Different drought-tolerance strategies of tree species to cope with increased water stress under climate change in a mixed forest.

Trees' functional strategies to cope with extreme drought are essential under climate change. In a mixed Mediterranean forest, we analyzed the functional strategy in response to drought of four co-occurring species (Pinus pinea, Pinus pinaster, Juniperus oxycedrus, and Quercus ilex) during two years. Specifically, we assessed functional traits related to tree water status, leaf water relations, and gas exchange. Different trait-syndrome metrics and the functional strategies under water stress observed suggested a species drought-tolerance differentiation, with the more anysohidric Q. ilex and J. oxycedrus showing a much higher drought tolerance than the more isohydric P. pinea and P. pinaster. All species recovered from negative leaf turgor reached during peak water stress in summer. Q. ilex and J. oxycedrus kept lower leaf osmotic potentials and lower sensitivity of leaf gas exchange and leaf photochemistry to water stress. In contrast, the pine species exhibited more drought-avoidant and water-conservative strategies, yet this behavior was less effective in mitigating water stress's impact on their physiology. The pine species were the most affected by drought, with prolonged near-zero net photosynthesis during summer. P. pinaster was more isohydric than P. pinea and exhibited a lower capacity to maintain leaf turgor. Physiological processes regulating leaf turgor under drought constitute a key functional strategy involved in the carbon and water-related mechanisms, ultimately inducing mortality under hot drought. The currently observed mortality dynamics for P. pinaster, and to a lower extent in P. pinea, may be exacerbated by loss of functional homeostasis.

Snowpack permanence shapes the growth and dynamic of non-structural carbohydrates in Juniperus communis in alpine tundra.

Climate warming is altering snowpack permanence in alpine tundra, modifying shrub growth and distribution. Plant acclimation to snowpack changes depends on the capability to guarantee growth and carbon storage, suggesting that the content of non-structural carbohydrates (NSC) in plant organs can be a key trait to depict the plant response under different snow regimes. To test this hypothesis, we designed a 3-years long manipulative experiment aimed at evaluating the effect of snow melt timing (i.e., early, control, and late) on NSC content in needles, bark and wood of Juniperus communis L. growing at high elevation in the Alps. Starch evidenced a general decrease from late spring to summer in control and early melting, while starch was low but stable in plants subjected to a late snow melt. Leaves, bark and wood have different level of soluble NSC changing during growing season: in bark, sugars content decreased significantly in late summer, while there was no seasonal effect in needles and wood. Soluble NSC and starch were differently related with the plant growth, when considering different tissues and snow treatment. In leaf and bark we observed a starch depletion in control and early melting plants, consistently to a higher growth (i.e., twig elongation), while in late snow melt, we did not find any significant relationship between growth and NSC concentration. Our findings confirmed that snowpack duration affects the onset of the growing season promoting a change in carbon allocation in plant organs and, between bark and wood in twigs. Finally, our results suggest that plants, at this elevation, could take advantage from an early snow melt caused by climate warming, most likely due to photosynthetic activity by maintaining the level of reserves and enhancing the carbon investment for growth.

Dynamic regulation of water potential in Juniperus osteosperma mediates ecosystem carbon fluxes.

Some plants exhibit dynamic hydraulic regulation, in which the strictness of hydraulic regulation (i.e. iso/anisohydry) changes in response to environmental conditions. However, the environmental controls over iso/anisohydry and the implications of flexible hydraulic regulation for plant productivity remain unknown. In Juniperus osteosperma, a drought-resistant dryland conifer, we collected a 5-month growing season time series of in situ, high temporal-resolution plant water potential ( Ψ ) and stand gross primary productivity (GPP). We quantified the stringency of hydraulic regulation associated with environmental covariates and evaluated how predawn water potential contributes to empirically predicting carbon uptake. Juniperus osteosperma showed less stringent hydraulic regulation (more anisohydric) after monsoon precipitation pulses, when soil moisture and atmospheric demand were high, and corresponded with GPP pulses. Predawn water potential matched the timing of GPP fluxes and improved estimates of GPP more strongly than soil and/or atmospheric moisture, notably resolving GPP underestimation before vegetation green-up. Flexible hydraulic regulation appears to allow J. osteosperma to prolong soil water extraction and, therefore, the period of high carbon uptake following monsoon precipitation pulses. Water potential and its dynamic regulation may account for why process-based and empirical models commonly underestimate the magnitude and temporal variability of dryland GPP.

