Chemical Characteristic and Sensory Evaluation of Biscuit Prepared from Wheat and Aleppo Pine Seeds Flour.

This study aimed to explore the potential use of Aleppo pine seed flour as a nutritious ingredient in biscuit production. Aleppo pine seed flour was blended with wheat flour to create biscuits with varying proportions: 15%, 30%, and 45% Aleppo pine seed flour. The analysis of the chemical composition of the biscuits revealed that increasing the proportion of Aleppo pine seed flour led to higher levels of protein, fat, and ash, while the carbohydrate content decreased. Sensory evaluation showed that biscuits with 15% Aleppo pine seed flour and 85% wheat flour had favorable characteristics in terms of color, flavor, crispness, and overall appeal. These findings indicate that incorporating Aleppo pine seed flour into wheat flour can enhance the nutritional profile of biscuits, offering higher protein, fat, and mineral content. The study suggests that a blend containing 15% Aleppo pine seed flour is optimal for producing biscuits with desirable sensory properties, making it a promising alternative ingredient for healthier biscuit formulations. Overall, this research highlights the potential of Aleppo pine seed flour to improve the nutritional quality of biscuits without compromising their sensory qualities, paving the way for its use in health-conscious baking practices.

Elaboration of an innovative plant biomaterial for its valorization in the treatment of wastewater.

The global scientific community is deeply concerned about the deterioration of water quality resulting from the release of industrial effluents. This issue is of utmost importance as it serves to safeguard the environment and combat water pollution. The objective of this work is to elaborate a biomaterial of vegetable origin, based on the twigs of Aleppo pine, and to use it as an abundant and less expensive material for the treatment of wastewater. For this reason, the twigs were treated physically to get the powder called biomaterial FPA (Aleppo pine fiber), which was characterized by physicochemical, and spectroscopic analyses namely scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction (XRD). The crystallinity index of FPA was evaluated by the peak height method. The findings indicate that the FPA powder has an acidic nature, exhibiting a porous structure that promotes the adsorption and binding of molecules. Additionally, it has a zero charge point of 5.8 and a specific surface area of 384 m2.g-1. It is primarily composed of hydroxyl, carboxyl, and amine functional groups, along with mineral compounds and organic compounds, including cellulose and other mineral elements such as Ca, Mg, Fe, Na, P, Al, K, Ni, and Mo. Combining these characteristics, FPA biomaterial has considerable potential for use as an effective adsorbent biomaterial for various wastewater pollutants. Its abundance and relatively low cost make it an attractive solution to the growing challenges of water pollution worldwide.

Transpiration Dynamics of Esparto Grass (Macrochloa tenacissima (L.) Kunth) in a Semi-Arid Mediterranean Climate: Unraveling the Impacts of Pine Competition.

Macrochloa tenacissima (M. tenacissima), or esparto, is a perennial tussock grass that coexists with Pinus halepensis (P. halepensis) in semi-arid Mediterranean woodlands. This research was carried out to explore diurnal transpiration at leaf level in esparto grass under different levels of pine-esparto competition and in contrasting environmental soil water conditions. The measurement period spanned from the summer of 2020 to the spring of 2021. The relationship between transpiration and competition was conducted in open and closed P. halepensis stands, and the type of leaf (green, senescent) and the maturity of the esparto grass were taken into account. We observed a higher control of transpiration in green leaves, and the correlations between the transpiration and pine competition were noted exclusively in this type of leaf. Our results demonstrated a significant impact of pine competitors (closed stands) on the transpiration of esparto grass, particularly during seasons characterized by scenarios of high water demand: the summer drought period and the commencement of the growing and flowering period (spring). Furthermore, our findings revealed a greater response to transpiration in mature bushes compared to young ones under severe water stress, indicating a higher adaptation to drought by esparto as it ages. Although our results confirmed that PAR increased transpiration in all seasons and in both stands, which is attributable to the heliophilia of esparto grass, the site effects on transpiration could also be attributable to competition for water, especially during periods of drought. These results may have important implications for the dynamics and management of these semi-arid mixed woodlands, as well as the planning of reforestation programs aimed at restoring esparto grass formations.

Bacillus subtilis SPB1 lipopeptide biosurfactant: antibacterial efficiency against the phytopathogenic bacteria Agrobacterium tumefaciens and compared production in submerged and solid state fermentation systems.

