Allocation of 14C assimilated in late spring to tissue and biochemical stem components of cork oak (Quercus suber L.) over the seasons

Carbon distribution in the stem of 2-year-old cork oak plants was studied by 14CO2 pulse labeling in late spring in order to trace the allocation of photoassimilates to tissue and biochemical stem components of cork oak. The fate of 14C photoassimilated carbon was followed during two periods: the first 72 h (short-term study) and the first 52 weeks (long-term study) after the 14CO2 photosynthetic assimilation. The results showed that 14C allocation to stem tissues was dependent on the time passed since photoassimilation and on the season of the year. In the first 3 h all 14C was found in the polar extractives. After 3 h, it started to be allocated to other stem fractions. In 1 day, 14C was allocated mostly to vascular cambium and, to a lesser extent, to primary phloem; no presence of 14C was recorded for the periderm. However, translocation of 14C to phellem was observed from 1 week after 14CO2 pulse labeling. The phellogen was not completely active in its entire circumference at labeling, unlike the vascular cambium; this was the tissue that accumulated most photoassimilated 14C at the earliest sampling. The fraction of leaf-assimilated 14C that was used by the stem peaked at 57% 1 week after 14CO2 plant exposure. The time lag between C photoassimilation and suberin accumulation was ∼8 h, but the most active period for suberin accumulation was between 3 and 7 days. Suberin, which represented only 1.77% of the stem weight, acted as a highly effective sink for the carbon photoassimilated in late spring since suberin specific radioactivity was much higher than for any other stem component as early as only 1 week after 14C plant labeling. This trend was maintained throughout the whole experiment. The examination of microautoradiographs taken over 1 year provided a new method for quantifying xylem growth. Using this approach it was found that there was more secondary xylem growth in late spring than in other times of the year, because the calculated average cell division time was much shorter.

Improved walnut mass micropropagation through the combined use of phloroglucinol and FeEDDHA

Despite the socioeconomic importance of walnut trees, poor rooting and recalcitrance to in vitro culture have hampered the establishment of high-yield clonal plantations. To improve walnut micropropagation, we introduced several modifications to current methods and evaluated the effects on microshoot performance and acclimatization. Nine selected genotypes (13-year-old trees) of the commercial hybrid Juglans major 209 x J. regia were cultured in vitro on DKW-C medium supplemented with 4.4 µM BA and 50 µM IBA. A protocol was developed that relies on the use of 0.40 mM phloroglucinol during shoot multiplication, 0.20 mM previous root induction, and 6.81 mg/L Fe3+ (FeEDDHA). Moreover, the addition of 83.2 µM glucose during the root expression phase significantly improved plant survival during acclimatization. Phloroglucinol promoted microshoot elongation but inhibited rooting, especially at concentrations above 0.40 mM. Replacing FeEDTA by FeEDDHA diminished chlorotic symptoms and improved rooting, with up to 90% microshoots developing viable roots. Likewise, glucose was more efficient than sucrose or fructose in promoting plant survival. At the proposed working concentrations, neither glucose nor FeEDDHA caused any noticeable deleterious effect on walnut micropropagation. Microscopic analysis revealed the physical continuity between adventitious roots and stem pericycles. Analysis of leaf genomic DNA with eight polymorphic microsatellite markers was supportive of the clonal fidelity and genetic stability of the micropropagated material. Successful clonal plantations (over 5,800 ramets) have been established by applying this protocol.

