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Pairwise trade-offs among the main dimensions of life history of plants, namely growth, reproduction and defence, have repeatedly tested with inconclusive results. Relationships between life history dimensions mediate the evolution of reproductive strategies, such as masting, as they are key to understand resource allocation, predator satiation and the response of plants to resource and climate variation. We carried out two long-term field experiments with 81 clonally replicated individuals from 6 genotypes to assess their response to increased resource availability (fertilization and control treatments) and simulated herbivory (low intensity mechanical wounding, high intensity mechanical wounding, needle clipping, jasmonate application and control treatments). The data set comprises 6 csv files containing between 6 and 19 variables each, with data on tree (ramet) growth (DBH), reproductive effort and performance (female and male strobili production, number of ripe cones, number of healthy seeds, total number of seeds and mean dry weight of seeds), nutrient content (P, C, N and C/N), production of defensive compounds (total resin and total phenolics) and realized herbivory damage (damage level produced by Pissodes castaneus) during six years (four reproductive cohorts).

Pairwise trade-offs among the main dimensions of life history of plants, namely growth, reproduction and defence, have been hypothesized and repeatedly tested. Experimental evidences, however, are far from conclusive, probably as a consequence of different methodological approaches, but also of an underlying multivariate complexity that is not considered in pairwise analyses. We carried out two long-term field experiments with 81 ramets clonally replicated from 6 genotypes of maritime pine (Pinus pinaster) to assess the allocation to growth, reproduction and defence in response to increased resource availability (fertilization and control treatments) and simulated herbivory (low intensity mechanical wounding, high intensity mechanical wounding, needle clipping, jasmonate application and control treatments). Fertilization increased tree growth and N and P concentration, and altered the production of resin and polyphenolics in complex ways, contingent on time and tissue. A natural event of infestation by Pissodes castaneus was more intense on fertilized trees. Fertilization increased both reproductive effort and output in terms of quantity (number of seeds) but not quality (dry weight of seeds). Needle clipping and methyl-jasmonate, but not mechanical wounding, increased the concentration of needle resin and phenols shortly after treatment, while concentrations in the phloem remain unaltered. Damage by P. castaneus did not differ across induction treatments. Generally, needle clipping had the strongest effect on reproductive variables, and mechanical wounding the mildest, with a high variability on intensity and direction of effects. Needle clipping decreased tree growth, while the rest of herbivory treatments did increase it. Complex interactions among life-history dimensions arose, showing that pairwise approaches are too simplistic for unravelling these complexities. This three-way relationship imposes new environmental and physiological constraints to the evolutio

The potential of pine resin as a renewable non-timber product is furthering the socioeconomic relevance of the resin industry in Europe, where maritime pine (Pinus pinaster Ait.) is the main tapped species. Characterizing maritime pine genetic resources in terms of resin yield potential and its covariation with tree growth is crucial to implement efficient tree breeding programs aiming to maximize resin yield without negatively impact tree growth. We explored genetic variation in resin yield and tree growth among provenance regions, populations within regions, and families of maritime pine. We used a progeny trial located at Central Spain to estimate additive genetic variation and heritability of resin yield, and a common garden located at Northwest Spain to explore variation among provenance regions and populations within regions. Resin yield was assessed in both trials with and without sulphuric-based stimulant paste. Association between growth and resin yield at different levels (phenotypes, families, populations and provenance regions) was also explored. Growth and resin yield were highly variable within populations, with narrow-sense heritability estimates being particularly high for resin yield (hi2 = 0.49). Growth varied significantly across provenance regions and populations within regions, while resin yield was only variable among provenance regions. Genetic variation in resin yield was similar in trees with and without stimulant paste. We did not find evidence of genetic correlations between growth and resin yield. However, at the phenotype level resin yield and growth were significantly correlated, although the sign of such association switched between trials. Across provenance regions, growth and resin yield where negatively correlated. Altogether, the high narrow-sense heritability estimate, the highly consistent genetic variation and the lack of negative genetic correlation with growth evidence the opportunity for implementing breeding programs to incr

Persistence of tree populations in the face of global change relies on their capacity to respond to biotic and abiotic stressors through plastic or adaptive changes. Genetic adaptation will depend on the additive genetic variation within populations and the heritability of traits related to stress tolerance. Because traits can be genetically linked, selective pressure acting on one trait may lead to correlated responses in other traits.