Bacillus subtilis SPB1 derived biosurfactants (BioS) proved its bio-control activity against Agrobacterium tumefaciens using tomato plant. Almost 83% of disease symptoms triggered by Agrobacterium tumefaciens were reduced. Aiming potential application, we studied lipopeptide cost-effective production in both fermentations systems, namely the submerged fermentation (SmF) and the solid-state fermentation (SSF) as well as the use of Aleppo pine waste and confectionery effluent as cheap substrates. Optimization studies using Box-Behnken (BB) design followed by the analysis with response surface methodology were applied. When using an effluent/sea water ratio of 1, Aleppo pine waste of 14.08 g/L and an inoculum size of 0.2, a best production yield of 17.16 ± 0.91 mg/g was obtained for the SmF. While for the SSF, the best production yield of 27.59 ± 1.63 mg/g was achieved when the value of Aleppo pine waste, moisture, and inoculum size were, respectively, equal to 25 g, 75%, and 0.2. Hence, this work demonstrated the superiority of SSF over SmF.

Proteomic and Metabolic Analysis of Pinus halepensis Mill. Embryonal Masses Induced under Heat Stress.

Understanding the physiological and molecular adjustments occurring during tree stress response is of great importance for forest management and breeding programs. Somatic embryogenesis has been used as a model system to analyze various processes occurring during embryo development, including stress response mechanisms. In addition, "priming" plants with heat stress during somatic embryogenesis seems to favor the acquisition of plant resilience to extreme temperature conditions. In this sense, Pinus halepensis somatic embryogenesis was induced under different heat stress treatments (40 °C for 4 h, 50 °C for 30 min, and 60 °C for 5 min) and its effects on the proteome and the relative concentration of soluble sugars, sugar alcohols and amino acids of the embryonal masses obtained were assessed. Heat severely affected the production of proteins, and 27 proteins related to heat stress response were identified; the majority of the proteins with increased amounts in embryonal masses induced at higher temperatures consisted of enzymes involved in the regulation of metabolism (glycolysis, the tricarboxylic acid cycle, amino acid biosynthesis and flavonoids formation), DNA binding, cell division, transcription regulation and the life-cycle of proteins. Finally, significant differences in the concentrations of sucrose and amino acids, such as glutamine, glycine and cysteine, were found.

In situ, direct observation of seasonal embolism dynamics in Aleppo pine trees growing on the dry edge of their distribution.

Xylem embolism impairs hydraulic conductivity in trees and drives drought-induced mortality. While embolism has been monitored in vivo in potted plants, and research has revealed evidence of embolism in field-grown trees, continuous in situ monitoring of cavitation in forests is lacking. Seasonal patterns of embolism were monitored in branchlets of Aleppo pine (Pinus halepensis) trees growing in a dry Mediterranean forest. Optical visualization (OV) sensors were installed on terminal branches, in addition to monthly sampling for micro computed tomography scans. We detected 208 cavitation events among four trees, which represented an embolism increase from zero to c. 12% along the dry season. Virtually all the cavitation events occurred during daytime hours, with 77% occurring between 10:00 and 17:00 h. The probability for cavitation in a given hour increased as vapor pressure deficit (VPD) increased, up to a probability of 42% for cavitation when VPD > 5 kPa. The findings uniquely reveal the instantaneous environmental conditions that lead to cavitation. The increased likelihood of cavitation events under high VPD in water-stressed pines is the first empirical support for this long hypothesized relationship. Our observations suggest that low levels of embolism are common in Aleppo pine trees at the dry edge of their distribution.

Heat Stress in Pinus halepensis Somatic Embryogenesis Induction: Effect in DNA Methylation and Differential Expression of Stress-Related Genes.

