast, oaks in valley sites maintained much higher EMF colonization (>19%) in upper roots during drought. EMF root colonization increased sharply at both topographic positions during the ensuing wet year (78% in valley, 49% in hill), which indicates that the mycorrhizal status of roots in upper rhizosphere soil is highly responsive to interannual rainfall variability. Across sites and years, percentage EMF colonization and soil hyphal density and viability were strongly positively correlated with soil moisture potential, but percentage AMF root colonization was not. Interestingly, changes in percentage EMF root colonization and density of viable hyphae between a wet and a dry year were proportionally much greater in xeric hill sites than in mesic valley sites. The mycorrhizal status of oak trees was particularly responsive to changes in soil moisture at the hill sites, where roots in upper rhizosphere soil shifted from almost exclusively AMF during severe drought to predominantly EMF during the ensuing wet year. By contrast, the mycorrhizal status of oaks in the valley sites was less strongly coupled to current meteorological conditions, as roots in upper soil layers remained predominantly EMF during both a dry and a wet year. Canopy shading and hydraulic lift by oaks in valley sites likely contributed to maintain the integrity and viability of EMF roots and extraradical hyphae in upper rhizosphere soil during extended drought. Our results suggest that oak woodlands in water-limited ecosystems may become increasingly reliant on the AMF symbiosis under future climate change scenarios for the U.S. southwest and other world regions.

Plant isotopic composition provides insight into mechanisms underlying growth stimulation by AM fungi in a semiarid environment.

We hypothesised that improved plant water status and enhanced transpiration are key mechanisms involved in plant growth stimulation by native arbuscular mycorrhizal fungi (AMF) in semiarid calcareous soils. Seedlings of the dryland shrubs Pistacia lentiscus L. and Retama sphaerocarpa L. were pre-inoculated with a mixture of eight native Glomus spp. fungi, or left un-inoculated, before transplanting into a degraded site in south-eastern Spain. Pre-inoculated Pistacia and Retama shrubs grew faster after transplanting, despite spontaneous colonisation of control plants by local AMF. Pre-inoculation enhanced shoot water content and shoot δ15N in both shrub species. Increased potassium uptake and improved water relations were key mechanisms behind growth stimulation by native AMF in Pistacia. Shoot δ18O (a proxy measure of stomatal conductance) was significantly less negative in AMF-inoculated than in control Pistacia seedlings, indicating enhanced cumulative transpiration in the former. In contrast, shoot δ18O was unaffected by AMF inoculation in Retama, a leafless leguminous shrub with photosynthetic stems. Growth stimulation by native AMF in Retama was attributed to increased phosphorus uptake, enhanced atmospheric nitrogen fixation and a largely nutrient-mediated improvement of plant water status. Shoot δ13C was not significantly influenced by AMF inoculation in either shrub species, thus suggesting roughly parallel upshifts in photosynthetic and transpiration rates which did not affect plant water use efficiency.

Corrigendum to: Plant isotopic composition provides insight into mechanisms underlying growth stimulation by AM fungi in a semiarid environment.

We hypothesised that improved plant water status and enhanced transpiration are key mechanisms involved in plant growth stimulation by native arbuscular mycorrhizal fungi (AMF) in semiarid calcareous soils. Seedlings of the dryland shrubs Pistacia lentiscus L. and Retama sphaerocarpa L. were pre-inoculated with a mixture of eight native Glomus spp. fungi, or left un-inoculated, before transplanting into a degraded site in south-eastern Spain. Pre-inoculated Pistacia and Retama shrubs grew faster after transplanting, despite spontaneous colonisation of control plants by local AMF. Pre-inoculation enhanced shoot water content and shoot ´15N in both shrub species. Increased potassium uptake and improved water relations were key mechanisms behind growth stimulation by native AMF in Pistacia. Shoot ´18O (a proxy measure of stomatal conductance) was significantly less negative in AMF-inoculated than in control Pistacia seedlings, indicating enhanced cumulative transpiration in the former. In contrast, shoot ´18O was unaffected by AMF inoculation in Retama, a leafless leguminous shrub with photosynthetic stems. Growth stimulation by native AMF in Retama was attributed to increased phosphorus uptake, enhanced atmospheric nitrogen fixation and a largely nutrient-mediated improvement of plant water status. Shoot ´13C was not significantly influenced by AMF inoculation in either shrub species, thus suggesting roughly parallel upshifts in photosynthetic and transpiration rates which did not affect plant water use efficiency.