Quantifying intraspecific genetic variation and phenotypic plasticity of traits involved in drought tolerance is essential to forecast forest tree vulnerability to climate change. Tree ring analysis was applied to retrospectively assess the resistance, recovery and resilience of a Mediterranean pine in the face of extreme climate episodes. We combined a dendrochronological approach with the analysis of common-garden tests to disentangle genetic, environment and genetic-by-environment effects in growth responses of 10 P. pinaster populations to two extreme climatic events. Trees were 43 years old and had experienced two intense droughts, in 1995 and 2005. Drought events caused drastic reductions in secondary growth, but trees showed high capacity to recover pre-drought growth rates. The differences in the characteristics of the two drought events and the environmental distance between sites strongly modulated maritime pine responses to extreme droughts. However, a common among-population signal across sites and events was detected in the drought response strategy. Among-population variation in response to extreme droughts was evident for the resistance and recovery components, two strategies that appeared to trade-off between each other. Populations from Atlantic climates showed higher resistance but lower recovery capacity, whereas Mediterranean origins prioritised recovery over resistance. Mediterranean populations showed a more conservative strategy that indicated an adaptive advantage under water stress, reflected in greater long-term survival. The abovementioned relationships were clear in the site where the impact of the drought events was strongest, but not in the more favourable site. Differences in relationships between sites reflect that strategies of populations to cope with drought are strongly context dependent. Based on these results, we infer that future extreme droughts will differentially affect P. pinaster populations across the natural range of the

The ability of trees to overcome the new environmental conditions will depend on the extant intraspecific variation in traits with adaptive value. Although genetic variation in life history traits related to productivity and abiotic stress has been better studied, little is still known about defensive traits. Resin ducts are the structures that store and produce the oleoresin in conifers, and because their role in resistance against biotic stress, they are a valuable proxy to study the defensive investment. In the current study, we explored the inducibility and the genetic variation of resin ducts among populations in Pinus pinaster and evaluated whether this variation is explained by the abiotic environment from the origin of populations. To meet our objectives, we used a clonal genetic collection of P. pinaster in a greenhouse experiment, including 79 clonally-replicated genotypes from 10 populations covering the species distribution range. We measured resin duct features in phloem and xylem in one clonal replicate and their inducibility in the other. Genetic variation among populations was explored by fitting mixed models, and correlation analysis were performed between resin duct features and climatic variables at the site of origin of populations after accounting for the population relatedness, which was quantified by genotyping 126 SNPs in the experimental material. We found that intraspecific genetic variation in the phenotypic expression of resin ducts was explained by the climate at origin. Moreover, we found that the differentiation of resin ducts in the xylem was inducible, although inducibility did not varied among populations.

Breeding for resistance to forest pests and pathogens is emerging as a promising tool for minimising the impact of the increasing biotic threats that our forests are experiencing as a consequence of global change. Efficient phenotyping protocols of resistance are urgently needed. Here we present the results of two experiments aimed to determine whether the variation in resistance to the pine weevil Hylobius abietis, a harmful pest of European conifers, can be inferred by nondestructive bioassays using excised plant material collected in forest genetic trials. Weevil damage and amount of nonvolatile resin induced by weevil feeding were assessed in young trees and in branches of adult trees using several phenotyping procedures (bioassays using either living trees, excised plant material and cut stem twigs) on four pine species (Pinus pinaster, P. radiata, P. sylvestris and P. pinea). Half of the plants were previously induced with methyl jasmonate (MJ), a treatment that is known to affect resistance to the pine weevil. In Experiment 1, living and excised plants showed parallel results: MJ treatment significantly reduced weevil damage, and saplings responded to weevil damage locally increasing the nonvolatile resin (NVR) in the stems proportionally to the damage suffered. This response was, however, slightly lower in excised than in living saplings. On the contrary, patterns of weevil feeding on stem twigs completely departed from those observed in living and excised seedlings. Moreover, cut stem twigs were unable to respond to weevil feeding increasing NVR according to the weevil damage. In Experiment 2, assessment of weevil damage on excised branches explained around 50% of variation in damage on living branches. This relationship became much more pronounced (R2 = 0.81) when explored at the mean treatment level; branch manipulation did not alter the patterns of variation in resistance across pine species or MJ treatments. Irrespective of the assessment procedure, MJ

Interspecific phenotypic variation in plant secondary metabolites (PSM) is often explained by biotic and abiotic factors. However, patterns of variation within species do not clearly fit the theoretical predictions. Exploring how genetics, environment and demographic processes shape such variation among and within populations is crucial for understanding evolution of PSM, particularly in long‐lived plants such as forest trees. Here, we quantified genetic variation in PSM among and within populations, and explored drivers of local adaptation by studying the role of climate as a source of population differentiation in PSM of maritime pine. Constitutive profile and concentrations of 63 PSM and their herbivory‐associated inducibility were determined in the bark of 130 clonally replicated genotypes with known familial structure from 10 populations covering the distribution range of the species. We compared neutral and quantitative population genetic differentiation of PSM (FST and QST). Also, we accounted for population genetic structure and kinship among individuals when exploring climate–trait relationships. We found large population differentiation and additive genetic variation in constitutive PSM. Many PSM were inducible, although very low genetic variation was observed with respect to their inducibility. QST–FST comparisons suggest that differentiation of most diterpenes, monoterpenes, and phenolics can be explained by neutral demographic processes. Spatially heterogeneous selection across populations leading to local adaptation was only found for total constitutive sesquiterpenes and a few individual PSM. After accounting for population genetic structure, only the constitutive concentration of two sesquiterpenes showing signs of diversifying selection was predicted by climate, with decreasing concentrations along a growth‐prone climatic gradient. Synthesis. Evolutionary patterns of plant secondary metabolites depended on their chemical nature, with neutral differe

