Interactive effects of ocean acidification, elevated temperature, and reduced salinity on early-life stages of the Pacific oyster

Ocean acidification (OA) effects on larvae are partially attributed for the rapidly declining oyster production in the Pacific Northwest region of the United States. This OA effect is a serious concern in SE Asia, which produces >80% of the world's oysters. Because climate-related stressors rarely act alone, we need to consider OA effects on oysters in combination with warming and reduced salinity. Here, the interactive effects of these three climate-related stressors on the larval growth of the Pacific oyster, Crassostrea gigas, were examined. Larvae were cultured in combinations of temperature (24 and 30 °C), pH (8.1 and 7.4), and salinity (15 psu and 25 psu) for 58 days to the early juvenile stage. Decreased pH (pH 7.4), elevated temperature (30 °C), and reduced salinity (15 psu) significantly delayed pre- and post-settlement growth. Elevated temperature lowered the larval lipid index, a proxy for physiological quality, and negated the negative effects of decreased pH on attachment and metamorphosis only in a salinity of 25 psu. The negative effects of multiple stressors on larval metamorphosis were not due to reduced size or depleted lipid reserves at the time of metamorphosis. Our results supported the hypothesis that the C. gigas larvae are vulnerable to the interactions of OA with reduced salinity and warming in Yellow Sea coastal waters now and in the future.

Growth dynamics and yield of melon as influenced by nitrogen fertilizer

Nitrogen (N) is an important nutrient for melon (Cucumis melo L.) production. However there is scanty information about the amount necessary to maintain an appropriate balance between growth and yield. Melon vegetative organs must develop sufficiently to intercept light and accumulate water and nutrients but it is also important to obtain a large reproductive-vegetative dry weight ratio to maximize the fruit yield. We evaluated the influence of different N amounts on the growth, production of dry matter and fruit yield of a melon ‘Piel de sapo’ type. A three-year field experiment was carried out from May to September. Melons were subjected to an irrigation depth of 100% crop evapotranspiration and to 11 N fertilization rates, ranging 11 to 393 kg ha –1 in the three years. The dry matter production of leaves and stems increased as the N amount increased. The dry matter of the whole plant was affected similarly, while the fruit dry matter decreased as the N amount was increased above 112, 93 and 95 kg ha –1 , in 2005, 2006 and 2007, respectively. The maximum Leaf Area Index (LAI), 3.1, was obtained at 393 kg ha –1 of N. The lowest N supply reduced the fruit yield by 21%, while the highest increased the vegetative growth, LAI and Leaf Area Duration (LAD), but reduced yield by 24% relative to the N93 treatment. Excessive applications of N increase vegetative growth at the expense of reproductive growth. For this melon type, rates about 90-100 kg ha –1 of N are sufficient for adequate plant growth, development and maximum production. To obtain fruit yield close to the maximum, the leaf N concentration at the end of the crop cycle should be higher than 19.5 g kg –1

Reduced three-wave model to study the hard transition to chaotic dynamics in Alfven wave-fronts

The derivative nonlinear Schrödinger (DNLS) equation, describing propagation of circularly polarized Alfven waves of finite amplitude in a cold plasma, is truncated to explore the coherent, weakly nonlinear, cubic coupling of three waves near resonance, one wave being linearly unstable and the other waves damped. In a reduced three-wave model (equal damping of daughter waves, three-dimensional flow for two wave amplitudes and one relative phase), no matter how small the growth rate of the unstable wave there exists a parametric domain with the flow exhibiting chaotic dynamics that is absent for zero growth-rate. This hard transition in phase-space behavior occurs for left-hand (LH) polarized waves, paralelling the known fact that only LH time-harmonic solutions of the DNLS equation are modulationally unstable.