Mechanical behaviour and rupture of normal and pathological human ascending aortic wall

The mechanical properties of aortic wall, both healthy and pathological, are needed in order to develop and improve diagnostic and interventional criteria, and for the development of mechanical models to assess arterial integrity. This study focuses on the mechanical behaviour and rupture conditions of the human ascending aorta and its relationship with age and pathologies. Fresh ascending aortic specimens harvested from 23 healthy donors, 12 patients with bicuspid aortic valve (BAV) and 14 with aneurysm were tensile-tested in vitro under physiological conditions. Tensile strength, stretch at failure and elbow stress were measured. The obtained results showed that age causes a major reduction in the mechanical parameters of healthy ascending aortic tissue, and that no significant differences are found between the mechanical strength of aneurysmal or BAV aortic specimens and the corresponding age-matched control group. The physiological level of the stress in the circumferential direction was also computed to assess the physiological operation range of healthy and diseased ascending aortas. The mean physiological wall stress acting on pathologic aortas was found to be far from rupture, with factors of safety (defined as the ratio of tensile strength to the mean wall stress) larger than six. In contrast, the physiological operation of pathologic vessels lays in the stiff part of the response curve, losing part of its function of damping the pressure waves from the heart.

A computational model coupling mechanics and electrophysiology in spinal cord injury

Traumatic brain injury and spinal cord injury have recently been put under the spotlight as major causes of death and disability in the developed world. Despite the important ongoing experimental and modeling campaigns aimed at understanding the mechanics of tissue and cell damage typically observed in such events, the differenti- ated roles of strain, stress and their corresponding loading rates on the damage level itself remain unclear. More specif- ically, the direct relations between brain and spinal cord tis- sue or cell damage, and electrophysiological functions are still to be unraveled. Whereas mechanical modeling efforts are focusing mainly on stress distribution and mechanistic- based damage criteria, simulated function-based damage cri- teria are still missing. Here, we propose a new multiscale model of myelinated axon associating electrophysiological impairment to structural damage as a function of strain and strain rate. This multiscale approach provides a new framework for damage evaluation directly relating neuron mechanics and electrophysiological properties, thus provid- ing a link between mechanical trauma and subsequent func- tional deficits.

1H HR MAS NMR metabolomic and non-destructive 2D NMR relaxometry to assess internal quality in apples.

NMR can be considered a multi-scale multidimensional technology in the sense that it provides both spatial insight at macroscopic (MRI) or microscopic level (relaxometry), together with chemical characterization (HR-MAS). In this study 296 apples (from 4 cultivars) were MRI screened (20 slices per fruit) among which 7 fruits were used for metabolomic study by 1H HR MAS in order to assess various chemical shifts: malic acid, sucrose, glucose, fructose and ethanol. On the first season, tissue samples were taken from the sound and affected apples (near the core, centre and outer part of the mesocarp) belonging to sound and affected locations, while on the second season, tissue samples were focused on the comparison between sound and affected tissue. Beside, MRI and 2D non-destructive relaxometry (on whole fruits, and localized tissue) where performed on 72 and 12 apples respectively in order to compare features at macroscopic (tissue) and microscopic (subcellular) level. HR MAS shows higher content of ?-glucose, ?-glucose, malic acid and aromatic compounds in watercore affected tissues from both seasons, while sound tissue reflects higher sucrose. Microscopic (subcellular) degradation of tissue varies according to disorder development and is in good accordance with macroscopic characterization with MRI.

Respuesta fisiológica y agronómica del olivar en seto (cv. Arbequina) a la orientación de filas y al ancho de calle: relación entre la producción y calidad del aceite y la radiación interceptada