Las coníferas, de gran tamaño y longevidad, están expuestas a múltiples estreses bióticos y abióticos. El cambio global conlleva a un incremento de eventos de sequía y aparición de nuevos estreses bióticos. Los pinos responden a la sequía mediante cierre estomático, reduciendo la entrada de carbono necesario para la síntesis de compuestos defensivos. El objetivo de este trabajo es conocer si el estrés hídrico afecta a la capacidad defensiva de Pinus halepensis frente a Hylobius abietis. Para ello, se sometió a P. halepensis a tres niveles de estrés hídrico: control, moderado y severo y dos niveles de herbivoría: control y herbívoro. Los resultados obtenidos mostraron un aumento del daño causado por el insecto de un 75 % en estrés moderado, volviendo a los mismos niveles que los de plantas control en estrés severo. La herbivoría redujo la concentración de fenoles y taninos e incrementó la de diterpenos. La interacción de ambos tuvo efecto a nivel cuantitativo y cualitativo en fenoles y terpenos. Estos resultados aportan luz a posibles respuestas que puedan presentar los pinos mediterráneos a diferentes estreses asociados al cambio global.

During their long lifespan, pines must cope with simultaneous abiotic and biotic stresses such as drought andherbivory. Mediterranean pines are isohydric species that rapidly close their stomata in response to droughtreducing carbonfixation. In such situations, the synthesis of chemical defences could be impaired. Here, wetested the hypothesis that drought stress may constrain the capability of Mediterranean pines to defend againstherbivory and to induce chemical defences. For this purpose, we subjected three contrasting populations ofAleppo pine (Pinus halepensisMill.) to three levels of drought stress, thereafter exposing the seedlings to theherbivoreHylobius abietisL. A suite of ecophysiological and defensive traits was measured to explore the in-teraction between both stresses. Drought significantly affected the13C signature and reduced starch and fatty acids concentration. Damagecaused by the insect was affected by drought stress, being 75% higher at the moderate stress level but returningunder severe stress to similar values as control seedlings. Seedlings responded to herbivory by decreasing theconcentration of total polyphenolics and condensed tannins, increasing the concentration of total diterpenes, andmodifying the profile of major terpenes. Induced responses to herbivory were, as expected, altered by drought.Inducibility of polyphenolics decreased as drought stress increased while for diterpenes it was higher at mod-erate stress. Moreover, a significant drought × herbivory interaction was found on the multivariate terpeneprofile.These results should be considered for predicting responses of pine forests to the forecasted increase of abioticand biotic risks associated with global change.

In Mediterranean regions, temperature and drought frequency will likely increase by the end of this century. Biotic stress caused by plant pathogens and herbivores is also predicted to rise associated with global change. The ability of tree species to overcome the new environmental conditions will depend on both, their environmental plasticity and the extant intraspecific adaptive variation in functional traits related to resistance and tolerance against environmental stressors. Phenotypic variation for traits related to growth, productivity and abiotic stress tolerance have been well studied, but little is still know about defensive traits. Aditionally, most studies focus on the assesment of defensive function as punctual snapshots, whereas defensive investment is influenced by past climatic and biotic events during the tree lifespan. Resin ducts are specialized structures for the synthesis and accumulation of oleoresin, the main defensive compound of conifer trees against pest and pathogens. The abundance of resin ducts have been related with the resistance to specific herbivores and pathogens, thus becoming a trait with potential adaptive value. Resin duct production is costly, and as any other trait, their phenotypic expression may rely on genotypic and environmental factors. Therefore, understanding the interactive effects of these factors on this major defensive mechanism becomes crucial to predict the performance and survival of Mediterranean trees in the near future. Tree–ring anatomy is a valuable source of ecological information. Traditionally, tree rings have been used to asses the influence of climatic factors over traits related to growth, but inter-annual variability of any tree-ring-related feature can be quantified. Such is the case of axial resin ducts which reflect the variation of defensive investment of long –lived pine trees across their lifespan. Here we applied dendroecological methods to retrospectively asses the phenotypic variation in the y

