Growth rate and TRI5 gene expression profiles of Fusarium equiseti strains isolated from Spanish cereals cultivated on wheat and barley media at different environmental conditions

Fusarium equiseti is a toxigenic species that often contaminates ce real crops from diverse climatic regions such as Northern and Southern Europe. Previous results suggested the existence of two distinct populations within this species with differences in toxin pro file which largely corresponded to North and South Europe (Spain). In this work, growth rate profiles of 4 F. equiseti strains isolated from different cereals and distinct Spanish regions were determined on wheat and barley based media at a range of temperatures (15, 20, 25, 30, 35 and 40 °C) and water potentialregimens(−0.7,−2.8,−7.0,and −9.8MPa,correspondingto 0.99,0.98,0.95 and 0.93aw values).Growth was observed at all temperatures except at 40 °C, and at all the solute potential values except at−9.8 MPa when combined with 15 °C. Optimal growth was observed at 20– 30 °C and −0.7/−2.8 MPa. The effect of these factors on trichothecene biosynthesis was examined on a F. equiseti strain using a newly developed real time RT-PCR protocol to quantify TRI5 gene expression at 15, 25 and 35 °C and −0.7, −2.8, − 7.0 and −9.8 MPa on wheat and barley based media. Induction of TRI5 expression was detected between 25 and 35 °C and −0.7 and − 2.8 MPa, with maximum values at 35 °C and −2.8 MPa being higher in barley than in wheat medium. These results appeared to be consistent with a population well adapted to the present climatic conditions and predicted scenarios for Southern Europe and suggested some differences depending on the cereal considered. These are also discussed in relation to other Fusarium species co-occurring in cereals grown in this region and to their significance for prediction and control strategies of toxigenic risk in future scenarios of climate change for this region.

Type of grinding of the main cereal of the diet affects production of brown egg-laying hens

The influence of method of grinding of the cereal of the diet on production and egg quality was studied in 420 Hy-line brown egg-laying hens. The design was completely randomized with six treatments arranged as a 3 × 2 factorial with three cereals (barley, dented maize and soft wheat) and two grinding procedures (hammer mill vs. roller mill). Each treatment was replicated seven times and the experimental unit was an enriched cage with ten hens. Production was recorded every four weeks from 24 to 59 weeks of age and egg quality was measured at 40 and 56 weeks of age. For the entire experiment, feed intake was higher in hens fed wheat or maize than in hens fed barley (110.8 and 110.7 vs. 109.7 g/d; P = 0.014) but most of the differences were observed when the cereal was roller milled (P = 0.009 for the interaction). Also, egg production was similar for the three diets when the cereal was hammer milled but tended to be lower for the barley than for the wheat or maize diets when the cereal was roller milled (P = 0.09 for the interaction). None of the other productive or egg quality traits was affected by dietary treatment. We conclude that roller mills are useful to grind low fiber cereals, such as maize or wheat. However, the use of the roller mill might not be adequate when barley is the main cereal in diets for egg-laying hens.

Factores nutricionales que influyen sobre las variables productivas y el desarrollo del tracto gastrointestinal en pollitas comerciales rubias = Nutritional factors affecting productive performance and gastrointestinal tract traits in commercial brown-egg laying pullets