En las últimas dos décadas, los productores han plantado olivares en seto para lograr la mecanización de la poda y en especial de la cosecha, reducir los costes de mano de obra y permitir intervenciones de manejo rápidas y oportunas. Los olivares se desarrollaron en ausencia del conocimiento científico, sobre el diseño óptimo de la estructura de la copa, necesario para incrementar la producción y calidad del aceite. En contraste, con los árboles muy espaciados y distribuidos uniformemente de las plantaciones tradicionales, en el olivar en seto hay una marcada variabilidad espacial y temporal de la radiación disponible en función del diseño de la plantación. Así, conocer la respuesta fisiológica y productiva del olivo a la radiación resulta fundamental en el olivar en seto. La orientación de las filas y el ancho de calle son aspectos que se deciden en el diseño de las plantaciones en seto. Ambos aspectos modifican la radiación interceptada por la canopia y, por lo tanto, pueden incidir en la productividad y calidad del aceite. Una vez realizada la plantación no pueden ser modificados, y así las ventajas o desventajas permanecerán fijas durante toda la vida productiva del olivar. A pesar de esto, el impacto de la orientación de las filas y el ancho de calle han recibido poca atención en olivos y en la mayoría de los frutales conducidos en seto. Por todo ello, los objetivos principales de esta tesis fueron, (i) evaluar el efecto de la orientación del seto y del ancho de calle, sobre la productividad y calidad del aceite, (ii) evaluar un modelo que estime la radiación dentro de la canopia. Este modelo permitirá cuantificar las relaciones entre la radiación y los componentes del rendimiento y calidad del aceite de olivares en setos con un amplio rango de estructuras y (iii) conocer la variabilidad en las características de las hojas (morfológicas y fisiológicas) y de los tejidos del fruto (tamaño y composición) en diferentes posiciones de la copa de los setos. Para ello, se dispuso de 3 ensayos de olivar en seto (cv. Arbequina) implantados en 2008 en el municipio de La Puebla de Montalbán, Toledo. La primera cosecha fue en 2010 y a partir del 2012 los setos formaron una copa continua. A partir de ese año, los setos se mantuvieron mediante poda, con similar ancho (~1 m) y altura (~2,5 m), acordes a las dimensiones de la cosechadora vendimiadora. En los años 2012 y 2013 se estudió en profundidad la respuesta de las plantas de estos ensayos. En el ensayo 1, los setos fueron plantados con cuatro orientaciones de filas: N–S, NE–SO, NO–SE y E–O y el mismo ancho de calle (4 m). En los otros dos ensayos, los setos fueron plantados con tres anchos de calle (5,0, 4,0 y 2,5 m), y con filas orientadas N–S (ensayo 2) y E–O (ensayo 3). La respuesta de la orientación de las filas se evaluó a nivel de seto y de estratos del seto (alturas y caras), a través de mediciones del crecimiento de brotes, componentes reproductivos, características y temperatura del fruto, estado hídrico del suelo y de las plantas, fotosíntesis neta de las hojas y contenido de ácidos grasos. Los setos orientados NE–SO (2,7 t/ha) lograron la mayor producción de aceite, que fue significativamente más alta que la de los setos E–O (2,3 t/ha). La producción de aceite de los setos E–O no se diferenció estadísticamente de los setos N–S (2,5 t/ha). Las diferencias productivas entre orientaciones fueron explicadas por el número de frutos en cosecha, a su vez la variación en el número de frutos estuvo asociada al efecto de la orientación de las filas sobre el número de yemas desarrolladas y el porcentaje de inflorescencias fértiles. Las hojas en las caras iluminadas de los setos NE–SO y N–S presentaron mayor tasa fotosintética a la mañana (~10.0 h) que los setos E–O, en el año 2012, pero no en 2013. La orientación de las filas no tuvo un efecto significativo en el contenido de ácidos grasos de los aceites extraídos, esto ocurrió a pesar de variaciones en la temperatura interna de los frutos (3 °C) y de la radiación (40%) entre las distintas caras de los setos. La orientación del seto afectó significativamente al contenido relativo de agua del suelo, donde setos E–O presentaron valores más altos (12%) que setos N–S durante el verano y otoño. Sin embargo, el potencial hídrico de tallo fue similar entre orientaciones. En los ensayos 2 y 3, se evaluó el efecto que produce, a nivel de seto y de estratos (caras y