The ability of tree species to overcome the new environmental conditions associated to climate change, such as drought increase and rising biotic stress, will depend on their phenotypic plasticity, and the extant intraspecific adaptive variation in functional traits. While both sources of variation have been well studied in traits related to productivity and abiotic stress tolerance, little is still know about defensive traits. The abundance and size of resin ducts in several conifer species have been associated with resistance to pest and pathogens. As any other trait, their phenotypic expression may rely on genotypic and environmental factors. Understanding the interactive effects of these factors on this major defensive mechanism is crucial to forecast at what extent tree species will persist in a challenging environment. Usually, most studies focus on assessment of defensive function as punctual snapshots and little is still known about defensive variability along ontogenic development. Here, we applied dendroecological methods to retrospectively asses the intraspecific genetic variation in the yearly expression of resin ducts in Pinus pinaster. We used stem cores collected from nine populations of the whole natural distribution range established in two 40 years-old replicated genetic trials with contrasted climatic conditions. We measured tree-ring width series and separately counted the axial resin ducts in both the latewood (LW) and the earlywood (EW) for the last 30 years on every core. Among-population variation in resin duct production within the annual tree rings was high, but more importantly, differences among populations largely varied along the ontogenetic development (significant population x year interaction). Population investment in defensive structures drastically changed between early and late developmental stages. Additionally, differences between populations regarding defensive investment were also related to the distribution of resin ducts wi

Resistance to herbivores and pathogens is considered a key plant trait with strong adaptive value in trees, usually involving high concentrations of a diverse array of plant secondary metabolites (PSM). Intraspecific genetic variation and plasticity of PSM are widely known. However, their ecology and evolution are unclear, and even the implication of PSM as traits that provide direct effective resistance against herbivores is currently questioned. We used control and methyl jasmonate (MJ) induced clonal copies of genotypes within families from ten populations of the main distribution range of maritime pine to exhaustively characterize the constitutive and induced profile and concentration of PSM in the stem phloem, and to measure insect herbivory damage as a proxy of resistance. Then, we explored whether genetic variation in resistance to herbivory may be predicted by the constitutive concentration of PSM, and the role of its inducibility to predict the increase in resistance once the plant is induced. We found large and structured genetic variation among populations but not between families within populations in resistance to herbivory. The MJ-induction treatment strongly increased resistance to the weevil in the species, and the genetic variation in the inducibility of resistance was significantly structured among populations, with greater inducibility in the Atlantic populations. Genetic variation in resistance was largely explained by the multivariate concentration and profile of PSM at the genotypic level, rather than by bivariate correlations with individual PSM, after accounting for genetic relatedness among genotypes. While the constitutive concentration of the PSM blend did not show a clear pattern of resistance to herbivory, specific changes in the chemical profile and the increase in concentration of the PSM blend after MJ induction were related to increased resistance. To date, this is the first example of a comprehensive and rigorous approach in which i

Because of their huge size and longevity, pines are exposed to numerous pests and pathogens, and have to cope with periods of severe abiotic stress such as water scarcity. Climate models predict an increase in mean annual temperatures and a decrease in precipitation, resulting in longer and more intense periods of drought. Associated to global change, an increase of the incidence of pests and pathogens has also been predicted. Understanding how pines are able to cope, at the same time, with both biotic and abiotic stress is, thus, necessary to know how they will finally respond to global change. Pines are isohydric species whose main mechanism to prevent water losses is the closure of stomata. Closing stomata under water deficit may imply a decrease in the carbon uptake needed for growth, reproduction and defenses. Hormonal cascading in response to both biotic and abiotic threats may also interact among each other altering the ability to tolerate each single stress. The aim of this study is to test how drought stress can constrain pine defensive responses to insect herbivory. For this purpose, 8 month-old seedlings of Maritime pine (Pinus pinaster Ait.) originating from three different populations known to differ in their tolerance to drought (San Ciprián, Coca and Oria) were submitted to three levels of water scarcity (control, moderate and severe drought stress) during four months and then subjected to herbivory (control and the pine weevil Hylobius abietis L.) for five days. Then, seedlings were harvested and biomass allocation, water potential, 13C concentration, non-structural carbohydrates and chemical defensive composition were studied. Water treatments promoted drought stress, measurable by a less discrimination to 13C, a decrease in water potential and reduced seedling dry weight in all populations. Moreover, the insect herbivore inflicted less damage on water-stressed than on well-watered plants. Pine responses to insect damage included alterations of the