Gone with the waves: the role of sea currents as key dispersal mechanism for Mediterranean coastal and inland plant species.

Thalassochory, the dispersal of propagules through marine currents, is a key long-distance dispersal (LDD) mechanism with implications for global biogeography and particularly for island colonization. The propagules of coastal plant species are generally assumed to be better adapted for sea dispersal than those of inland plants, but this hypothesis remains largely untested. We conducted experiments on four genera (Juniperus, Daucus, Ferula, and Pancratium) and compared traits among nine species with different habitats and distributions. Our results showed that Juniperus spp. and P. maritimum have strong thalassochorous potential within the Mediterranean Basin. Interestingly, we did not find a clear association on the thalassochorous potential of coastal versus inland species within all the tested genera, apart from P. maritimum compared with the endemic inland P. illyricum. These findings suggest that thalassochory may be a more common dispersal mechanism than previously assumed. The apparently weak link of dispersal syndrome with species ecology broadens the possibility of dispersal by the sea also for inland plants, although considered to be poorly salt-tolerant. Moreover, our results reveal significant differences in sea dispersal between endemic and widespread species, but do not rule out an important role of thalassochory in shaping the distribution patterns of archipelago endemic flora. The presented method is largely replicable and could be used for further studies with a larger set of species to better delineate trends of sea dispersal syndrome among species with different ecology or dispersal traits.

Heavy metal accumulation characteristics and physiological response of Sabina chinensis and Platycladus orientalis to atmospheric pollution.

Heavy metal (Cu, Mn, Zn, Pb, and Cd) concentrations were measured in the leaves of Sabina chinensis and Platycladus orientalis collected from urban, suburban, and rural sites in Tianjin, China. Photosynthetic pigment contents, reactive oxygen species content, malondialdehyde (MDA) content and antioxidant enzyme activity were investigated, providing physiological response parameters. Our comparison of the sites revealed that urbanization significantly influenced the heavy metal concentrations in both plant leaves. At the rural site, both plant leaves exhibited the lowest heavy metal accumulation. The highest Cu, Mn, and Zn concentrations were found in S. chinensis leaves from the urban site; the highest Pb and Cd concentrations were found in P. orientalis leaves from the urban site. These results indicate that the urban site contained larger heavy metal concentrations in the plant leaves that may reflect the anthropogenic emission gradient. It is also found that S. chinensis may be used to monitor airborne heavy metal pollution because it is highly quick response to heavy metals, while P. orientalis may be used for mitigation due to its high resistance. The results of this study can contribute to the development of monitoring and environmental management plans by providing information on sensitive and resistant tree species for city greening in North China.

Foliar respiration is related to photosynthetic, growth and carbohydrate response to experimental drought and elevated temperature.

Short-term plant respiration (R) increases exponentially with rising temperature, but drought could reduce respiration by reducing growth and metabolism. Acclimation may alter these responses. We examined if species with different drought responses would differ in foliar R response to +4.8°C temperature and -45% precipitation in a field experiment with mature piñon and juniper trees, and if any differences between species were related to differences in photosynthesis rates, shoot growth and nonstructural carbohydrates (NSCs). Short-term foliar R had a Q10 of 1.6 for piñon and 2.6 for juniper. Piñon foliar R did not respond to the +4.8°C temperatures, but R increased 1.4× for juniper. Across treatments, piñon foliage had higher growth, lower NSC content, 29% lower photosynthesis rates, and 44% lower R than juniper. Removing 45% precipitation had little impact on R for either species. Species differences in the response of R under elevated temperature were related to substrate availability and stomatal response to leaf water potential. Despite not acclimating to the higher temperature and having higher R than piñon, greater substrate availability in juniper suggests it could supply respiratory demand for much longer than piñon. Species responses will be critical in ecosystem response to a warmer climate.

The higher availability of nutrients increases the production but decreases the quality of pollen grains in Juniperus communis L.