alturas), reducir el ancho de calle de 5,0 a 4,0 y 2,5 m, en un seto orientado N–S y otro E–O, respectivamente. La relación entre altura/ancho de calle libre aumentó 0,6 a 0,8 y 1,6, al reducir 5,0, 4,0 y 2,5 m el ancho de calle, mientras la longitud de seto y el volumen de copa por hectárea incrementó 100% al reducir de 5,0 a 2,5 m, el ancho de calle. En los setos orientados N–S, la producción de aceite por ha acumulada en 4 campañas, incrementó significativamente un 52 %, al reducir de 5,0 a 2,5 m el ancho de calle. Los setos N–S con calle más estrecha (2,5 m) tuvieron un 19% menos frutos que los setos con calle más ancha (5,0 m) y a su vez el 60% de los mismos se localizaron los estratos altos de la canopia de los setos con calles estrecha en comparación al 40% en setos con calle de 5,0 m. En los estratos más bajos de los setos con calles de 2,5m hubo menor crecimiento de los brotes y los frutos tuvieron menor peso seco, contenido de aceite y madurez, que los frutos en los estratos bajos de los setos a 5,0 m. Los componentes del rendimiento y características de los frutos (agua y madurez) fueron similares entre la caras E y O, independientemente del ancho de calle. En los setos orientados E–O, la producción de aceite por ha acumulada en 4 campañas, no respondió significativamente al ancho de calle, debido a una disminución significativa en el número de frutos y producción de aceite por m de seto, al reducir de 5,0 a 2,5 m, el ancho de calle. En los setos orientados E–O, con calles de 5,0 m, los frutos presentaron similar peso seco, contenido de aceite y agua, en las caras S y N, sin embargo, cuando la calle fue reducida a 2,5, los frutos de la cara S fueron más pesado y maduros que en la cara N. Independientemente del ancho de calle y de la orientación del seto, el aceite presentó mayor contenido de ácidos palmitoleico, palmítico, esteárico y linoleico en los frutos del estrato más alto de la canopia disminuyendo hacia la base. En contraste, el contenido de ácido oleico aumentó desde el estrato más alto hacia la base de los setos. Las diferencias en el contenido de ácidos grasos entre la parte alta y baja de los setos, incrementó al reducir el ancho de calle en los setos N–S, pero no en los E-O. En conclusión, en olivares en seto, reducir el ancho de calle permite incrementar la producción de aceite, en setos orientados N–S, pero no en E–O. Un modelo que estima la cantidad y distribución de la radiación en toda la copa del seto, fue utilizado para estimar la radiación interceptada en distintos estratos del seto. El modelo requiere un valor del coeficiente de extinción (k) para estimar la transmisión de radiación a través de la copa, el cual fue obtenido experimentalmente (k=1,2). Utilizando los datos del ensayo 1, un único modelo lineal relacionó el peso seco y el rendimiento graso de setos con la radiación interceptada por los distintos estratos de setos con cuatro orientaciones de filas. La densidad de frutos fue también relacionada con la radiación, pero más débilmente. En los setos orientados N–S, plantados con tres anchos de calles, (ensayo 2) el contenido de ácidos palmitoleico y linoleico del aceite incrementó linealmente con el incremento de la radiación interceptada, mientras el contenido ácido oleico disminuyó linealmente con el incremento de la radiación. El contenido de ácidos grasos del aceite no estuvo relacionado con la radiación interceptada en setos orientados E–O (Ensayo 3). En los setos N–S y E–O, plantados con anchos de calle de 2,5 m, se estudiaron las interacciones entre la radiación y características de las hojas, número de fruto, tamaño y composición de los frutos a nivel de órgano, tejido y células. Independientemente de la orientación del seto, el área y el contenido de clorofila de las hojas incrementaron significativamente en los estratos más bajos de los setos. Mientras, las hojas de los estratos medios del seto presentaron mayor capacidad fotosintética que en los estratos bajos y alto de los setos. Los estratos del seto que interceptaron más radiación produjeron frutos con mayor tamaño y contenido de aceite en el mesocarpo, sin efectos sobre el tamaño y composición del endocarpo. A nivel celular, los frutos expuestos a mayor nivel de radiación desarrollaron en el mesocarpo células de mayor tamaño en comparación a frutos menos expuestos, mientras el número de células no fue afectado. Adicionalmente, el número y tamaño de las células estuvo