El pino marítimo es capaz de responder a herbívoros y patógenos modificando cualitativa y cuantitativamente su arsenal de defensas químicas. También se sabe que presenta variación genética en compuestos defensivos, como sucede con otros caracteres de historia vital, sin embargo, se conoce poco en qué medida la plasticidad (inducibilidad) de las defensas químicas al estrés biótico es variable a nivel intraespecífico y cómo la posible variación en plasticidad de dichos compuestos químicos está estructurada dentro y entre poblaciones. Con el objetivo de evaluar la variación genética intraespecífica en las defensas constitutivas y su inducibilidad realizamos un experimento en invernadero en el que estudiamos el perfil y concentración de terpenos y fenoles en réplicas clonales de diez poblaciones del rango de distribución natural de la especie con estructura familiar (colección CLONAPIN®). El daño por herbivoría fue simulado mediante metil-jasmonato, un análogo hormonal involucrado en la señalización del daño biótico. Hemos identificado y cuantificado 118 metabolitos secundarios en 260 plantas experimentales. Nuestros resultados muestran la especie presenta una gran variación dentro y entre poblaciones en el metaboloma defensivo constitutivo, mientras que la inducción con metil-jasmonato incrementó la concentración de forma generalizada en el conjunto de defensas químicas y no en compuestos individuales.

La producción de defensas químicas en los pinos es costosa y debe balancearse con la inversión en otras funciones vitales. Cambios ambientales que alteren el proceso fotosintético, como la falta de luz, podrían afectar la capacidad defensiva de las plantas frente a herbívoros. En trabajos previos se observó que el daño por Hylobius abietis, importante herbívoro en la etapa juvenil de Pinus pinaster, es mayor en oscuridad. Nuestro objetivo es conocer por qué la disponibilidad de luz modifica el resultado de la interacción planta-insecto, algo que podría deberse a restricciones fisiológicas en la planta o al hábito nocturno del insecto. Para ello expusimos pinos de tres procedencias sometiendo planta y/o insecto a luz u oscuridad durante 6 días en un diseño factorial completo con muestreos destructivos, estudiando las respuestas de la planta y el consumo del insecto. La privación de luz sobre la planta no afectó a la intensidad de la respuesta defensiva (P=0.616); el daño por el insecto fue mayor cuando éste se encontraba en condiciones de oscuridad (P=0.003) y difirió significativamente entre procedencias (P=0.044). Estos resultados sugieren un efecto directo de la luz el comportamiento del insecto, sin restricciones fisiológicas defensivas de la planta en oscuridad.

This dataset compiles quantitative information about the resistance of Norway spruce (Picea abies) half-sibs to the pine weevil Hylobius abietis collected in two naturally infested progeny trials established in Southern Sweden. Growth and diverse components of weevil resistance were assessed in each plant.

Light is a major environmental factor that may determine the interaction between plants and herbivores in several ways, including top-down effects through changes in herbivore behavior and bottom-up effects mediated by alterations of plant physiology. Here we explored the relative contribution of these two regulation processes to the outcome of the interaction of pine trees with a major forest pest, the pine weevil (Hylobius abietis). We studied to what extent light availability influence insect feeding behavior and/or the ability of pines to produce induced defenses in response to herbivory. For this purpose, 3-year old Pinus pinaster plants from three contrasting populations were subjected to 6 days of experimental herbivory by the pine weevil under two levels of light availability (complete darkness or natural sunlight) independently applied to the plant and to the insect in a fully factorial design. Light availability strongly affected the pine weevil feeding behavior. The pine weevil fed more and caused larger feeding scars in darkness than under natural sunlight. Besides, under the more intense levels of weevil damage (i.e., those registered with insects in darkness), light availability also affected the pine’s ability to respond to insect feeding by producing induced resin defenses. These results were consistent across the three studied populations despite they differed in weevil susceptibility and inducibility of defenses. Morocco was the most damaged population and the one that induced more defensive compounds. Overall, results indicate that light availability modulates the outcome of the pine–weevil interactions through both bottom-up and top-down regulation mechanisms.

Seed size is a relevant predictor of seed survival, seedling phenotype and species recruitment and it is generally accepted that larger seeds give rise to seedlings with improved performance. In pioneer tree species the success in seedling survival is strongly determined by resource availability, which largely varies at diverse spatial scales. Here, using a pioneer Mediterranean pine species as a model (Maritime pine, Pinus pinaster Ait.), we hypothesized that the positive effect of seed provisioning on seedling establishment and early performance may be stronger under environmental harassment, and that the progeny of lineages investing in bigger seeds would be favored in such stressful conditions. We explored the effect of seed mass on seedling traits related to germination, survival, growth and biomass partitioning by sowing pre-weighted seeds of known genetic background and culturing the seedlings under a range of abiotic stress conditions by experimentally modifying phosphorus (four levels, 5260 seeds) and water availability (three levels, 975 seeds) in two independent experiments. Phenotypic variation in seed mass was extremely large, with more than a tenfold change between the smallest and the largest seed. Seed mass variation showed a strong genetic component with large and significant variation both between and within populations. As expected, seed mass was a positive predictor of germination time and early seedling performance. However, contrary to our expectations, populations from more stressful sites produced smaller seeds, and the positive effect of seed mass on seedling performance was not magnified under resource limitation. Our results thus does not support that early seedling performance is favored by greater seed provisioning under resource limiting conditions.