Next to global warming, excessive deposition of nitrogen has an alarming environmental impact on forest ecology, especially within dioecious species. Resource availability affects seed quality and can affect the distribution of plant species. Lower seed productivity can also be a result of limited pollen availability or lower pollen quality. A few studies have assessed the effect of nutrient availability on the quantity and quality of pollen grains produced. Therefore, rooted shoots of dioecious, male Juniperus communis L. grown in different nutritional conditions were used to assess the impact on productivity and quality attributes of produced pollen grains (pollen volume, morphology, germination, and chemical composition). The results indicated that nutrient availability impacts pollen grain development. Male plants growing in nutrient-rich environments appear to compensate for the lower quality of produced pollen grains by producing a higher number of male cones and thus a greater quantity of pollen. In contrast, the opposite was observed in plants growing in nutrient-poor environments. The availability of nitrogen and other nutrients will probably continue to impact soils in the foreseeable future due to anthropogenic activity and can be one of the drivers that can impact the reproduction and distribution of plants.

Conifers depend on established roots during drought: results from a coupled model of carbon allocation and hydraulics.

Trees may survive prolonged droughts by shifting water uptake to reliable water sources, but it is unknown if the dominant mechanism involves activating existing roots or growing new roots during drought, or some combination of the two. To gain mechanistic insights on this unknown, a dynamic root-hydraulic modeling framework was developed that set up a feedback between hydraulic controls over carbon allocation and the role of root growth on soil-plant hydraulics. The new model was tested using a 5 yr drought/heat field experiment on an established piñon-juniper stand with root access to bedrock groundwater. Owing to the high carbon cost per unit root area, modeled trees initialized without adequate bedrock groundwater access experienced potentially lethal declines in water potential, while all of the experimental trees maintained nonlethal water potentials. Simulated trees were unable to grow roots rapidly enough to mediate the hydraulic stress, particularly during warm droughts. Alternatively, modeled trees initiated with root access to bedrock groundwater matched the hydraulics of the experimental trees by increasing their water uptake from bedrock groundwater when soil layers dried out. Therefore, the modeling framework identified a critical mechanism for drought response that required trees to shift water uptake among existing roots rather than growing new roots.

Variations in leaf traits of Juniperus przewalskii from an extremely arid and cold environment.

How leaf traits vary with environmental and climatic variables in cold and arid environments is an essential issue in environmental ecology. Here, we analyzed the variations in leaf nitrogen (N) and phosphorus (P) stoichiometry and leaf dry matter content (LDMC) in Qilian juniper (Juniperus przewalskii Kom.) growing in 14 environmentally different plots on the northeastern Tibetan Plateau. The results showed that the N and P concentrations, N:P ratio and LDMC of Qilian juniper were 10.89 mg.g-1, 1.04 mg.g-1, 10.80 and 483.06 mg.g-1, respectively. The spatial coefficients of the variations in leaf N and P stoichiometry were significantly higher than the seasonal ones, and the correlations of leaf N and P concentrations with spatial variables were stronger than their correlations with the season. During the growing season, only the leaf N concentration and N:P ratio significantly increased. Soil nutrients were highly positively significantly correlated with leaf P concentrations but negatively correlated with the N:P ratio and LDMC. However, leaf N concentrations showed no significant correlations with soil nutrients. We suggest that the effects of temperature on the N concentration and LDMC were stronger than the effects of drought, while those on the P concentration and N:P ratio were weaker. Drought reduced leaf N and P concentrations and increased the N:P ratio and LDMC. In the arid region, with an increasing mean annual temperature (MAT), leaf N concentration significantly decreased, and LDMC significantly increased. In the semi-arid region, as MAT increased, leaf N and P concentrations significantly increased and LDMC and the N:P ratio significantly decreased. These opposite results supported the growth rate hypothesis that plant N and P concentrations increase while the N:P ratio and LDMC decrease as the growth rate increases.

Prolonged warming and drought modify belowground interactions for water among coexisting plants.

Understanding how climate alters plant-soil water dynamics, and its impact on physiological functions, is critical to improved predictions of vegetation responses to climate change. Here we analyzed how belowground interactions for water shift under warming and drought, and associated impacts on plant functions. In a semi-arid woodland, adult trees (piñon and juniper) and perennial grasses (blue grama) were exposed to warming and precipitation reduction. After 6 years of continuous treatment exposure, soil and plant water isotopic composition was measured to assess plant water uptake depths and community-level water source partitioning. Warming and drought modified plant water uptake depths. Under warming, contrasting changes in water sources between grasses and trees reduced belowground water source partitioning, resulting in higher interspecific competition for water. Under drought, shifts in trees and grass water sources to deeper soil layers resulted in the maintenance of the naturally occurring water source partitioning among species. Trees showed higher water stress, and reduced water use and photosynthesis in response to warming and drought. This case study demonstrates that neighboring plants shift their competitive interactions for water under prolonged warming and drought, but regardless of whether changes in moisture sources will result in increased competition among species or maintained partitioning of water resources, these competitive adaptations may easily be overridden by climate extremes.