El objetivo general de esta Tesis doctoral fue estudiar la influencia de diversos factores nutricionales sobre los parámetros productivos y el desarrollo del tracto digestivo de pollitas rubias destinadas a la producción de huevos comerciales. Para alcanzar este objetivo se realizaron tres experimentos donde se estudió el cereal principal, el tamaño de partícula del cereal y el nivel de energía y la presentación de los piensos. En el experimento 1 se estudió la influencia del cereal (piensos con enzimas) y la presentación del pienso sobre los parámetros productivos y las características del tracto digestivo en 576 pollitas rubias de 1 a 120 d de edad. De 1 a 45 d de la edad, se utilizaron 4 piensos experimentales organizados de forma factorial con 2 cereales al 50% de inclusión (maíz vs. trigo) y 2 presentaciones del pienso (harina vs. gránulo de 2- mm de diámetro). Cada tratamiento se replicó 6 veces (24 pollitas por réplica). De 46 a 120 d de edad todas las dietas (maíz o trigo) se ofrecieron en harina y por tanto, la única diferencia entre tratamientos fue el cereal utilizado. De 1 a 120 d de edad, las pollitas que recibieron los piensos basados en maíz tuvieron una ganancia de peso vivo (PV) superior (P < 0,05) que las que recibieron los piensos basados en trigo, pero el índice de conversión (IC) fue similar para ambos grupos. De 1 a 45 d de edad, las pollitas alimentadas con gránulo consumieron más pienso (P < 0,001) y tuvieron una ganancia de peso superior (P < 0,001) que las pollitas alimentadas con harina. Gran parte de los efectos beneficiosos de la granulación sobre los parámetros productivos se mantuvieron al final de la prueba (120 d de edad). A los 45 d de edad, el peso relativo de la molleja (PR; g/kg PV) fue superior (P < 0,01) en pollitas alimentadas con maíz que en pollitas alimentadas con trigo. La alimentación en gránulo redujo el PR del tracto gastro intestinal (TGI) y de la molleja (P < 0,001), así como la longitud relativa (LR; cm/kg PV) del intestino delgado (P< 0.01) a ambas edades (45 y 120 d de edad). El tipo de cereal utilizado no afectó al pH del contenido de la molleja a 120 d de edad pero fué inferior (P < 0,01) en las pollitas que recibieron el pienso en harina de 1 a 45 d de la edad que en las que recibieron el pienso en gránulo. Se concluye que el trigo puede substituir al maíz en piensos para pollitas si se acepta una ligera reducción en la ganancia de peso. Asímismo, la alimentación en gránulo de 1 a 45 d de edad aumentó la ganancia de peso a esta edad y al final de la prueba, así como el pH de la molleja a 120 d de edad. La presentación del pienso en gránulo redujo el PR de la molleja y la LR del TGI a 120 d de edad. En el experimento 2 se utilizaron un total de 864 pollitas rubias Hy-Line de 1 d de edad para estudiar la influencia del cereal de la dieta (500 g de maíz o trigo/kg) y el tamaño de partícula del mismo (molienda con molino de martillos con un diámetro de criba de 6, 8, o 10-mm) sobre los parámetros productivos y las características del TGI de 1 a 120 d de edad. Cada uno de los 6 tratamientos se replicó 6 veces (24 pollitas por réplica). De 1 a 45 d de edad, la ganancia de PV aumentó (P< 0,001) y el IC se mejoró (P < 0,05) al reducir el tamaño de partícula del cereal, pero no se observaron diferencias en el periodo crecimiento de 45 a 120 d de edad. A los 45 d de vida, las pollitas alimentadas con maíz tendieron (P < 0,10) a tener un mayor PR del TGI y del proventrículo y una mayor LR del intestino delgado que las pollitas alimentadas con trigo. Asímismo, el PR del TGI a esta edad, aumentó (P < 0,05) a medida que aumentaba el tamaño de partícula del cereal utilizado. A los 120 d de edad, el tratamiento no afectó el PR de ninguno de los órganos del TGI ni al pH de la molleja. Sin embargo, la LR del intestino delgado fue superior (P < 0,05) para las pollitas alimentadas con trigo que para las pollitas alimentadas con maíz. La LR del TGI se redujó (P < 0,05) al aumentar el tamaño de partícula del cereal. Se concluye que el trigo puede incluirse 500 g/kg en piensos de pollitas de 1 a 120 días de edad y que el tamaño de partícula de los cereales afecta el crecimiento de las pollitas durante los primeros 45 d de vida, pero no después. Por lo tanto, se recomienda moler el cereal utilizado al inicio del período de recría (1 a 45 d de edad) con una criba de diámetro igual o inferior a 8 mm. En el experimento 3 se utilizaron un total de 1.152 pollitas rubias Hy-Line de 1 d de edad para estudiar la influencia del nivel de energía de la dieta y la presentación del pienso sobre la productividad y las características del TGI. De 1 a 45 d de edad se utilizaron 6 piensos organizados de forma factorial con 3 concentraciones energéticas (baja: 11,44 MJ; media: 12,05 MJ y alta: 12,66 MJ/kg) y 2 presentaciones del pienso (harina vs. gránulo). De 45 a 120 d todos los piensos experimentales se suministraron en forma de harina y por tanto, la única diferencia entre tratamientos fue el nivel de EMAn utilizado. Cada uno de los 6 tratamientos se replicó 8 veces y la unidad experimental fue la jaula con 24 pollitas. De 1 a 120 d de edad, la ganancia de PV y el IC mejoraron a medida que aumentó la EMAn del pienso (P < 0,001). Las pollitas alimentadas con gránulo de 1 a 45 d de edad comieron mas y tuvieron una ganancia de peso superior (P < 0,001) que las alimentadas con harina. En el global de la prueba, la ganancia de PV fue mayor (P < 0,01) para las pollitas alimentadas con piensos en gránulo. A los 45 d de edad, el PR de todos los segmentos del TGI estudiados fue inferior para las pollitas alimentadas con piensos de alta energía que para las pollitas alimentadas con piensos de media o baja energía. A 120 d de edad, el PR de la molleja fue superior (P < 0,01) para las pollitas alimentadas con piensos de baja energía que con los otros piensos. Sin embargo, la LR del TGI no se vió afectada por el nivel de energía de los piensos. A los 45 d de edad, la alimentación con gránulo redujo el PR del proventrículo (P < 0,05), de la molleja (P < 0,001) y del TGI (P < 0.001), así como la LR del intestino delgado (P < 0,05) y de los ciegos (P < 0,001). A pesar de que las pollitas solo recibieron los piensos en gránulo durante los primeros 45 d de vida, la alimentación con gránulos redujo el PR de la molleja y del proventrículo a 120 d de edad. Se concluye que la alimentación con gránulos durante los primeros 45 d de vida mejora el consumo de pienso y el PV de las pollitas a 120 d de edad. Un aumento del nivel de energía de la dieta de 12,0 a 12,7 MJ/kg mejora los parámetros productivos de 1 a 120 d de edad pero reduce el tamaño del proventrículo y de la molleja. En base de estos resultados concluimos que maíz y trigo con enzimas pueden utilizarse indistintamente en piensos para pollitas de 1 a 120 d de edad con sólo una ligera disminución del PV final con trigo. La granulación y la reducción del tamaño de partícula del cereal del pienso de primera edad (1 a 45 d de vida) y el uso de piensos de alta densidad energética, mejoran los PV a 120 d de edad. Por lo tanto, es recomendable moler los cereales con cribas de no más de 8-mm de diámetro. También, la granulación del pienso y el uso de piensos de alta energía (pobres en fibra bruta) pueden reducir el desarrollo del TGI especialmente de la molleja, lo que puede perjudicar el consumo posterior de pienso durante el inicio del ciclo de puesta. ABSTRACT The general objective of this Thesis was to study the effect of different nutritional factors on productive performance and the development of the gastrointestinal tract (GIT) of commercial brown egg-laying pullets from 1 to 120 d of age. In this respect, the influence of type and particle size of the cereal, and feed form, and energy content of the die,t were studied in 3 experiments. In experiment 1, the influence of the main cereal and feed form of the diet on performance and GIT traits was studied in 576 brown-egg laying pullets from 1 to 120 d of age. From 1 to 45 d of age, 4 diets arranged factorially with 2 cereals (maize vs. wheat) and 2 feed forms (mash vs. pellets) were used. Each treatment was replicated 6 times (24 pullets per replicate). From 46 to 120 d of age, all diets were offered in mash form and therefore, the only difference among diets was the cereal used. Cumulatively, pullets fed the maize diets had higher body weight (BW) gain (P< 0.05) but similar feed conversion ratio (FCR) than pullets fed the wheat diets. From 1 to 45 d of age, pullets fed pellets consumed more feed (P < 0.001) and had higher BW gain (P < 0.001) than pullets fed mash. Most of the beneficial effects of pelleting on productive performance of the birds were still evident at 120 d of age. At 45 d of age, gizzard relative weight (RW; g/kg BW) was higher (P < 0.01) in pullets fed maize than in pullets fed wheat. Feeding pellets reduced the RW of the GIT and the gizzard (P < 0.001) as well as the relative length (RL; cm/kg BW) of the small intestine (SI, P < 0.01) at both ages. The pH of the gizzard contents at 120 d of age was not affected by the main cereal of the diet, but was lower in pullets fed mash from 1 to 45 d of age (P < 0.01) than in pullets fed pellets. We conclude that wheat supplemented with enzymes can be used in substitution of maize in pullet diets with only a slight reduction in BW gain at 120 d of age. Also, feeding pellets from 1 to 45 d of age increased BW gain and pH of the gizzard, and reduced the RW of the gizzard and the RL of the GIT at 120 d of age. In experiment 2, a total of 864 brown-egg laying pullets was used to study the effects of the main cereal of the diet (500 g maize or wheat/kg) and particle size of the cereal (hammer milled to pass through a 6, 8, and 10-mm screen) on growth performance and GIT traits from 1 to 120 d of age. Each of the 6 treatments was replicated 6 times (24 pullets per replicate). Type of cereal did not affect pullet performance at any age. From 1 to 45 d of age, BW gain was increased (P < 0.001) and FCR was improved (P < 0.05) as the particle size of the cereal was reduced, but no effects were observed after this age. At 45 d of age, pullets fed maize tended (P < 0.10) to have a heavier RW of the GIT and proventriculus and a higher relative length (RL, cm/kg BW) of the SI than pullets fed wheat. Also at this age, the RW of the GIT increased (P < 0.05) with increases in particle size of the cereal. At 120 d of age, dietary treatment did not affect the RW of any of the organs studied or gizzard pH, but the RL of the SI was higher (P < 0.05) for pullets fed wheat than for pullets fed maize. Also, the RL of the SI was reduced (P < 0.05) as the particle size of the cereal increased. We conclude that 500 g wheat/kg can be included in pullet feeds from 1 to 120 d of age, and that particle size of the cereal affects pullet performance during the first 45 d of life but not thereafter. Therefore, it is recommended to grind the cereal used in this period with a screen size of no more than 8-mm. In experiment 3, a total of 1,152 one-day-old Hy-Line Brown egg laying pullets were used to study the influence of the energy content of the diet and feed form on productive performance and on several GIT traits. From 1 to 45 d of age, there were 6 diets arranged factorially with 3 concentrations of AMEn (low: 11.66 MJ/kg, medium: 12.05 MJ/kg and high: 12.66 MJ/kg) of the diet and 2 feed forms (mash vs. pellets). From 45 to 120 d all diets were fed in mash form and therefore, the only difference among treatments in this period was the energy content of the diets. Each of the 6 treatments was replicated 8 times and the experimental unit was formed by 24 pullets. Cumulatively, BW gain and FCR improved as the AMEn of the diet increased (P < 0.001). Also, pullets fed pellets from 1 to 45 d of age had higher feed intake and BW gain (P < 0.001) in this period and higher cumulative BW gain (P < 0.01) than pullets fed mash. At 45 d of age, the RWof all the segments of the GIT was lower for pullets fed the high- than for pullets fed the medium- or low- energy diets. At 120 d of age, the RW of the gizzard was higher (P < 0.01) for pullets fed the low energy diets than for pullets fed the other diets. However, the RL of the GIT was not affected by the energy content of the diet. Feeding pellets reduced the RW of the proventriculus (P < 0.05), gizzard (P < 0.001), and GIT (P < 0.001), as well as the RL of the small intestine (P < 0.05) and the ceaca (P < 0.001) at 45 d of age. The effects of feeding pellets on RW of gizzard and proventriculus were still evident at 120 d of age. We concluded that feeding pellets from 1 to 45 d of age improved feed intake and BW of pullets at 120 d of age and that an increase in the energy content of the diet increased pullet performance at all ages but reduced the RW of the proventriculus and gizzard. We conclude that maize and wheat can be used indistinctly in diets for egg laying pullets from 1 to 120 d of age, with only a slight reduction in final BW when wheat is used. Also, particle size of the cereal affects pullet performance during the first 45 d of life but not thereafter. Pelleting of the feeds, and grinding the cereal with a screen size of no more than 8-mm from 1 to 45 d of age, and the use of high density energy diets are recommended in order to achieve adequate target BW at 120 d of age. However, pelleting of the feed, very fine grinding, and the use of high AMEn diets might hinder the development of the GIT, especially that of the gizzard, which might affect feed intake of laying hens especially at the beginning of the production cycle.