Pine tree chemical defence is based on constitutive and inducible carbon-rich substances (mainly oleoresin and phenolic compounds) present in pine tissues in conspicuously high concentrations. Resistance at early stages is essential for future fitness, and allocation of carbon resources to chemical defences may signify a relevant carbon sink for juvenile pines. However there is still poor knowledge about the extent to which allocation priorities of recently assimilated and stored C pools to chemical defences are plastic in response to biotic stress. We aimed to determine the extent to which the production of induced chemical defences in pine trees in response to herbivory depends on the availability of current photosynthates or if they can be built upon stored C pools. Two years-old trees of three pine species (Pinus pinaster, Pinus sylvestris and Pinus radiata) were exposed to real herbivory by the pine weevil Hylobius abietis, a bark-chewing insect, during one week under conditions of either natural sunlight or complete light deprivation. Weevil damage and the concentration of chemical defences in the whole stem were measured in a time-course design with destructive sampling. We analysed, using an ANCOVA approach, whether the ability to increase the concentration of chemical defences in response to weevil damage was altered by light deprivation. We hypothesised that if induced defences are built mainly upon current photoassimilates, then carbon starvation by light deprivation should strongly hamper the production of new defences. For all pine species, concentration of non-volatile resin significantly increased in response to weevil damage, which was more intense under dark conditions. ANCOVA analysis showed that light deprivation did not constrain the ability of young pines to respond to weevil damage by increasing non-volatile resin content in the stems. No significant covariation was found for total polyphenolics. By evaluating chemical defences in the whole ste

The combination of defensive traits leads to the evolution of ‘plant defense syndromes’ which should provide better protection against herbivores than individual traits on their own. Defense syndromes can be generally driven by plant phylogeny and/or biotic and abiotic factors. However, we lack a solid understanding of (i) the relative importance of shared evolution vs. convergence due to similar ecological conditions and (ii) the role of induced defense strategies in shaping defense syndromes. We investigate the relative roles of evolutionary and ecological factors shaping the deployment of pine defense syndromes including multiple constitutive and induced chemical defense traits. We performed a greenhouse experiment with seedlings of eighteen species of Pinaceae family, and measured plant growth rate, constitutive chemical defenses and their inducibility. Plant growth rate, but not phylogenetic relatedness, determined the deployment of two divergent syndromes. Slow-growing pine species living in harsh environments where tissue replacement is costly allocated more to constitutive defenses (energetically more costly to produce than induced). In contrast, fast-growing species living in resource-rich habitats had greater inducibility of their defenses, consistent with the theory of constitutive-induced defense trade-offs. This study contributes to a better understanding of evolutionary and ecological factors driving the deployment of defense syndromes.

La lucha contra plagas y enfermedades forestales lleva preocupando a los selvicultores desde los comienzos de la gestión forestal. Tradicionalmente todo el esfuerzo se ha centrado en combatir al agente causante, buscando reducir las poblaciones o el impacto de éstas, ya sea mediante aplicación de productos químicos (insecticidas y fungicidas), captura directa con feromonas, medidas de sanea-miento o las técnicas más recientes de control biológico. En general, salvo en el caso de insecticidas y fungicidas, las medidas de control no suelen ser totalmente eficaces, y sólo la combinación de múltiples métodos, por lo general costosos y difíciles de aplicar, consigue reducir de forma significativa el problema. A su vez, las restricciones ambientales, operativas y económicas inherentes a la gestión forestal, limitan de forma drástica la posibilidad de aplicación de productos fitosanitarios específicos y selectivos. En este contexto, y considerando el alarmante aumento de problemas sanitarios durante las últimas décadas en nuestras masas forestales, el estudio científico de los mecanismos de resistencia innatos de nuestras especies forestales a plagas y enfermedades adquiere una especial relevancia. Tras millones de generaciones, la convivencia con múltiples organismos nocivos ha desencadenado la evolución de complejos y eficaces mecanismos de resistencia. Aunque todavía poco estudiados en especies arbóreas por las dificultades asociadas a la investigación en organismos de vida larga y gran tamaño, hoy sabemos que la expresión de estos mecanismos de resistencia es extremadamente variable. Los mecanismos de resistencia dependen tanto de factores genéticos, como de factores ambientales abióticos y bióticos e incluso de complejas interacciones que involucran a múltiples componentes del ecosistema. Esta enorme variabilidad abre las puertas a la investigación de nuevas formas de gestión forestal sostenible orientada a maximizar la expresión de resistencia y tolerancia en las mas