In the current context of climate change, plants need to develop different mechanisms of stress tolerance and adaptation to cope with changing environmental conditions. Temperature is one of the most important abiotic stresses that forest trees have to overcome. Recent research developed in our laboratory demonstrated that high temperatures during different stages of conifer somatic embryogenesis (SE) modify subsequent phases of the process and the behavior of the resulting ex vitro somatic plants. For this reason, Aleppo pine SE was induced under different heat stress treatments (40 °C for 4 h, 50 °C for 30 min, and 60 °C for 5 min) in order to analyze its effect on the global DNA methylation rates and the differential expression of four stress-related genes at different stages of the SE process. Results showed that a slight decrease of DNA methylation at proliferating embryonal masses (EMs) can correlate with the final efficiency of the process. Additionally, different expression patterns for stress-related genes were found in EMs and needles from the in vitro somatic plants obtained; the DEHYDRATION INDUCED PROTEIN 19 gene was up-regulated in response to heat at proliferating EMs, whereas HSP20 FAMILY PROTEIN and SUPEROXIDE DISMUTASE [Cu-Zn] were down-regulated in needles.

Assessing model performance via the most limiting environmental driver in two differently stressed pine stands.

Climate change will impact forest productivity worldwide. Forecasting the magnitude of such impact, with multiple environmental stressors changing simultaneously, is only possible with the help of process-based models. In order to assess their performance, such models require careful evaluation against measurements. However, direct comparison of model outputs against observational data is often not reliable, as models may provide the right answers due to the wrong reasons. This would severely hinder forecasting abilities under unprecedented climate conditions. Here, we present a methodology for model assessment, which supplements the traditional output-to-observation model validation. It evaluates model performance through its ability to reproduce observed seasonal changes of the most limiting environmental driver (MLED) for a given process, here daily gross primary productivity (GPP). We analyzed seasonal changes of the MLED for GPP in two contrasting pine forests, the Mediterranean Pinus halepensis Mill. Yatir (Israel) and the boreal Pinus sylvestris L. Hyytiälä (Finland) from three years of eddy-covariance flux data. Then, we simulated the same period with a state-of-the-art process-based simulation model (LandscapeDNDC). Finally, we assessed if the model was able to reproduce both GPP observations and MLED seasonality. We found that the model reproduced the seasonality of GPP in both stands, but it was slightly overestimated without site-specific fine-tuning. Interestingly, although LandscapeDNDC properly captured the main MLED in Hyytiälä (temperature) and in Yatir (soil water availability), it failed to reproduce high-temperature and high-vapor pressure limitations of GPP in Yatir during spring and summer. We deduced that the most likely reason for this divergence is an incomplete description of stomatal behavior. In summary, this study validates the MLED approach as a model evaluation tool, and opens up new possibilities for model improvement.

Effective Defense of Aleppo Pine Against the Giant Scale Marchalina hellenica Through Ecophysiological and Metabolic Changes.

Aleppo pine (Pinus halepensis) is widely distributed in the Mediterranean region and in other areas of the world, where it has been introduced due to its adaptive capacity to xerothermic conditions. The giant pine scale Marchalina hellenica often infests Aleppo pine, as well as other pines, in several southeastern European countries, causing pine declines. When combined with the expected intensified heat and drought events in eastern Mediterranean, the impact of this biotic parameter on the host pines may be exacerbated. The importance of understanding the defense mechanisms of Aleppo pine is emphasized by the recent invasion of the pine scale in new regions, like Australia, lacking the insect's natural enemies, where more intense negative effects on pine species may occur. To date, Aleppo pine's physiological responses to the infestation by M. hellenica are largely unknown. This study aimed at assessing the responses of Aleppo pine to the giant pine scale attack, both on an ecophysiological and a metabolic level. For this purpose, gas exchange, needle water status, and carbon and nitrogen content were measured during 1 year on healthy and infested adult trees. M etabolic profiling of Aleppo pine needles was also performed before, during, and after the high feeding activity of the insect. The maintenance of stable relative water content, δ13C signatures, and chlorophyll fluorescence in the needles of infested pines indicated that infestation did not induce drought stress to the host pines. At the peak of infestation, stomatal closure and a pronounced reduction in assimilation were observed and were associated with the accumulation of sugars in the needles, probably due to impaired phloem loading. At the end of the infestation period, tricarboxylic acids were induced and phenolic compounds were enhanced in the needles of infested pines. These metabolic responses, together with the recovery of photosynthesis after the end of M. hellenica intense feeding, indicate that in the studied region and under the current climate, Aleppo pine is resilient to the infestation by the giant pine scale. Future research should assess whether these promising defense mechanisms are also employed by other host pines, particularly in regions of the world recently invaded by the giant pine scale, as well as under more xerothermic regimes.