Crop rotation and residues influence N2O emission and N efficiency rather than tillage under a rainfed Mediterranean agroecosystem

Conservation tillage and crop rotation have spread during the last decades because promotes several positive effects (increase of soil organic content, reduction of soil erosion, and enhancement of carbon sequestration) (Six et al., 2004). However, these benefits could be partly counterbalanced by negative effects on the release of nitrous oxide (N2O) (Linn and Doran, 1984). There is a lack of data on long-term tillage system study, particularly in Mediterranean agro-ecosystems. The aim of this study was to evaluate the effects of long-term (>17 year) tillage systems (no tillage (NT), minimum tillage (MT) and conventional tillage (CT)); and crop rotation (wheat (W)-vetch (V)-barley (B)) versus wheat monoculture (M) on N2O emissions. Additionally, Yield-scaled N2O emissions (YSNE) and N uptake efficiency (NUpE) were assessed for each treatment.

The family of DOF transcription factors in Brachypodium distachyon: phylogenetic comparison with rice and barley DOFs and expression profiling

ABSTRACT: Transcription factors (TFs) are proteins that have played a central role both in evolution and in domestication, and are major regulators of development in living organisms. Plant genome sequences reveal that approximately 7% of all genes encode putative TFs. The DOF (DNA binding with One Finger) TF family has been associated with vital processes exclusive to higher plants and to their close ancestors (algae, mosses and ferns). These are seed maturation and germination, light-mediated regulation, phytohormone and plant responses to biotic and abiotic stresses, etc. In Hordeum vulgare and Oryza sativa, 26 and 30 different Dof genes, respectively, have been annotated. Brachypodium distachyon has been the first Pooideae grass to be sequenced and, due to its genomic, morphological and physiological characteristics, has emerged as the model system for temperate cereals, such as wheat and barley. RESULTS: Through searches in the B. distachyon genome, 27 Dof genes have been identified and a phylogenetic comparison with the Oryza sativa and the Hordeum vulgare DOFs has been performed. To explore the evolutionary relationship among these DOF proteins, a combined phylogenetic tree has been constructed with the Brachypodium DOFs and those from rice and barley. This phylogenetic analysis has classified the DOF proteins into four Major Cluster of Orthologous Groups (MCOGs). Using RT-qPCR analysis the expression profiles of the annotated BdDof genes across four organs (leaves, roots, spikes and seeds) has been investigated. These results have led to a classification of the BdDof genes into two groups, according to their expression levels. The genes highly or preferentially expressed in seeds have been subjected to a more detailed expression analysis (maturation, dry stage and germination). CONCLUSIONS: Comparison of the expression profiles of the Brachypodium Dof genes with the published functions of closely related DOF sequences from the cereal species considered here, deduced from the phylogenetic analysis, indicates that although the expression profile has been conserved in many of the putative orthologs, in some cases duplication followed by subsequent divergence may have occurred (neo-functionalization).

Role of maize stover incorporation on nitrogen oxide emissions in a non-irrigated Mediterranean barley field.