Lack of acclimation of leaf area:sapwood area ratios in piñon pine and juniper in response to precipitation reduction and warming.

The leaf area to sapwood area ratios of trees (Al:AS) can shift to maintain homeostatic gas exchange per unit leaf area in response to climate variability. We tested the hypothesis that trees alter their Al:AS ratios in response to long-term warming and reduced precipitation in order to maintain leaf-specific gas exchange rates under more stressful conditions. Whole-tree Al:AS was measured on mature piñon pine (Pinus edulis Engelm.) and one-seed juniper (Juniperus monosperma (Engelm.) Sarg.) trees after 5 years (2012-16) of chronic exposure to increased temperature (+4.8 °C), precipitation reduction (-45%), or both simultaneously. No difference was found in Al:As among treatments for either species. Associated with this lack of shift in Al:As were large changes in pre-dawn leaf water potential and stomatal conductance, consistent with theoretical expectations of interactions between leaf and whole-tree hydraulic supply. Our results suggest that a lack of whole-tree acclimation in Al:As results in the reductions in plant gas exchange and water status associated with long-term warming and reduced precipitation in semi-arid woodlands.

Enhanced gross primary production and evapotranspiration in juniper-encroached grasslands.

Woody plant encroachment (WPE) into grasslands has been occurring globally and may be accelerated by climate change in the future. This land cover change is expected to alter the carbon and water cycles, but it remains uncertain how and to what extent the carbon and water cycles may change with WPE into grasslands under current climate. In this study, we examined the difference of vegetation indices (VIs), evapotranspiration (ET), gross primary production (GPP), and solar-induced chlorophyll fluorescence (SIF) during 2000-2010 between grasslands and juniper-encroached grasslands. We also quantitatively assessed the changes of GPP and ET for grasslands with different proportions of juniper encroachment (JWPE). Our results suggested that JWPE increased the GPP, ET, greenness-related VIs, and SIF of grasslands. Mean annual GPP and ET were, respectively, ~55% and ~45% higher when grasslands were completely converted into juniper forests under contemporary climate during 2000-2010. The enhancement of annual GPP and ET for grasslands with JWPE varied over years ranging from about +20% GPP (~+30% for ET) in the wettest year (2007) to about twice as much GPP (~+55% for ET) in the severe drought year (2006) relative to grasslands without encroachment. Additionally, the differences in GPP and ET showed significant seasonal dynamics. During the peak growing season (May-August), GPP and ET for grasslands with JWPE were ~30% and ~40% higher on average. This analysis provided insights into how and to what degree carbon and water cycles were impacted by JWPE, which is vital to understanding how JWPE and ecological succession will affect the regional and global carbon and water budgets in the future.

Reductions in tree performance during hotter droughts are mitigated by shifts in nitrogen cycling.

Climate warming should result in hotter droughts of unprecedented severity in this century. Such droughts have been linked with massive tree mortality, and data suggest that warming interacts with drought to aggravate plant performance. Yet how forests will respond to hotter droughts remains unclear, as does the suite of mechanisms trees use to deal with hot droughts. We used an ecosystem-scale manipulation of precipitation and temperature on piñon pine (Pinus edulis) and juniper (Juniperus monosperma) trees to investigate nitrogen (N) cycling-induced mitigation processes related to hotter droughts. We found that while negative impacts on plant carbon and water balance are manifest after prolonged drought, performance reductions were not amplified by warmer temperatures. Rather, increased temperatures for 5 years stimulated soil N cycling under piñon trees and modified tree N allocation for both species, resulting in mitigation of hotter drought impacts on tree water and carbon functions. These findings suggest that adjustments in N cycling are likely after multi-year warming conditions and that such changes may buffer reductions in tree performance during hotter droughts. The results highlight our incomplete understanding of trees' ability to acclimate to climate change, raising fundamental questions about the resistance potential of forests to long-term, compound climatic stresses.

Stem radial growth and water storage responses to heat and drought vary between conifers with differing hydraulic strategies.