Effect of type of dietary carbohydrates on digestion, intestinal barrier and growth performance in 25-d weaned rabbits

Los objetivos globales de esta tesis han sido estudiar el efecto que los carbohidratos de la dieta ejercen sobre los rendimientos productivos, la barrera intestinal, y la digestión de animales destetados a 25 días de edad. Además se ha estudiado cuál es el mejor periodo para determinar la digestibilidad fecal tras el destete a esta edad. En el primer experimento se estudió el efecto de la fibra neutro detergente soluble (FNDS) sobre la barrera intestinal, digestión, microbiota intestinal y rendimientos productivos de gazapos en gazapos en la fase post-destete. Se diseñaron tres piensos isonutritivos en los que la única fuente de variación fueron los niveles de fibra soluble. A partir de una dieta control (AH) con 103 g/kg de materia seca de FNDS y alfalfa como fuente principal de fibra, se sustituyó la mitad de esta alfalfa por una mezcla de pulpa de remolacha y pulpa de manzana (75:25) en el pienso B-AP y por una mezcla de cascarilla y concentrado de proteína de soja (88:12) en el pienso OH, obteniéndose 131 y 79 g/kg de FNDS sobre materia seca, respectivamente. Los conejos se destetaron a 25 días y fueron alimentados con los piensos experimentales hasta los 35 días de edad, momento en el que se sacrificaron para la determinación de la digestibilidad ileal aparente (DIA) de la materia seca (MS), proteína bruta (PB) y almidón, la morfología de la mucosa, y actividad enzimática en el yeyuno, el tejido linfoide asociado a la mucosa, así como la microbiota intestinal. Para la determinación de la morfología de la mucosa se utilizaron adicionalmente 19 animales lactantes de 35 días de edad. Para el estudio de la tasa de mortalidad, se utilizaron 118 animales más por tratamiento que recibieron los piensos experimentales durante las dos semanas post-destete y posteriormente un pienso comercial hasta los 60 días de edad. Los animales recibieron durante todo el experimento medicación en el agua de bebida (100 ppm de apramicina sulfato y 120 ppm de tilosina tartrato). El nivel de fibra soluble mejoró los parámetros que se utilizaron para la caracterización del estado de la barrera intestinal. Los conejos alimentados con el mayor nivel de FNDS en el pienso presentaron una mayor longitud de los villi (P=0.001), un mayor ratio longitud villi/profundidad de las criptas (8.14; P=0.001), una mayor actividad disacaridásica (8671 μmol de glucosa/g de proteína; P=0.019), así como una mayor digestibilidad ileal (96.8%; P=0.002), observándose una reducción en el flujo ileal de almidón a medida que se incrementó el nivel de fibra soluble en el pienso (1,2 vs 0,5 g/d; P=0.001). Los animales lactantes a 35 días de edad presentaron un ratio longitud de villi/profundidad de las criptas menor que el observado en aquéllos alimentados con el pienso B-AP (6.70), pero superior al de los piensos AH y OH. Niveles inferiores de NDFS tendieron (P=0.074) a incrementar la respuesta inmune de tipo celular (linfocitos CD8+). El pienso también afectó a la producción de IL2 (CD25+; P=0.029; CD5+CD25+; P=0.057), pero sin llegar a establecerse una clara relación con el nivel de fibra soluble. La diversidad de la microbiota intestinal no se vio afectada por el pienso (P ≥ 0.38). Los animales alimentados con las piensos B-AP y AH presentaron una reducción en la frecuencia de detección de Clostridium perfringens tanto en íleon (P=0.062) como en ciego (4.3 vs. 17.6%, P =0.047), comparado con el pienso OH. Además la tasa de mortalidad (118 gazapos/pienso) disminuyó de 14.4% en el pienso OH a 5.1% en el pienso B-AP. Entre los 32 y los 35 días de edad se determinó la digestibilidad fecal aparente (14/pienso) de la materia seca (MS), energía bruta (EB), proteína bruta (PB), fibra neutro detergente (FND), fibra ácido detergente (FAD) y almidón. Este grupo, junto con otros nueve animales por tratamiento se utilizaron para determinar el peso del estómago y el ciego, la concentración cecal de ácidos grasos volátiles (AGV) y amoniaco (NH3), así como las tasas de similitud de la microbiota intestinal. Además se estudiaron los rendimientos productivos (35 animales/tratamiento) de los gazapos durante todo el período de cebo, consumiendo los piensos experimentales desde el destete hasta los 35 días y posteriormente un pienso comercial hasta los 60 días de edad. Niveles crecientes