Aims Agricultural soils in semiarid Mediterranean areas are characterized by low organic matter contents and low fertility levels. Application of crop residues and/or manures as amendments is a cost-effective and sustainable alternative to overcome this problem. However, these management practices may induce important changes in the nitrogen oxide emissions from these agroecosystems, with additional impacts on carbon dioxide emissions. In this context, a field experiment was carried out with a barley (Hordeum vulgare L.) crop under Mediterranean conditions to evaluate the effect of combining maize (Zea mays L.) residues and N fertilizer inputs (organic and/or mineral) on these emissions. Methods Crop yield and N uptake, soil mineral N concentrations, dissolved organic carbon (DOC), denitrification capacity, N2O, NO and CO2 fluxes were measured during the growing season. Results The incorporation of maize stover increased N2O emissions during the experimental period by c. 105 %. Conversely, NO emissions were significantly reduced in the plots amended with crop residues. The partial substitution of urea by pig slurry reduced net N2O emissions by 46 and 39 %, with and without the incorporation of crop residues respectively. Net emissions of NO were reduced 38 and 17 % for the same treatments. Molar DOC:NO 3 − ratio was found to be a robust predictor of N2O and NO fluxes. Conclusions The main effect of the interaction between crop residue and N fertilizer application occurred in the medium term (4–6 month after application), enhancing N2O emissions and decreasing NO emissions as consequence of residue incorporation. The substitution of urea by pig slurry can be considered a good management strategy since N2O and NO emissions were reduced by the use of the organic residue.

Amino acid availability in ruminants of cereals and cereal co-products

Microbial corrected in situ estimatesof the ruminal undegraded fraction (RU) and intestinal effectivedigestibility (IED) of amino acids (AA), except tryptophan, of rye, wheat and corn grains, wheat bran, wheat and barley distilled dried grains and corn gluten feed were measured on three rumen- and duodenum-cannulated wethers using 15N-labelling techniques and considering ruminal rates of particle comminution and outflow.

Function of C1A barley peptidases and their inhibitors (cystatins) in barley seed germination : Funcion de las peptidasas C1A de cebada y sus inhibidores (cistatinas) en la germinacion de la semilla

Plant proteolysis is a metabolic process where specific enzymes called peptidases degrade proteins. In plants, this complex process involves broad metabolic networks and different sub-cellular compartments. Several types of peptidases take part in the proteolytic process, mainly cysteine-, serine-, aspartyl- and metallo- peptidases. Among the cysteine-peptidases, the papain-like or C1A peptidases (family C1, clan CA) are extensively present in land plants and are classified into catepsins L-, B-, H- and Flike. The catalytic mechanism of these C1A peptidases is highly conserved and involves the three amino acids Cys, His and Asn in the catalytic triad, and a Gln residue which seems essential for maintaining an active enzyme conformation. These proteins are synthesized as inactive precursors, which comprise an N-terminal signal peptide, a propeptide, and the mature protein. In barley, we have identified 33 cysteine-peptidases from the papain-like family, classifying them into 8 different groups. Five of them corresponded to cathepsins L-like (5 subgroups), 1 cathepsin B-like group, 1 cathepsin F-like group and 1 cathepsin H-like group. Besides, C1A peptidases are the specific targets of the plant proteinaceous inhibitors known as phytocystatins (PhyCys). The cystatin inhibitory mechanism is produced by a tight and reversible interaction with their target enzymes. In barley, the cystatin gene family is comprised by 13 members. In this work we have tried to elucidate the role of the C1A cysteine-peptidases and their specific inhibitors (cystatins) in the germination process of the barley grain. Therefore, we selected a representative member of each group/subgroup of C1A peptidases (1 cathepsin B-like, 1 cathepsin F-like, 1 cathepsin H-like and 5 cathepsins L-like). The molecular characterization of the cysteine-peptidases was done and the peptidase-inhibitor interaction was analyzed in vitro and in vivo. A study in the structural basis for specificity of pro-peptide/enzyme interaction in barley C1A cysteine-peptidases has been also carried out by inhibitory assays and the modeling of the three-dimensional structures. The barley grain maturation produces the accumulation of storage proteins (prolamins) in the endosperm which are mobilized during germination to supply the required nutrients until the photosynthesis is fully established. In this work, we have demonstrated the participation of the cysteine-peptidases and their inhibitors in the degradation of the different storage protein fractions (hordeins, albumins and globulins) present in the barley grain. Besides, transgenic barley plants overexpressing or silencing cysteine-peptidases or cystatins were obtained by Agrobacterium-mediated transformation of barley immature embryos to analyze their physiological function in vivo. Preliminary assays were carried out with the T1 grains of several transgenic lines. Comparing the knock-out and the overexpressing lines with the WT, alterations in the germination process were detected and were correlated with their grain hordein content. These data will be validated with the homozygous grains that are being produced through the double haploid technique by microspore culture. Resumen La proteólisis es un proceso metabólico por el cual se lleva a cabo la degradación de las proteínas de un organismo a través de enzimas específicas llamadas proteasas. En plantas, este complejo proceso comprende un entramado de rutas metabólicas que implican, además, diferentes compartimentos subcelulares. En la proteólisis participan numerosas proteasas, principalmente cisteín-, serín-, aspartil-, y metalo-proteasas. Dentro de las cisteín-proteasas, las proteasas tipo papaína o C1A (familia C1, clan CA) están extensamente representadas en plantas terrestres, y se clasifican en catepsinas tipo L, B, H y F. El mecanismo catalítico de estas proteasas está altamente conservado y la triada catalítica formada por los aminoácidos Cys, His y Asn, y a un aminoácido Gln, que parece esencial para el mantenimiento de la conformación activa de la proteína. Las proteasas C1A se sintetizan como precursores inactivos y comprenden un péptido señal en el extremo N-terminal, un pro-péptido y la proteína madura. En cebada hemos identificado 33 cisteín-proteasas de tipo papaína y las hemos clasificado filogenéticamente en 8 grupos diferentes. Cinco de ellos pertenecen a las catepsinas tipo L (5 subgrupos), un grupo a las catepsinas tipo-B, otro a las catepsinas tipo-F y un último a las catepsinas tipo-H. Las proteasas C1A son además las dianas específicas de los inhibidores protéicos de plantas denominados fitocistatinas. El mecanismo de inhibición de las cistatinas está basado en una fuerte interacción reversible. En cebada, se conoce la familia génica completa de las cistatinas, que está formada por 13 miembros. En el presente trabajo se ha investigado el papel de las cisteín-proteasas de cebada y sus inhibidores específicos en el proceso de la germinación de la semilla. Para ello, se seleccionó una proteasa representante de cada grupo/subgrupo (1 catepsina tipo- B, 1 tipo-F, 1 tipo-H, y 5 tipo-L, una por cada subgrupo). Se ha llevado a cabo su caracterización molecular y se ha analizado la interacción enzima-inhibidor tanto in vivo como in vitro. También se han realizado estudios sobre las bases estructurales que demuestran la especificidad en la interacción enzima/propéptido en las proteasas C1A de cebada, mediante ensayos de inhibición y la predicción de modelos estructurales de la interacción. Finalmente, y dado que durante la maduración de la semilla se almacenan proteínas de reserva (prolaminas) en el endospermo que son movilizadas durante la germinación para suministrar los nutrientes necesarios hasta que la nueva planta pueda realizar la fotosíntesis, en este trabajo se ha demostrado la participación de las cisteínproteasas y sus inhibidores en la degradación de las diferentes tipos de proteínas de reserva (hordeinas, albúmins y globulinas) presentes en el grano de cebada. Además, se han obtenido plantas transgénicas de cebada que sobre-expresan o silencian cistatinas y cisteín-proteasas con el fin de analizar la función fisiológica in vivo. Se han realizado análisis preliminares en las semillas T1 de varias líneas tránsgenicas de cebada y al comparar las líneas knock-out y las líneas de sobre-expresión con las silvestres, se han detectado alteraciones en la germinación que están además correlacionadas con el contenido de hordeinas de las semillas. Estos datos serán validados en las semillas homocigotas que se están generando mediante la técnica de dobles haploides a partir del cultivo de microesporas.