We investigated stem radial growth and water storage dynamics of 2 conifer species differing in hydraulic carbon strategies, Juniperus monosperma and Pinus edulis, under conditions of ambient, drought (∼45% reduction in precipitation), heat (∼4.8 °C temperature increase), and the combination of drought + heat, in 2013 and 2014. Juniper maintained low growth across all treatments. Overall, the relatively isohydric piñon pine showed significantly greater growth and water storage recharge than the relatively anisohydric juniper across all treatments in the average climate year (2014) but no differences in the regionally dry year (2013). Piñon pine ceased growth at a constant predawn water potential across all treatments and at a less negative water potential threshold than juniper. Heat has a greater negative impact on piñon pines' growth and water storage than drought, whereas juniper was, in contrast, unaffected by heat but strongly impacted by drought. The whole-plant hydraulic carbon strategies, in this case captured using the isohydric/anisohydric concept, translate into alternative growth and water storage strategies under drought and heat conditions.

Is desiccation tolerance and avoidance reflected in xylem and phloem anatomy of two coexisting arid-zone coniferous trees?

Plants close their stomata during drought to avoid excessive water loss, but species differ in respect to the drought severity at which stomata close. The stomatal closure point is related to xylem anatomy and vulnerability to embolism, but it also has implications for phloem transport and possibly phloem anatomy to allow sugar transport at low water potentials. Desiccation-tolerant plants that close their stomata at severe drought should have smaller xylem conduits and/or fewer and smaller interconduit pits to reduce vulnerability to embolism but more phloem tissue and larger phloem conduits compared with plants that avoid desiccation. These anatomical differences could be expected to increase in response to long-term reduction in precipitation. To test these hypotheses, we used tridimensional synchroton X-ray microtomograph and light microscope imaging of combined xylem and phloem tissues of 2 coniferous species: one-seed juniper (Juniperus monosperma) and piñon pine (Pinus edulis) subjected to precipitation manipulation treatments. These species show different xylem vulnerability to embolism, contrasting desiccation tolerance, and stomatal closure points. Our results support the hypothesis that desiccation tolerant plants require higher phloem transport capacity than desiccation avoiding plants, but this can be gained through various anatomical adaptations in addition to changing conduit or tissue size.

Co-occurring woody species have diverse hydraulic strategies and mortality rates during an extreme drought.

From 2011 to 2013, Texas experienced its worst drought in recorded history. This event provided a unique natural experiment to assess species-specific responses to extreme drought and mortality of four co-occurring woody species: Quercus fusiformis, Diospyros texana, Prosopis glandulosa, and Juniperus ashei. We examined hypothesized mechanisms that could promote these species' diverse mortality patterns using postdrought measurements on surviving trees coupled to retrospective process modelling. The species exhibited a wide range of gas exchange responses, hydraulic strategies, and mortality rates. Multiple proposed indices of mortality mechanisms were inconsistent with the observed mortality patterns across species, including measures of the degree of iso/anisohydry, photosynthesis, carbohydrate depletion, and hydraulic safety margins. Large losses of spring and summer whole-tree conductance (driven by belowground losses of conductance) and shallower rooting depths were associated with species that exhibited greater mortality. Based on this retrospective analysis, we suggest that species more vulnerable to drought were more likely to have succumbed to hydraulic failure belowground.

Impacts of long-term precipitation manipulation on hydraulic architecture and xylem anatomy of piñon and juniper in Southwest USA.

Hydraulic architecture imposes a fundamental control on water transport, underpinning plant productivity, and survival. The extent to which hydraulic architecture of mature trees acclimates to chronic drought is poorly understood, limiting accuracy in predictions of forest responses to future droughts. We measured seasonal shoot hydraulic performance for multiple years to assess xylem acclimation in mature piñon (Pinus edulis) and juniper (Juniperus monosperma) after 3+ years of precipitation manipulation. Our treatments consisted of water addition (+20% ambient precipitation), partial precipitation-exclusion (-45% ambient precipitation), and exclusion-structure control. Supplemental watering elevated leaf water potential, sapwood-area specific hydraulic conductivity, and leaf-area specific hydraulic conductivity relative to precipitation exclusion. Shifts in allocation of leaf area to sapwood area enhanced differences between irrigated and droughted KL in piñon but not juniper. Piñon and juniper achieved similar KL under ambient conditions, but juniper matched or outperformed piñon in all physiological measurements under both increased and decreased precipitation treatments. Embolism vulnerability and xylem anatomy were unaffected by treatments in either species. Absence of significant acclimation combined with inferior performance for both hydraulic transport and safety suggests piñon has greater risk of local extirpation if aridity increases as predicted in the southwestern USA.