de FNDS mejoraron la digestibilidad fecal de la materia seca (MS) y energía (P<0.001). La inclusión FNDS aumentó de manera lineal el peso del contenido cecal (P=0.001) y el peso del aparato digestivo completo (P=0.008), y en los días previos al sacrificio disminuyó de manera lineal el consumo medio diario (P=0.040). Se observó además, una disminución lineal (P≤0.041) del pH del estómago. No se encontró relación entre el pH, la concentración y proporciones molares de AGV y el nivel de FNDS. El pienso pareció tener un efecto, incluso superior al de la madre, sobre la tasa de similitud de la microbiota, y los efectos fueron mayores a nivel cecal que ileal. La eficacia alimenticia aumentó de manera lineal en un 12% entre piensos extremos tras el destete (25- 39d) y en un 3% en el período global de cebo con niveles mayores de NDFS. El consumo medio diario durante la fase post-destete y durante todo el período de cebo, tendió a aumen tar (P≤0.079) con niveles mayores de FNDS, sin embargo no se apreció efecto sobre la ganancia media diaria (P≥0.15). En conclusión, el incremento del nivel de fibra soluble en el pienso parece resultar beneficioso para la salud del animal ya que mejora la integridad de la mucosa, y reduce la frecuencia de detección de potenciales patógenos como C. perfringens y Campylobacter spp. Conforme a estos resultados, debería tenerse en cuenta el contenido en fibra soluble en la formulación de piensos de conejos en la fase post-destete. El objetivo del segundo experimento fue determinar el efecto de la fuente de almidón sobre la digestión, la microbiota intestinal y los rendimientos productivos en conejos destetados con 25 días de edad. Se formularon tres piensos isonutritivos en los que se modificaron las principales fuentes de almidón: trigo crudo, trigo cocido y una combinación de trigo y arroz cocido. Dos grupos de 99 y 193 animales se destetaron con 25 días de edad. El primero de ellos se utilizó para la determinación de los parámetros productivos conforme al mismo protocolo seguido en el experimento anterior. El segundo de los grupos se utilizó para la determinación de la digestibilidad fecal de 32 a 35 d, la digestibilidad ileal aparente (DIA) a 35 d, la morfología de la mucosa intestinal, los parámetros de fermentación cecal; así como, la caracterización de la microbiota intestinal. Se utilizaron además dos grupos adicionales de animales 384 (medicados) y 177 (no medicados) para estudiar el efecto de la suplementación con antibióticos en el agua de bebida sobre la mortalidad. El procesado térmico del trigo mejoró ligeramente la digestibilidad ileal del almidón (P=0.020) pero no modificó el flujo final de almidón que alcanzó el ciego, observándose una mayor frecuencia de detección de Campylobacter spp. y Ruminococcus spp. en ciego (P≤0.023), pero sin cambios a nivel ileal. El procesado térmico del trigo no afectó tampoco a los parámetros productivos, la mortalidad, la digestibilidad ileal y fecal o la morfología de la mucosa. La sustitución parcial del trigo cocido por arroz cocido, penalizó la digestibilidad ileal del almidón (P=0.020) e incrementó el flujo ileal de este nutriente al ciego (P=0.007). Sin embargo no afectó a la mortalidad, pese a que se detectaron cambios en la microbiota tanto a nivel ileal como cecal, disminuyendo la frecuencia de detección de Campylobacter spp. (en íleon y ciego), Helicobacter spp. (en íleon) y Ruminococcus spp (en ciego) e incrementando Bacteroides spp. (en ciego) (P≤0.046). El empleo de arroz cocido en las piensos post-destete no tuvieron efectos sobre los parámetros productivos, la mortalidad, la digestibilidad ileal y fecal a excepción del almidón, o la morfología de la mucosa. La suplementación con antibiótico redujo la fre cuencia de detección de la mayoría de las bacterias estudiadas (P≤0.048), sobre todo para Campylobacter spp., Clostridium perfringens y Propionibacterium spp. (P≤0.048), observándose un efecto mayor a nivel ileal que cecal, lo que se asoció a la bajada significativa (P<0.001) de la mortalidad. En conclusión, los resultados de este experimento indican que la fuente de almidón afecta a la microbiota intestinal pero no influiye sobre la salud del animal. En relación al procesado, el uso de trigo cocido junto con arroz cocido no mejora los resultados obtenidos con trigo duro, si bienserían necesarios más experimentos que confirmaran este punto. El último de los