Effect of fibers and whole grain content on quality attributes

Incorporation of fiber in cereals may lead to quality issues, thus decreasing consumer acceptance. This is partially due to deterioration of the microstructure, one of the primary quality attributes of cereals. The objective of this study was to better understand the mechanisms by which dietary fibers affect the quality of cereal products during extrusioncooking. The study quantified the effect of amount and type of fiber and whole grain on (i) texture, (ii) structure, and (iii) rehydration properties of extruded cereals. New innovative methods were applied and combined with traditional techniques to characterize both the structure and the rehydration properties. Extruded cereals were produced using a starch-based recipe (whole and wheat flours) and two sources of fibers (oat bran concentrate and wheat bran). The oat and wheat bran levels used in this study were 0, 10, and 20%. The different mixtures were extruded in a pilot twinscrew extruder BC21 (Clextral) and then sugar coated after drying. Mechanical properties of extruded cereals were investigated by compression test. The cellular structure was observed by X-ray tomography. The quality of coating (thickness, homogeneity) was analyzed by optical coherence tomography. The rehydration properties of such cereals in milk were evaluated by magnetic resonance imaging and optical coherence tomography. This work revealed that structure assessment of extruded cereals may lead to a better understanding of the effect of fiber addition on texture and rehydration properties. The application of innovative methods, such as optical coherence tomography and magnetic resonance imaging, was found to be useful to quantify the structural properties.

Effects of the main cereal and type of fat of the diet on productive performance and egg quality of brown egg laying hens from twenty two to fifty four weeks of age

The influence of the main cereal and supplemental fat of the diet on productive performance and egg quality was studied in 756 brown-egg laying hens from 22 to 54 weeks of age. The experiment was conducted as a completely randomized design with 9 treatments arranged factorially with 3 cereals (dented corn, soft wheat, and barley) and 3 types of fat [soy oil (SBO), acidulated soapstocks (AOS), and lard].

CSM-IXIM: A New Maize Simulation Model for DSSAT Version 4.5

Th e CERES-Maize model is the most widely used maize (Zea mays L.) model and is a recognized reference for comparing new developments in maize growth, development, and yield simulation. Th e objective of this study was to present and evaluate CSMIXIM, a new maize simulation model for DSSAT version 4.5. Code from CSM-CERES-Maize, the modular version of the model, was modifi ed to include a number of model improvements. Model enhancements included the simulation of leaf area, C assimilation and partitioning, ear growth, kernel number, grain yield, and plant N acquisition and distribution. Th e addition of two genetic coeffi cients to simulate per-leaf foliar surface produced 32% smaller root mean square error (RMSE) values estimating leaf area index than did CSM-CERES. Grain yield and total shoot biomass were correctly simulated by both models. Carbon partitioning, however, showed diff erences. Th e CSM-IXIM model simulated leaf mass more accurately, reducing the CSM-CERES error by 44%, but overestimated stem mass, especially aft er stress, resulting in similar average RMSE values as CSM-CERES. Excessive N uptake aft er fertilization events as simulated by CSM-CERES was also corrected, reducing the error by 16%. Th e accuracy of N distribution to stems was improved by 68%. Th ese improvements in CSM-IXIM provided a stable basis for more precise simulation of maize canopy growth and yield and a framework for continuing future model developments

Influence of nutritional and management practices on productive performance and egg quality of brown egg-laying hens = Influencia de factores nutricionales y de manejo sobre la productividad y calidad del huevo en gallinas ponedoras rubias