relacionado con la composición del mesocarpo en términos de aceite, agua y peso seco menos aceite. Esta tesis, contribuye, desde una perspectiva integral del cultivo del olivo, a cuantificar el impacto de la orientación y ancho de calle sobre la producción y calidad del aceite en olivares conducidos en setos. El análisis y discusión de la relación entre la radiación y los componentes del rendimiento y calidad del aceite, puede ayudar a diseñar plantaciones en seto con dimensiones óptimas para la intercepción de la radiación. ABSTRACT In the last two decades, olive hedgerow system has been established by commercial growers to allow continuous mechanized pruning and especially harvest, reduce costs of manual labour and allow more rapid and timely management interventions. The adoption of hedgerow was done in the absence of adequate scientific knowledge of the impact of this orchard structure and associated mechanization on tree response, yield and quality, after centuries in low-density orchards and open-formed trees. The row orientation and width alley are fundamental aspects in the hedgerow design and have been scarcely studied in olive. Both aspects modify the radiation intercepted by the canopy, and consequently the productivity and oil quality, and once defined in orchard planting cannot be changed, so advantages and disadvantages remain fixed for the lifespan of the orchard. The main objectives of this thesis were to (i) evaluate the impact of the row orientation and width alley on productivity and oil quality by the measurements of profile of the determining processes of shoot growth, fruit temperature, yield components and fruit and oil characteristics on opposite sides of olive hedgerows. Additionally, the effect of row orientation on the plant water status was also evaluated; (ii) evaluate a mathematical model for estimating the radiation within the canopy and quantify the relationships between the radiation estimated and yield components and oil quality in olive hedgerows under wide range of structures and; (iii) determine the variability in the characteristics of the leaves (morphological and physiological) and fruit tissues (size and composition) in different positions of the hedgerows canopy. Three plots of olive hedgerows (cv. Arbequina) planted in 2008 in La Puebla de Montalbán, Toledo were evaluated during the 2012 and 2013 seasons. The hedgerows were maintained by lateral pruning and topping with the same width (1 m) and height (2.5 m) compatible with the intended harvester. In a plot (experiment 1), the hedgerows were planted with the same width alley (4 m) and four row orientations: N–S, NE–SW, NW–SE and E–W. Other two plots (Experiments 2 and 3) separated by approximately 100 m were planted with N–S and E–O oriented rows and three alley widths in each orientation: 5.0, 4.0 and 2.5 m. In the exp. 1, maximum fruit yield were achieved by NE–SW and NW–SW (15.7 t/ha). Of these, NE–SW achieved the highest oil yield (2.7 t/ha). There were no differences in fruit or oil yield between N–S (2.5 t oil/ha) and E–W (2.3 t oil/ha) orientations. Fruit number was the most important component to explain these differences, by previous influence on number of bud developed and percentage of fertile inflorescences. Fruit maturity and oil quality on both sides of the hedgerows were not affected by row orientation. This occurred despite significant variations in the internal fruit temperature, which was closely related to the irradiance received by the canopy and the time of day. Additionally, row orientation significantly affected the relative water content of the soil, where E–W oriented hedgerows showed consistently higher values than N–S during summer-autumn season. The stem water potential at midday, however, was similar between orientations, revealing possible lower water consumption of E–W than N–S oriented hedgerows. In the exp. 2, regardless of row orientation, reduction of row spacing from 5.0 to 4.0 and 2.5 m increases the ratio of canopy depth to free alley width (Al/An) from 0.6 to 0.8 and 1.6, respectively, and ads 25 and 100 % more hedgerow length per ha. In N–S oriented hedgerows, oil production per ha increased significantly by 14 and 52 % in 4.0 m and 2.5 m relative to 5.0 m row spacing, the effect being proportionally less than the increase in hedgerow length per ha. Hedgerows spaced 2.5 m with Al/An = 1.6 produced relatively fewer fruits per