Intraspecific responses to climate reveal nonintuitive warming impacts on a widespread thermophilic conifer.

Many ecologically important forest trees from dry areas have been insufficiently investigated for their ability to adapt to the challenges posed by climate change, which hampers the implementation of mitigation policies. We analyzed 14 common-garden experiments across the Mediterranean which studied the widespread thermophilic conifer Pinus halepensis and involved 157 populations categorized into five ecotypes. Ecotype-specific tree height responses to climate were applied to projected climate change (2071-2100 ad), to project potential growth patterns both locally and across the species' range. We found contrasting ecotypic sensitivities to annual precipitation but comparatively uniform responses to mean temperature, while evidence of local adaptation for tree height was limited to mesic ecotypes. We projected intriguing patterns of response range-wide, implying either height inhibition or stimulation of up to 75%, and deduced that the ecotype currently experiencing more favorable (wetter) conditions will show the largest inhibition. Extensive height reductions can be expected for coastal areas of France, Greece, Spain and northern Africa. Our findings underline the fact that intraspecific variations in sensitivity to precipitation must be considered when projecting tree height responses of dry forests to future climate. The ecotype-specific projected performances call for management activities to ensure forest resilience in the Mediterranean through, for example, tailored deployment strategies.

Use of Autochthonous Lactobacilli to Increase the Safety of Zgougou.

Seeds of Pinus halepensis are used for preparing zgougou, a spontaneously fermented matrix giving juice and seeds debris, consumed in many Arabian countries, including Tunisia. In the same way as all the food processes based on spontaneous fermentation, zgougou hides health risks due to eventual pathogenic microorganisms and derived toxins. This study aimed at investigating the effect of the use of autochthonous Lactobacillus paraplantarum A1 and Lactobacillus plantarum A2, as fermentation starters, on the microbiological characteristics, profiles of volatile organic compounds (VOC), antibacterial and antioxidant activities of juice and seeds debris from zgougou. The starter lactobacilli inhibited undesired bacteria (e.g., Enterobacter and Aeromonas) and coccus-shaped lactic acid bacteria, as shown by culture-dependent and-independent methods. The inhibitory effect was more evident in juice than in seeds debris. Some VOC (ethanol, acetoin, phenol,2-methoxy and caryophyllene) were present at higher concentrations in juice and seeds obtained upon spontaneous fermentation, compared to the samples deriving from fermentation with lactobacilli. The latter samples were characterized by higher concentrations of acetic acid, decane, 1-nonanol, bornyl acetate and bornyl formate. In addition, they showed a wider spectrum of antibacterial activity than spontaneously fermented juice and seeds. The use of autochthonous lactobacilli did not relevantly affect the antioxidant activity of zgougou. When juice from lactobacilli-driven fermentation was used to prepare a traditional Tunisian pudding ("Assidat-Zgougou"), it improved color and odor with respect to the pudding containing juice from spontaneous fermentation. This study showed that the use, at laboratory scale, of autochthonous lactobacilli is a feasible biotechnological tool to outgrow undesired bacteria, thus improving the safety of zgougou juice. Future studies should be undertaken to confirm the observed benefits at industrial scale.

Allelopathic effects of volatile organic compounds released from Pinus halepensis needles and roots.

The Mediterranean region is recognized as a global biodiversity hotspot. However, over the last decades, the cessation of traditional farming in the north part of the Mediterranean basin has given way to strong afforestation leading to occurrence of abandoned agricultural lands colonized by pioneer expansionist species like Pinus halepensis. This pine species is known to synthesize a wide range of secondary metabolites, and previous studies have demonstrated strong allelopathic potentialities of its needle and root leachates. Pinus halepensis is also recognized to release significant amounts of volatile organic compounds (VOC) with potential allelopathic effects that have never been investigated. In this context, the objectives of the present study were to improve our knowledge about the VOC released from P. halepensis needles and roots, determine if these VOC affect the seed germination and root growth of two herbaceous target species (Lactuca sativa and Linum strictum), and evaluate if soil microorganisms modulate the potential allelopathic effects of these VOC. Thirty terpenes were detected from both, needle and root emissions with ß-caryophyllene as the major volatile. Numerous terpenes, such as ß-caryophyllene, δ-terpinene, or α-pinene, showed higher headspace concentrations according to the gradient green needles < senescent needles < needle litter. Seed germination and root growth of the two target species were mainly reduced in presence of P. halepensis VOC. In strong contrast with the trend reported with needle leachates in literature, we observed an increasing inhibitory effect of P. halepensis VOC with the progress of needle physiological stages (i.e., green needle < senescent needle < needle litter). Surprisingly, several inhibitory effects observed on filter paper were also found or even amplified when natural soil was used as a substrate, highlighting that soil microorganisms do not necessarily limit the negative effects of VOC released by P. halepensis on herbaceous target species.