Following an herbivore attack plants can reduce damage by inducing a wide variety of highly specific chemical and physical defences. Plants are able to recognize, integrate and respond accordingly in a specific manner to the biotic stimuli provoked by different herbivore species. This specificity of induced defences is particularly important for long-lived plants (e.g. pines) which have to face against multiple biological aggressions during their life cycle. In this sense, it has been recently documented that induction of mono- and sesquiterpenes (the volatile fraction of oleoresin) in pine trees is specific to plant species, herbivore identity and plant tissue. However, much less attention has been paid to understand the sources of phenotypic variation of diterpenes, the non-volatile fraction of oleoresin that seals the wounds after oxidization.

Pine trees are long -lived, widespread organisms that have to cope with multiple biotic stresses during their life cycle. To face against biological aggressions pine trees harbour terpenoids and phenolics as major chemical defences that may avoid or deter the attack providing resistance. Mono and sesquiterpenes, the volatile fraction of oleoresin, have been found to be adaptive resistance traits and to differ between and within species. However much less attention has been paid to sources of phenotypic variation in diterpenes, the non- volatile fraction of oleoresin that seals the stem wounds after oxidization. We examined the induced defensive responses of the diterpene fraction in two pine species, one native (Maritime pine) and one exotic (Monterey pine) to Europe, against two native insect herbivores, the bark chewing pine weevil and the pine processionary caterpillar; and explored the association between the induced chemical profile of diterpenes in each species and the subsequent damage. We found that the two pine species showed different constitutive and induced diterpene concentration in stem and needles. Whereas Pinus pinaster increased total di terpene concentration only in the stem against the two herbivores, P. radiata diterpene defences were locally induced in the damaged tissue due to herbivore feeding activity. Regarding to the chemical diterpene profile, the native P. pinaster produced a ge neral induced response increasing diterpene concentration against the two herbivores in the stem, whereas the exotic P. radiata showed a weevil -induced increase and a caterpillar -induced decrease in the most abundant diterpenes in the needles.

La producción de defensas químicas inducidas por el ataque de insectos y patógenos es un importante mecanismo de defensa en los pinos. Pero como el carbono asignado a defensas deja de estar disponible para otras funciones, para entender los costes asociados a la síntesis de defensas inducidas es necesario conocer el origen del carbono (almacenado o recién asimilado) dedicado a su síntesis. Nuestro objetivo fue doble: (i) conocer el efecto de la luz sobre el patrón de alimentación del curculiónido Hylobius abietis; y (ii) conocer si la ausencia de luz afecta a la capacidad de los pinos para producir defensas inducidas en respuesta al ataque por este insecto. Expusimos Pinus pinaster jóvenes de tres procedencias a herbivoría experimental bajo dos factores ambientales cruzados: restricción de luz en la zona de alimentación del insecto y restricción de luz en la planta, y medimos el daño por el insecto y la concentración de defensas químicas (resina y fenoles) en la planta tras 2 y 6 días de herbivoría. La tasa de alimentación del insecto fue marcadamente mayor en oscuridad en todas las procedencias, independientemente de que la planta estuviese en luz o en oscuridad. No encontramos evidencias de que ello estuviese relacionado con una menor capacidad de síntesis de defensas químicas en la planta en ausencia de luz, pues ésta, no afectó a la inducibilidad de resina de forma significativa. Esto sugiere una importante flexibilidad en la asignación de recursos a defensas en ausencia de fotosíntesis.

As any other living entity, forest trees must optimize resource investment to growth, reproduction and maintenance, but investment in one function may occur at the expense of other functions, resulting in trade-offs that limit maximization of all processes. Global change is imposing new challenges to Mediterranean forests, in particular combined abiotic and biotic stressors such as drought and epidemic outbreaks of pests and pathogens that may require increased defensive investment. Mediterranean pines are expected to harbour a valuable adaptive genetic diversity, already shaping the populations since long ago, for facing these challenges. Little is known, however, about how populations will respond to multiple challenges, and also about the correlated responses to a single stressor in other life history traits. Here we used as a model a Maritime pine (Pinus pinaster Ait.) population historically exploited by tapping for oleoresin extraction and artificially selected for increased turpentine and rosin production. In a common garden with 60 open-pollinated half-sib families (selected for resin production plus a control seedlot) we evaluated the constitutive and induced resin flux as a proxy of defensive investment and concentration of major groups of chemical defences in phloem-cortex and needles. We also measured a wide set of traits related to growth and maintenance (height, diameter and bark thickness), reproduction (cone counts and serotiny level) and drought-resistance (13C-based water-use efficiency). Our objectives were to determine the intrapopulation genetic variation in these key phenotypic traits and to explore the existence of tradeoffs among adaptive life history traits. Preliminary results showed the existence of significant genetic variation among families for most assessed variables, and distinct trade-offs between growth and maintenance, reproduction and defense. We will discuss to what extent variations in the carbonwater balance at the intra-specif