unit length than did wider spacings and were preferentially distributed in upper layers. Fruits located at the bottom of the canopy were smaller, with lower oil content and were less mature. In E–W oriented hedgerows, oil production per ha did not respond significantly to row spacing, despite the doubling of row length from the 5.0 to the 2.5 m row spacing. The explanation was found in fewer fruit per unit length of hedgerow and smaller oil content at 2.5 m than 5.0 m row spacing, averaged over the experimental period. In E–W hedgerows spaced at 5.0 m with Al/An = 0.6, the vertical profiles of fruit characteristics (mass, oil and water contents, and maturity) were similar between opposing sides, but at 4.0 m (Al/An= 0.8) and 2.5 m (Al/An=1.6) spacings, fruits on the S side were heavier and more mature than on N side. The oil extracted from fruits harvested at different heights of N–S and E–W oriented hedgerows showed higher palmitoleic, palmitic, stearic and linoleic contents at the canopy top decreasing toward base. The oleic content was reverse, increased from top to base. In N–S hedgerows, vertical gradients increased by reducing the alley width, but not in the E–W oriented hedgerows. The simulation of internal canopy irradiance was related in a single relationship (R2 = 0.63) to the vertical profiles of fruit weight and oil content of olive hedgerows with wide range of structures. The density of fruits was also associated with the irradiance but more weakly (R2 = 0.27), and revealed a more complex response involving changes in the vegetative structure by canopy management (topping) and the effect of radiation on the previous sequence that defines the number of fruits. The vertical profiles of oil quality traits were closely associated to canopy irradiance, but only when the N–S oriented hedgerows were considered. The contents of palmitoleic and linoleic acid in the oil increased linearly when intercepted irradiance increased from 9 to 19 mol PAR/m2. In contrast, oleic content decreased linearly in this irradiance range. Additionally, we advanced knowledge regarding the interactions among irradiance and leaf, fruit number, size and composition at organ-, tissue- and cellular- levels. The irradiance received at different positions in the canopy strongly affected the leaf area and chlorophyll content, and mesocarp size and composition (water and oil), without effects on endocarp size and composition. At the cellular level, light-exposed fruit developed larger mesocarp cells than shaded fruits, but cell number was not affected. Our results indicate that cell number and size are related to mesocarp composition in term of oil, water, and dry weight menus oil, although the specific manner in which they interact remains to be determined. This research contributes from an integral perspective of olive growing to quantify the impact of row orientation and width alley on productivity and oil quality in hedgerows systems. The analysis and discussion of the relationships between radiation and yield components and oil quality can help understand the impact of design olive hedgerows in general and in a wide range of environmental conditions.

A computational model coupling mechanics and electrophysiology in spinal cord injury

Traumatic brain injury and spinal cord injury have recently been put under the spotlight as major causes of death and disability in the developed world. Despite the important ongoing experimental and modeling campaigns aimed at understanding the mechanics of tissue and cell damage typically observed in such events, the differentiated roles of strain, stress and their corresponding loading rates on the damage level itself remain unclear. More specifically, the direct relations between brain and spinal cord tissue or cell damage, and electrophysiological functions are still to be unraveled. Whereas mechanical modeling efforts are focusing mainly on stress distribution and mechanistic-based damage criteria, simulated function-based damage criteria are still missing. Here, we propose a new multiscale model of myelinated axon associating electrophysiological impairment to structural damage as a function of strain and strain rate. This multiscale approach provides a new framework for damage evaluation directly relating neuron mechanics and electrophysiological properties, thus providing a link between mechanical trauma and subsequent functional deficits