Quantification of leaf-scale light energy allocation and photoprotection processes in a Mediterranean pine forest under extensive seasonal drought.

Photoprotection strategies in a Pinus halepensis Mill. forest at the dry timberline that shows sustained photosynthetic activity during 6-7 month summer drought were characterized and quantified under field conditions. Measurements of chlorophyll fluorescence, leaf-level gas exchange and pigment concentrations were made in both control and summer-irrigated plots, providing the opportunity to separate the effects of atmospheric from soil water stress on the photoprotection responses. The proportion of light energy incident on the leaf surface ultimately being used for carbon assimilation was 18% under stress-free conditions (irrigated, winter), declining to 4% under maximal stress (control, summer). Allocation of absorbed light energy to photochemistry decreased from 25 to 15% (control) and from 50% to 30% (irrigated) between winter and summer, highlighting the important role of pigment-mediated energy dissipation processes. Photorespiration or other non-assimilatory electron flow accounted for 15-20% and ~10% of incident light energy during periods of high and low carbon fixation, respectively, representing a proportional increase in photochemical energy going to photorespiration in summer but a decrease in the absolute amount of photorespiratory CO2 loss. Resilience of the leaf photochemical apparatus was expressed in the complete recovery of photosystem II (PSII) efficiency (ΦPSII) and relaxation of the xanthophyll de-epoxidation state on the diurnal cycle throughout the year, and no seasonal decrease in pre-dawn maximal PSII efficiency (Fv/Fm). The response of CO2 assimilation and photoprotection strategies to stomatal conductance and leaf water potential appeared independent of whether stress was due to atmospheric or soil water deficits across seasons and treatments. The range of protection characteristics identified provides insights into the relatively high carbon economy under these dry conditions, conditions that are predicted for extended areas in the Mediterranean and other regions due to global climate change.

Geographic Differences in Element Accumulation in Needles of Aleppo Pines (Pinus halepensis Mill.) Grown in Mediterranean Region.

Pine needles are widely used as bio-indicators due to their worldwide distribution and the ease of sample collection. In contrast to deciduous trees, conifers offer the possibility of monitoring long-term exposure through older needles. Pinus halepensis Miller is a pine species native to the Mediterranean region, which has been used for restoration activities in arid and semiarid areas leading to vast spatial expansion. Needles from pine trees collected in the southeastern to northwestern extension of Croatia's coastal area at twelve sampling sites were analysed for twenty-one metals and metalloids. Statistical evaluation of the obtained data revealed significant differences for Al, As, B, Ba, Ca, Cr, Fe, K, Mg, Mn, Na, Se, and Sr between the different regions. Needles from trees growing on islands did not show elevated levels of Mg and/or Na as a result of the sea spray influence. The differences in metal accumulation are supposed to be linked to the environmental conditions at the respective sampling site, since the species was the same everywhere. By comparing the elemental contents of the soil those of with needles, it can be clearly seen, that the root as well as the foliar uptake contribute to the final amount.

[Variation in Thaumetopoea pityocampa infestation rate of Aleppo pine: Effect on dendrometric parameters in the Djelfa region forests (Saharan Atlas, Algeria)]. / Variation du taux d'infestation par Thaumetopoea pityocampa du pin d'Alep : effet sur les paramètres dendrométriques dans les forêts de la région de Djelfa (Atlas saharien, Algérie).