El objetivo general de esta Tesis Doctoral fue estudiar la influencia de diversos factores nutricionales y de manejo sobre la productividad y la calidad del huevo en gallinas ponedoras comerciales rubias. Los factores estudiados fueron: 1) Cereal principal y tipo de grasa en la dieta; 2) Nivel de proteína bruta y grasa en la dieta; 3) Nivel energético de la dieta; 4) Peso vivo al inicio del período de puesta. En el experimento 1, la influencia del cereal principal en la dieta y el tipo de grasa suplementada en la dieta sobre los parámetros productivos y la calidad del huevo fue estudiado en 756 gallinas rubias de la estirpe Lohmann desde la sem 22 hasta las 54 de vida. El experimento se realizó mediante un diseño completamente al azar con 9 tratamientos ordenados factorialmente, con 3 cereales bases (maíz, trigo blando y cebada) y 3 tipos de grasa que variaban en su contenido en ácido linoléico (aceite de soja, oleína vegetal mezcla y manteca). Todas las dietas satisfacian las recomendaciones nutricionales para gallinas ponedoras rubias según el NRC (1994) y FEDNA (2008). La unidad experimental fue la jaula para todas las variables. Cada tratamiento fue replicado 4 veces, y la unidad experimental estuvo formada por 21 gallinas alojadas en grupos de 7. Las dietas fueron formuladas con un contenido nutritivo similar, excepto para el ácido linoléico, que varió en función del tipo de cereal y grasa utilizado. Así, dependiendo de la combinación de estos elementos el contenido de este ácido graso varió desde un 0.8% (dieta trigo-manteca) a un 3.4% (dieta maíz-aceite de soja). Este rango de ácido linoléico permitió estimar el nivel mínimo de este nutriente en el pienso que permite maximizar el peso del huevo. Los parámetros productivos y la calidad del huevo se controlaron cada 28 días y el peso de las aves se midió individualmente al inicio y al final del experimento con el objetivo de estudiar la variación en el peso vivo de los animales. No se observaron interacciones entre el tipo de cereal y grasa en la dieta para ninguna de las variables productivas estudiadas. Los tratamientos experimentales no afectaron a las principales variables productivas (porcentaje de puesta, peso del huevo y masa de huevo). Sin embargo, la ganancia de peso fue mayor en gallinas alimentadas con maíz o trigo que las gallinas alimentadas con cebada (243 vs. 238 vs. 202 g, respectivamente; P< 0.05). En el mismo sentido, las gallinas alimentadas con manteca obtuvieron una mayor ganancia de peso que las gallinas alimentadas con aceite de soja u oleína vegetal (251 vs. 221 vs. 210 g, respectivamente; P< 0.05). En cuanto a las variables estudiadas en relación con la calidad del huevo, ninguna de las variables estudiadas se vio afectada por el tratamiento experimental, salvo la pigmentación de la yema. Así, las gallinas alimentadas con maíz como cereal principal obtuvieron una mayor puntuación en relación con la escala de color que las gallinas alimentadas con trigo y con cebada (9.0 vs. 8.3 vs. 8.3, respectivamente; P< 0.001). La pigmentación de la yema también se vio afectada por el tipo de grasa en la dieta, así, las gallinas alimentadas con manteca obtuvieron una mayor puntuación de color en relación con la escala de color que las gallinas alimentadas con aceite de soja u oleína vegetal (8.9 vs. 8.5 vs. 8.2, respectivamente; P< 0.001). La influencia del contenido en ácido linoléico respecto al peso de huevo y masa de huevo fue mayor a medida que el contenido de dicho ácido graso se redujo en la dieta. Así, la influencia de la dieta en los radios peso de huevo/g linoléico ingerido y masa de huevo/g linoléico ingerido fue significativamente mayor a medida que el contenido en dicho ácido graso disminuyo en la dieta (P< 0.001). Los resultados del ensayo indican que las gallinas ponedoras rubias no necesitan más de un 1.0% de ácido linoléico en la dieta para maximizar la producción y el tamaño del huevo. Además, se pudo concluir que los 3 cereales y las 3 grasas utilizadas pueden sustituirse en la dieta sin ningún perjuicio productivo o referente a la calidad del huevo siempre que los requerimientos de los animales sean cubiertos. En el experimento 2, la influencia del nivel de proteína bruta y el contenido de grasa de la dieta sobre los parámetros productivos y la calidad del huevo fue estudiado en 672 gallinas ponedoras rubias de la estirpe Lohmann entre las sem 22 y 50 de vida. El experimento fue conducido mediante un diseño completamente al azar con 8 tratamientos ordenados factorialmente con 4 dietas y 2 pesos vivos distintos al inicio de puesta (1592 vs. 1860g). Tres de esas dietas diferían en el contenido de proteína bruta (16.5%, 17.5% y 18.5%) y tenían un contenido en grasa añadida de 1.8%. La cuarta dieta tenía el nivel proteico más elevado (18.5%) pero fue suplementada con 3.6% de grasa añadida en vez de 1.8%. Cada tratamiento fue replicado 4 veces y la unidad experimental consistió en 21 gallinas alojadas dentro de grupos de 7 animales en 3 jaulas contiguas. Todas las dietas fueron isocalóricas (2750 kcal EMAn/kg) y cubrieron las recomendaciones en aminoácidos para gallinas ponedoras rubias (Arg, Ile, Lys, Met, Thr, Trp, TSAA y Val) según el NRC (1994) y FEDNA (2008). Los efectos de los tratamientos sobre las variables productivas y la calidad de huevo fueron estudiados cada 28 días. La dieta no afecto a ninguna de las variables productivas estudiadas a lo largo del período productivo. Sin embargo, el peso inicial origino que las gallinas pesadas consumieran más (120.6 vs. 113.9 g; P< 0.001), obtuvieran un porcentaje de puesta mayor (92.5 vs. 89.8%; P< 0.01) y un peso del huevo mayor (64.9 vs. 62.4 g; P< 0.001) que las gallinas ligeras. El peso inicial de las gallinas no afecto al IC por kg de huevo ni a la mortalidad, sin embargo, la ganancia de peso fue mayor (289 vs. 233 g; P< 0.01) y el IC por docena de huevos fue mejor (1.52 vs. 1.57; P< 0.01) en las gallinas ligeras que en las gallinas pesadas. En cuanto a la calidad del huevo, la dieta no influyó sobre ninguna de las variables estudiadas. Los resultados del ensayo muestran que las gallinas ponedoras rubias, independientemente de su peso vivo al inicio de la puesta, no necesitan una cantidad de proteína bruta superior a 16.5% para maximizar la producción, asegurando que las dietas cubren los requerimientos en AA indispensables. Asimismo, se puedo concluir que las gallinas con un peso más elevado al inicio de puesta producen más masa de huevo que las gallinas con un peso más bajo debido a que las primeras producen más cantidad de huevos y más pesados. Sin embargo, ambos grupos de peso obtuvieron el mismo IC por kg de huevo y las gallinas más livianas en peso obtuvieron un mejor IC por docena de huevo que las pesadas. En el experimento 3 la influencia de la concentración energética sobre los parámetros productivos y la calidad del huevo fue estudiada en 520 gallinas ponedoras rubias de la estirpe Hy-Line en el período 24-59 sem de vida. Se utilizaron 8 tratamientos ordenados factorialmente con 4 dietas que variaron en el contenido energético (2650, 2750, 2850 y 2950 kcal EMAn/kg) y 2 pesos vivos distintos al inicio del período de puesta (1733 vs. 1606g). Cada tratamiento fue replicado 5 veces y la unidad experimental consistió en una jaula con 13 aves. Todas las dietas se diseñaron para que tuvieran una concentración nutritiva similar por unidad energética. Las variables productivas y de calidad de huevo se estudiaron mediante controles cada 28 días desde el inicio del experimento. No se observaron interacciones entre el nivel energético y el peso inicial del ave para ninguna de las variables estudiadas. Un incremento en la concentración energética de la dieta incrementó la producción de huevos (88.8 % vs. 91.2 % vs. 92.7 % vs. 90.5 %), masa de huevo (56.1 g/d vs. 58.1 g/d vs. 58.8 g/d vs. 58.1 g/d), y eficiencia energética (5.42 vs. 5.39 vs. 5.38 vs. 5.58 kcal EMA/g huevo) de forma lineal y cuadrática (P< 0.05) y afectó significativamente a la ganancia de peso (255 g vs. 300 g vs. 325 g vs. 359 g; P<0.05) . Sin embargo, un incremento en la concentración energética provocó un descenso lineal en el consumo de los animales (115 g vs. 114 g vs. 111 g vs. 110 g; P< 0.001) y un descenso lineal y cuadrático en el IC por kg de huevo (2.05 vs. 1.96 vs. 1.89 vs. 1.89; P< 0.01). En cuanto a la calidad del huevo, un incremento en el contenido energético de la dieta provocó una reducción en la calidad del albumen de forma