You'd better walk alone: Changes in forest composition affect pollination efficiency and pre-dispersal cone damage in Iberian Juniperus thurifera forests.

Changes in land-use patterns are a major driver of global environmental change. Cessation of traditional land-use practices has led to forest expansion and shifts in forest composition. Consequently, former monospecific forests maintained by traditional management are progressing towards mixed forests. However, knowledge is scarce on how the presence of other tree species will affect reproduction of formerly dominant species. We explored this question in the wind-pollinated tree Juniperus thurifera. We hypothesised that the presence of heterospecific trees would have a negative effect on cone production and on the proportion of cones attacked by specialised predators. We assessed the relative importance of forest composition on cone production, seed development and pre-dispersal cone damage on nine paired pure and mixed J. thurifera forests in three regions across the Iberian Peninsula. The effects of forest composition on crop size, cone and seed characteristics, as well as damage by pre-dispersal arthropods were tested using mixed models. Cone production was lower and seed abortion higher in mixed forests, suggesting higher pollination failure. In contrast, cone damage by arthropods was higher in pure forests, supporting the hypothesis that presence of non-host plants reduces damage rates. However, the response of each arthropod to forest composition was species-specific and the relative rates of cone damage varied depending on individual tree crops. Larger crop sizes in pure forests compensated for the higher cone damage rates, leading to a higher net production of sound seeds compared to mixed forests. This study indicates that ongoing changes in forest composition after land abandonment may impact tree reproduction.

Climate Change Increases Drought Stress of Juniper Trees in the Mountains of Central Asia.

Assessments of climate change impacts on forests and their vitality are essential for semi-arid environments such as Central Asia, where the mountain regions belong to the globally important biodiversity hotspots. Alterations in species distribution or drought-induced tree mortality might not only result in a loss of biodiversity but also in a loss of other ecosystem services. Here, we evaluate spatial trends and patterns of the growth-climate relationship in a tree-ring network comprising 33 juniper sites from the northern Pamir-Alay and Tien Shan mountain ranges in eastern Uzbekistan and across Kyrgyzstan for the common period 1935-2011. Junipers growing at lower elevations are sensitive to summer drought, which has increased in intensity during the studied period. At higher elevations, juniper growth, previously favored by warm summer temperatures, has in the recent few decades become negatively affected by increasing summer aridity. Moreover, response shifts are observed during all seasons. Rising temperatures and alterations in precipitation patterns during the past eight decades can account for the observed increase in drought stress of junipers at all altitudes. The implications of our findings are vital for the application of adequate long-term measures of ecosystem conservation, but also for paleo-climatic approaches and coupled climate-vegetation model simulations for Central Asia.

Pre-dispersal predation effect on seed packaging strategies and seed viability.

An increased understanding of intraspecific seed packaging (i.e. seed size/number strategy) variation across different environments may improve current knowledge of the ecological forces that drive seed evolution in plants. In particular, pre-dispersal seed predation may influence seed packaging strategies, triggering a reduction of the resources allocated to undamaged seeds within the preyed fruits. Assessing plant reactions to pre-dispersal seed predation is crucial to a better understanding of predation effects, but the response of plants to arthropod attacks remains unexplored. We have assessed the effect of cone predation on the size and viability of undamaged seeds in populations of Juniperus thurifera with contrasting seed packaging strategies, namely, North African populations with single-large-seeded cones and South European populations with multi-small-seeded cones. Our results show that the incidence of predation was lower on the single-large-seeded African cones than on the multi-small-seeded European ones. Seeds from non-preyed cones were also larger and had a higher germination success than uneaten seeds from preyed cones, but only in populations with multi-seeded cones and in cones attacked by Trisetacus sp., suggesting a differential plastic response to predation. It is possible that pre-dispersal seed predation has been a strong selective pressure in European populations with high cone predation rates, being a process which maintains multi-small-seeded cones and empty seeds as a strategy to save some seeds from predation. Conversely, pre-dispersal predation might not have a strong effect in the African populations with single-large-seeded cones characterized by seed germination and filling rates higher than those in the European populations. Our results indicate that differences in pre-dispersal seed predators and predation levels may affect both selection on and intraspecific variation in seed packaging.