The forests of the Saharan Atlas represent the southern edge of the natural range of the Aleppo pine (Pinus halepensis Mill.). These are exposed to climatic stress in relation to drought and attacks of the pine processionary moth (Thaumetopoea pityocampa Schiff., denoted by CP in the French text). The purpose of our work is to study the variation of the infestation rate by the pine processionary moth and its density with climatic factors and dendrometric parameters. The infestation rate (7.82%) by the pine processionary moth and its density (2.74±2.61nests/tree) were lower in our pine forests than in the northern ones. It appeared that the attacks of the moth increase mainly with the number of days of frost and heatwave. The dendrometric parameters were negatively correlated with the moth's density per tree. On the other hand, the latter was positively correlated with the defoliation rate. The results were discussed in a biogeographic framework in the light of the current knowledge of the distribution of the pine processionary moth in the Aleppo pine, in relation with the climatic conditions.

Temporal characterisation of soil-plant natural recovery related to fire severity in burned Pinus halepensis Mill. forests.

Despite Mediterranean ecosystems' high resilience to fire, both climate and land use change, and alterations in fire regimes increase their vulnerability to fire by affecting the long-term natural recovery of ecosystem services. The objective of this work is to study the effects of fire severity on biochemical soil indicators, such as chemical composition or enzymatic activity, related to time after fire and natural vegetation recovery (soil-plant interphase). Soil samples from three wildfires occurring 3, 15 and 21 years ago were taken in the south-eastern Iberian Peninsula (semiarid climate). Sampling included three fire severity levels in naturally regenerated (and changing to shrublands) Pinus halepensis Mill. forests. In the short-term post-fire period, phosphorus concentration, electrical conductivity and urease activity were positively linked to fire severity, and also influenced ß-glucosidade activity in a negative relationship. During the 15-21-year post-fire period, the effects related to medium-high fire severity were negligible and soil quality indicators were linked to natural regeneration success. The results showed that most soil properties recovered in the long term after fire (21 years). These outcomes will help managers and stakeholders to implement management tools to stabilise soils and to restore burned ecosystems affected by medium-high fire severity. Such knowledge can be considered in adaptive forest management to reduce the negative effects of wildfires and desertification, and to improve the resilience of vulnerable ecosystems in a global change scenario.

Short-term effects of spring prescribed burning on the understory vegetation of a Pinushalepensis forest in Northeastern Spain.

Since the 1970s, fire regimes have been modified in the Northern Mediterranean region due to profound landscape changes mostly driven by socioeconomic factors, such as rural abandonment and large-scale plantations. Both fuel accumulation and the increasing vegetation spatial continuity, combined with the expansion of the wildland-urban interface, have enhanced fire risk and the occurrence of large wildfires. This situation will likely worsen under the projected aridity increase resulting from climate change. Higher fire recurrences, in particular, are expected to cause changes in vegetation composition or structure and affect ecosystems' resilience to fire, which may lead to further land degradation. Prescribed burning is a common fuel reduction technique used for fire prevention, but for conservation and restoration purposes as well. It is still poorly accepted in the Mediterranean region since constrained by critical knowledge gaps about, in particular, its effects on the ecosystems (soil, vegetation). We studied the short-term (10months) effects on the understory vegetation of a spring prescribed burning conducted in a Pinushalepensis forest in Mediterranean climate (Northeastern Spain). Our results show that the understory plant community recovered after the burning without short term significant changes in either species richness, diversity, or floristic composition. Most vegetation structural characteristics were modified though. The burning strongly reduced shrub height, shrub and herbaceous percentage covers, and aerial shrub phytomass; especially its living fine fraction, thus resulting in a less flammable community. The treatment proved to be particularly effective for the short term control of Ulexparviflorus, a highly flammable seeder species. Moreover, the strong reduction of seeder shrubs frequency in relation to resprouters' likely promoted the resilience to fire of this plant community. From a fuel-oriented perspective, the burning caused a strong reduction of spatial continuity and surface fuel loads, leading to a less fire-prone fuel complex.

Drought Tolerance in Pinus halepensis Seed Sources As Identified by Distinctive Physiological and Molecular Markers.