lineal en forma de reducción de Unidades Haugh (88.4 vs. 87.8 vs. 86.3 vs. 84.7; P< 0.001), asimismo el incremento de energía redujo de forma lineal la proporción relativa de cáscara en el huevo (9.7 vs. 9.6 vs. 9.6 vs. 9.5; P< 0.001). Sin embargo, el incremento energético propició un incremento lineal en la pigmentación de la yema del huevo (7.4 vs. 7.4 vs. 7.6 vs. 7.9; P< 0.001). El peso vivo al inicio de la prueba afecto a las variables productivas y a la calidad del huevo. Así, los huevos procedentes de gallinas pesadas al inicio de puesta tuvieron una mayor proporción de yema (25.7 % vs. 25.3 %; P< 0.001) y menor de albumen (64.7 vs. 65.0; P< 0.01) y cáscara (9.5 vs. 9.6; P< 0.05) respecto de los huevos procedentes de gallinas ligeras. Consecuentemente, el ratio yema:albumen fue mayor (0.40 vs. 0.39; P< 0.001) para las gallinas pesadas. Según los resultados del experimento se pudo concluir que las actuales gallinas ponedoras rubias responden con incrementos en la producción y en la masa del huevo a incrementos en la concentración energética hasta un límite que se sitúa en 2850 kcal EMAn/kg. Asimismo, los resultados obtenidos entre los 2 grupos de peso al inicio de puesta demostraron que las gallinas pesadas al inicio de puesta tienen un mayor consumo y producen huevos más pesados, con el consecuente aumento de la masa del huevo respecto de gallinas más ligeras. Sin embargo, el IC por kg de huevo fue el mismo en ambos grupos de gallinas y el IC por docena de huevo fue mejor en las gallinas ligeras. Asimismo, la eficiencia energética fue mejor en las gallinas ligeras. Abstract The general aim of this PhD Thesis was to study the influence of different nutritional factors and management on the productivity and egg quality of comercial Brown laying hens. The factor studied were: 1) The effect of the main cereal and type of fat of the diet; 2) The effect of crude protein and fat content of the diet; 3) The effect of energy concentration of the diet; 4) The effect of initial body weight of the hens at the onset of lay period. In experiment 1, the influence of the main cereal and type of supplemental fat in the diet on productive performance and egg quality of the eggs was studied in 756 Lohmann brown-egg laying hens from 22 to 54 wk of age. The experiment was conducted as a completely randomized design with 9 treatments arranged factorially with 3 cereals (dented corn, soft wheat, and barley) and 3 types of fat (soy oil, acidulated vegetable soapstocks, and lard). Each treatment was replicated 4 times (21 hens per replicate). All diets were formulated according to NRC (1994) and FEDNA (2008) to have similar nutrient content except for linoleic acid that ranged from 0.8 (wheat-lard diet) to 3.4% (corn-soy bean oil) depending on the combination of cereal and fat source used. This approach will allow to estimate the minimum level of linoleic acid in the diets that maximizes egg weight. Productive performance and egg quality traits were recorded every 28 d and BW of the hens was measured individually at the beginning and at the end of the experiment. No significant interactions between main factors were detected for any of the variables studied. Egg production, egg weight, and egg mass were not affected by dietary treatment. Body weight gain was higher (243 vs. 238 vs. 202 g; P<0.05) for hens fed corn or wheat than for hens fed barley and also for hens fed lard than for hens fed soy oil or acidulated vegetable soapstocks (251 vs. 221 vs. 210 g; P< 0.05). Egg quality was not influenced by dietary treatment except for yolk color that was greater (9.0 vs. 8.3 vs. 8.3; P< 0.001) for hens fed corn than for hens fed wheat or barley and for hens fed lard than for hens fed soy oil or acidulated vegetable soapstocks (8.9 vs. 8.5 vs. 8.2, respectivamente; P< 0.001). The influence of linoleic acid on egg weight and egg mass was higher when the fatty acid was reduced in the diet. Thus, the influence of the diet in egg weight/g linoleic acid intake and egg mass/g linolec acid intake was higher when the amount of this fatty acid decreased in the diet (P< 0.001). It is concluded that brown egg laying hens do not need more than 1.0% of linoleic acid in the diet (1.16 g/hen/d) to maximize egg production and egg size. The 3 cereals and the 3 fat sources tested can replace each other in the diet provided that the linoleic acid requirements to maximize egg size are met. In experiment 2, the influence of CP and fat content of the diet on performance and egg quality traits was studied in 672 Lohmann brown egg-laying hens from 22 to 50 wk of age. The experiment was conducted as a completely randomized design with 8 treatments arranged factorially with 4 diets and 2 initial BW of the hens (1,592 vs. 1,860 g). Three of these diets differed in the CP content (16.5, 17.5, and 18.5%) and included 1.8% added fat. The fourth diet had also 18.5% CP but was supplemented with 3.6% fat instead of 1.8% fat. Each treatment was replicated 4 times and the experimental unit consisted of 21 hens allocated in groups of 7 in 3 adjacent cages. All diets were isocaloric (2,750 kcal AME/kg) and met the recommendations of brown egg-laying hens for digestible Arg, Ile, Lys, Met, Thr, Trp, TSAA, and Val. Productive performance and egg quality were recorded by replicate every 28-d. For the entire experimental period, diet did not affect any of the productive performance traits studied but the heavier hens had higher ADFI (120.6 vs. 113.9g; P< 0.001), egg production (92.5 vs. 89.8%; P< 0.01), and egg weight (64.9 vs. 62.4g; P< 0.001) than the lighter hens. Initial BW did not affect feed conversion per kilogram of eggs or hen mortality but BW gain was higher (289 vs. 233g; P< 0.01) and FCR per dozen of eggs was better (1.52 vs. 1.57; P< 0.01) for the lighter than for the heavier hens. None of the egg quality variables studied was affected by dietary treatment or initial BW of the hens. It is concluded that brown egg-laying hens, irrespective of their initial BW, do not need more than 16.5% CP to maximize egg production provided that the diet meet the requirements for key indispensable amino acids. Heavier hens produce more eggs that are larger than lighter hens but feed efficiency per kilogram of eggs is not affected. In experiment 3, the influence of AMEn concentration of the diet on productive performance and egg quality traits was studied in 520 Hy-Line brown egg-laying hens differing in initial BW from 24 to 59 wks of age. There were 8 treatments arranged factorially with 4 diets varying in energy content (2,650, 2,750, 2,850, and 2,950 kcal AMEn/kg) and 2 initial BW of the hens (1,733 vs. 1,606 g). Each treatment was replicated 5 times (13 hens per replicate) and all diets had similar nutrient content per unit of energy. No interactions between energy content of the diet and initial BW of the hens were detected for any trait. An increase in energy concentration of the diet increased (linear, P< 0.05; quadratic P< 0.05) egg production (88.8 % vs. 91.2 % vs. 92.7 % vs. 90.5 %), egg mass (56.1 g/d vs. 58.1 g/d vs. 58.8 g/d vs. 58.1 g/d), energy efficiency (5.42 vs. 5.39 vs. 5.38 vs. 5.58 kcal AMEn/g of egg), and BW gain (255 g vs. 300 g vs. 325 g vs. 359 g; P<0.05) but decreased ADFI (115 g vs. 114 g vs. 111 g vs. 110 g; P< linear, P< 0.001) and FCR per kg of eggs (2.05 vs. 1.96 vs. 1.89 vs. 1.89; linear, P< 0.01; quadratic P< 0.01). An increase in energy content of the diet reduced Haugh units (88.4 vs. 87.8 vs. 86.3 vs. 84.7; P< 0.01) and the proportion of shell in the egg (9.7 vs. 9.6 vs. 9.6 vs. 9.5; P< 0.001). Feed intake (114.6 vs. 111.1 g/hen per day), AMEn intake (321 vs. 311 kcal/hen per day), egg weight (64.2 vs. 63.0 g), and egg mass (58.5 vs. 57.0 g) were higher for the heavier than for the lighter hens (P<0.01) but FCR per kg of eggs and energy efficiency were not affected. Eggs from the heavier hens had higher proportion of yolk (25.7 % vs. 25.3 %; P< 0.001) and lower of albumen (64.7 vs. 65.0; P< 0.01) and shell (9.5 vs. 9.6; P< 0.05) than eggs from the lighter hens. Consequently, the yolk to albumen ratio was higher (0.40 vs. 0.39; P< 0.001) for the heavier hens. It is concluded that brown egg-laying hens respond with increases in egg production and egg mass, to increases in AMEn concentration of the diet up to 2,850 kcal/kg. Heavy hens had higher feed intake and produced heavier eggs and more egg mass than light hens. However, energy efficiency was better for the lighter hens.