Only in Sweden 360 million conifer seedlings are planted each year but a substantial proportion of these are killed by pine weevils (Hylobius abietis) and black spruce beetles (Hylastes cunicularius). A high survival rate is essential for a high primary production of forest raw materials and biomass. There is currently no suitable non-insecticide plant protection option for a large part of these seedlings. One option may, however, be to trigger the seedlings´ own induced defense by treating them with a plant activator at the nursery so that they are prepared when they later becomes attacked by pine weevils in the field. The induced defense is under natural conditions generally expressed after damage by, e.g. a pest insect, whereas constitutive defenses are permanently present. Constitutive defenses serves to inhibit an initial attack whereas the induced defense serves to actively and vigorously stop the attack. The induced defense is powerful but the time lag after an initial attack before the induced defenses are fully expressed may be detrimental for a plant when the pest pressure is high. This weakness of the induced defense can be circumvented by applying a plant activator to trigger the induced defense of the plant before it is exposed to attacks. There is a suitable plant activator to trigger the induced defense of conifers available. That is methyl jasmonate which is a substance that also is produced naturally when a plant is attacked. It triggers the production of traumatic resin channels and different types of chemical defense substances. In this talk I will present some recent results related to the potential of this method to protect conifer seedlings against pine weevils.

Maritime pine has many isolated and highly differentiated populations, ranging from stressful Mediterranean climates to benign Atlantic coastal areas in Southwestern Europe. Differentiation processes in phenotypic traits such as drought tolerance, reproductive behaviour and growth potential has been suggested to be adaptive. However little is known about the differentiation in defensive traits and strategies. In this paper we will summarize the results from several field and greenhouse experiments focused on identifying the sources of phenotypic variability of constitutive and inducible allocation to chemical defences in this model species. We have studied genetic variation within and across populations, resource availability and other factors such as ontogeny, within plant variation and herbivore identity. We usually find significant variation in defensive strategies across populations, and as well that families within populations differ in the constitutive and induced expression of defensive chemical traits. Particularly when analyzing terpenoids, we usually find high narrow sense heritability for the concentration of many individual terpenoids species. We also found that inducibility of those compounds (in response to real herbivory or methyl jasmonate application) is also usually genetically variable. Constitutive and inducible allocation to chemical defences are tissue specific, part specific and also compound specific and as well depends on the nature of the herbivore damage. Altogether, these results suggest that, in spite the pressure exerted by herbivores, existence of genetic variation in defensive traits is the common pattern in pine trees, and it is quantitatively relevant. However, although we have found that families differ in effective field resistance against several pests and pathogens, we usually fail to find significant genetic correlations among individual chemical traits and effective field resistance. Trade-offs between defences, between defens

All rights reserved. The pine wilt disease (PWD), caused by the pinewood nematode (PWN) Bursaphelenchus xylophilus (Steiner et Buhrer) Nickle, is one of the most serious threats to pine forests worldwide. Here we studied several components of susceptibility to PWN infection in a model group of pine species widely distributed in Europe (Pinus pinaster Ait., P. pinea L., P. sylvestris L. and P. radiata D. Don), specifically concerning anatomical and chemical traits putatively related to nematode resistance, whole-plant nematode population after experimental inoculation, and several biochemical and physiological traits indicative of plant performance, damage and defensive responses 60 days post inoculation (dpi) in 3-year-old plants. Pinus pinaster was the most susceptible species to PWN colonization, with a 13-fold increase in nematode population size following inoculation, showing up to 35-fold more nematodes than the other species. Pinus pinea was the most resistant species, with an extremely reduced nematode population 60 dpi. Axial resin canals were significantly wider in P. pinaster than in the other species, which may have facilitated nematode dispersal through the stem and contributed to its high susceptibility; nevertheless, this trait does not seem to fully determinate the susceptible character of a species, as P. sylvestris showed similar nematode migration rates to P. pinaster but narrower axial resin canals. Nematode inoculation significantly affected stem water content and polyphenolic concentration, and leaf chlorophyll and lipid peroxidation in all species. In general, P. pinaster and P. sylvestris showed similar chemical responses after infection, whereas P. radiata, which co-exists with the PWN in its native range, showed some degree of tolerance to the nematode. This work provides evidence that the complex interactions between B. xylophilus and its hosts are species-specific, with P. pinaster showing a strong susceptibility to the pathogen, P. pinea