Drought is one of the main constraints determining forest species growth, survival and productivity, and therefore one of the main limitations for reforestation or afforestation. The aim of this study is to characterize the drought response at the physiological and molecular level of different Pinus halepensis (common name Aleppo pine) seed sources, previously characterized in field trials as drought-sensitive or drought-tolerant. This approach aims to identify different traits capable of predicting the ability of formerly uncharacterized seedlings to cope with drought stress. Gas-exchange, water potential, photosynthetic pigments, soluble sugars, free amino acids, glutathione and proteomic analyses were carried out on control and drought-stressed seedlings in greenhouse conditions. Gas-exchange determinations were also assessed in field-planted seedlings in order to validate the greenhouse experimental conditions. Drought-tolerant seed sources presented higher values of photosynthetic rates, water use efficiency, photosynthetic pigments and soluble carbohydrates concentrations. We observed the same pattern of variation of photosynthesis rate and maximal efficiency of PSII in field. Interestingly drought-tolerant seed sources exhibited increased levels of glutathione, methionine and cysteine. The proteomic profile of drought tolerant seedlings identified two heat shock proteins and an enzyme related to methionine biosynthesis that were not present in drought sensitive seedlings, pointing to the synthesis of sulfur amino acids as a limiting factor for drought tolerance in Pinus halepensis. Our results established physiological and molecular traits useful as distinctive markers to predict drought tolerance in Pinus halepensis provenances that could be reliably used in reforestation programs in drought prone areas.

Soil moisture and its role in growth-climate relationships across an aridity gradient in semiarid Pinus halepensis forests.

In Mediterranean areas with limited availability of water, an accurate knowledge of growth response to hydrological variables could contribute to improving management and stability of forest resources. The main goal of this study is to assess the temporal dynamic of soil moisture to better understand the water-growth relationship of Pinus halepensis forests in semiarid areas. The estimates of modelled soil moisture and measured tree growth were used at four sites dominated by afforested Pinus halepensis Mill. in south-eastern Spain with 300 to 609mm mean annual precipitation. Firstly, dendrochronological samples were extracted and the widths of annual tree rings were measured to compute basal area increments (BAI). Secondly, soil moisture was estimated over 20 hydrological years (1992-2012) by means of the HYDROBAL ecohydrological model. Finally, the tree growth was linked, to mean monthly and seasonal temperature, precipitation and soil moisture. Results depict the effect of soil moisture on growth (BAI) and explain 69-73% of the variance in semiarid forests, but only 51% in the subhumid forests. This highlights the fact that that soil moisture is a suitable and promising variable to explain growth variations of afforested Pinus halepensis in semiarid conditions and useful for guiding adaptation plans to respond pro-actively to water-related global challenges.

Missing Rings in Pinus halepensis - The Missing Link to Relate the Tree-Ring Record to Extreme Climatic Events.

Climate predictions for the Mediterranean Basin include increased temperatures, decreased precipitation, and increased frequency of extreme climatic events (ECE). These conditions are associated with decreased tree growth and increased vulnerability to pests and diseases. The anatomy of tree rings responds to these environmental conditions. Quantitatively, the width of a tree ring is largely determined by the rate and duration of cell division by the vascular cambium. In the Mediterranean climate, this division may occur throughout almost the entire year. Alternatively, cell division may cease during relatively cool and dry winters, only to resume in the same calendar year with milder temperatures and increased availability of water. Under particularly adverse conditions, no xylem may be produced in parts of the stem, resulting in a missing ring (MR). A dendrochronological network of Pinus halepensis was used to determine the relationship of MR to ECE. The network consisted of 113 sites, 1,509 trees, 2,593 cores, and 225,428 tree rings throughout the distribution range of the species. A total of 4,150 MR were identified. Binomial logistic regression analysis determined that MR frequency increased with increased cambial age. Spatial analysis indicated that the geographic areas of south-eastern Spain and northern Algeria contained the greatest frequency of MR. Dendroclimatic regression analysis indicated a non-linear relationship of MR to total monthly precipitation and mean temperature. MR are strongly associated with the combination of monthly mean temperature from previous October till current February and total precipitation from previous September till current May. They are likely to occur with total precipitation lower than 50 mm and temperatures higher than 5°C. This conclusion is global and can be applied to every site across the distribution area. Rather than simply being a complication for dendrochronology, MR formation is a fundamental response of trees to adverse environmental conditions. The demonstrated relationship of MR formation to ECE across this dendrochronological network in the Mediterranean basin shows the potential of MR analysis to reconstruct the history of past climatic extremes and to predict future forest dynamics in a changing climate.