Phylogenetic analyses and toxigenic profiles of Fusarium equiseti and Fusarium acuminatum isolated from cereals from Southern Europe

Fusarium equiseti and Fusarium acuminatum are toxigenic species that contaminate cereal crops from diverse climatic regions. They are common in Spanish cereals. The information available on their phylogenetics and toxigenic profiles is, however, insufficient to assist risk evaluation. In this work, phylogenetic analyses were performed using partial sequences of the translation elongation factor gene (EF-1a) of F. equiseti and F. acuminatum strains isolated from barley and wheat from Spain and other countries. The Northern and Southern European F. equiseti strains largely separated into two phylogenetically distinct clusters. This suggests the existence of two distinct populations within this species, explaining its presence in these regions of markedly different climate. Production of type A and B trichothecenes by the Spanish strains, examined in wheat cultures using a multitoxin analytical method, indicated that F. equiseti could produce deoxynivalenol and nivalenol and other trichothecenes, at concentrations that might represent a significant risk of toxin contamination for Southern European cereals. F. acuminatum showed low intraspecific genetic variability and 58% of the strains could produce deoxynivalenol at low level. Neither species was found to produce T-2 or HT-2 toxins. The present results provide important phylogenetic and toxigenic information essential for the accurate prediction of toxigenic risk.

Calibration of WAVE in irrigated maize: fallow vs. cover crops.

Nitrate leaching decreases crop available N and increases water contamination. Replacing fallow by cover crops (CC) is an alternative to reduce nitrate contamination, because it reduces overall drainage and soil mineral N accumulation. A study of the soil N and nitrate leaching was conducted during 5 years in a semi-arid irrigated agricultural area of Central Spain. Three treatments were studied during the intercropping period of maize (Zea mays L.): barley (Hordeum vulgare L.), vetch (Vicia villosa L.), and fallow. Cover crops, sown in October, were killed by glyphosate application in March, allowing direct seeding of maize in April. All treatments were irrigated and fertilised following the same procedure. Soil water content was measured using capacity probes. Soil Nmin accumulation was determined along the soil profile before sowing and after harvesting maize. Soil analysis was conducted at six depths every 0.20m in each plot in samples from 0 to 1.2-m depth. The mechanistic water balance model WAVE was applied in order to calculate drainage and plant growth of the different treatments, and apply them to the N balance. We evaluated the water balance of this model using the daily soil water content measurements of this field trial. A new Matlab version of the model was evaluated as well. In this new version improvements were made in the solute transport module and crop module. In addition, this new version is more compatible with external modules for data processing, inverse calibration and uncertainty analysis than the previous Fortran version. The model showed that drainage during the irrigated period was minimized in all treatments, because irrigation water was adjusted to crop needs, leading to nitrate accumulation on the upper layers after maize harvest. Then, during the intercrop period, most of the nitrate leaching occurred. Cover crops usually led to a shorter drainage period, lower drainage water amount and lower nitrate leaching than the treatment with fallow. These effects resulted in larger nitrate accumulation in the upper layers of the soil after CC treatments.

Probabilistic analysis of water availability for agriculture and associated crop net margins.

Rising water demands are difficult to meet in many regions of the world. In consequence, under meteorological adverse conditions, big economic losses in agriculture can take place. This paper aims to analyze the variability of water shortage in an irrigation district and the effect on farmer?s income. A probabilistic analysis of water availability for agriculture in the irrigation district is performed, through a supply-system simulation approach, considering stochastically generated series of stream-flows. Net margins associated to crop production are as well estimated depending on final water allocations. Net margins are calculated considering either single-crop farming, either a polyculture system. In a polyculture system, crop distribution and water redistribution are calculated through an optimization approach using the General Algebraic Modeling System (GAMS) for several scenarios of irrigation water availability. Expected net margins are obtained by crop and for the optimal crop and water distribution. The maximum expected margins are obtained for the optimal crop combination, followed by the alfalfa monoculture, maize, rice, wheat and finally barley. Water is distributed as follows, from biggest to smallest allocation: rice, alfalfa, maize, wheat and barley.