experimentos se centró en un aspecto metodológico. Dado que, los conejos destetados presentan un patrón digestivo diferente al de un animal adulto resultado de su inmadurez digestiva, el objetivo buscado era tratar de determinar el mejor procedimiento para la determinación de la digestibilidad fecal en los gazapos en la fase post-destete. Para tal fin se utilizaron 15 animales/tratamiento de tres camadas diferentes que se destetaron con 25 días, suministrándoles un pienso comercial de crecimiento-cebo. Se registró el consumo medio diario y la excreción diaria de heces desde el día 25 hasta el día 40 de edad para la determinación de la digestibilidad de la MS. La camada afectó al consumo medio diario y la excreción de heces (P=0.013 y 0.014, respectivamente), observándose una tendencia (P=0.061) en la digestibilidad. La edad afectó (P<0.001) a todos estos factores, incrementándose de manera más evidente la excreción que la ingestión de materia seca en la primera semana de vida, para aumentar de forma paralela a partir de la segunda. La correlación entre el consumo medio diario fue mayor con la excreción de heces del mismo día que con la del día siguiente, por lo que se utilizó el primero para la determinación de la digestibilidad de la MS (MSd). La MSd disminuyó de manera lineal hasta los 32 días de edad (2.17±0.25 unidades porcentuales por día), mientras que permaneció constante desde los 32 a los 40 días (69.4±0.47%). Por otro lado, la desviación estándar de la MSd se redujo cuando se incrementó el período de recogida de 2 a 6 días en un 54%. Conforme a los resultados obtenidos, se puede concluir que no es aconsejable comenzar las pruebas de digestibilidad antes de los 32 días de edad y que el número de animales necesario para detectar diferencias significativas entre tratamientos dependerá del período de recogida de heces. ABSTRACT The global aim of this thesis has been to study the effect of dietary carbohydrates on growth, performance, digestion and intestinal barrier in 25-d weaned rabbits. In addition there has also been studied which is the best period to determine the fecal digestibility after weaning. The first experiment focused on the effect of Neutral Detergent Soluble Fibre (NDSF) on gut barrier function, digestion, intestinal microbiota and growth performance n rabbits in the post-weaning period. Three isonutritive diets which only varied in the levels of soluble fiber were formulated such as it described as follows: a control diet (AH) containing 103 g of neutral detergent soluble fiber, including alfalfa as main source of fiber, was replaced by a mixture of beet and apple pulp (75-25) in the B-AP diet and, by a mix of oat hulls and soybean protein concentrate (88:12) in the OH diet, resulting 131 and 79 g of NDFS/kg of dry matter, respectively. Rabbits, weaned at 25 days of age, were fed the experimental diets up to 35 days of age, moment in which they were slaughtered for apparent ileal digestibility (AID) of dry matter (DM), crude protein (CP) and starch, mucosa morphology, sucrose activity, characterization of lamina propria lymphocytes and intestinal microbiota. To assess mucosal morphology, 19 suckling 35-d-old rabbits were also used. For mortality study, besides these animals, 118 additional rabbits per treatment were fed the experimental diets for two weeks period and thereafter received a commercial diet until 60 days of age. Rabbits were water medicated during the whole experimental period (100 ppm de apramicine sulphate and 120 ppm of tylosine tartrate). Level of soluble fiber improved all the parameters used for the characterization of the intestinal barrier condition. Villous height of the jejunal mucosa increased with dietary soluble fiber (P=0.001). Villous height of jejunal mucosa increased with dietary soluble fiber (P = 0.001). Rabbits fed the highest level of soluble fiber (BA-P diet) showed the highest villous height/crypth depth ratio (8.14; P = 0.001), sucrase specific activity (8671 μmol glucose/ g protein; P = 0.019), and the greatest ileal starch digestibility (96.8%; P = 0.002). The opposite effects were observed in rabbits fed decreased levels of soluble fiber (AH and OH diets; 4.70, 5,848 μmol of glucose/g of protein, as average, respectively). The lowest ileal starch digestibility was detected for animal fed OH diet (93.2%). Suckling rabbits of the same age showed a lower villous height/crypt depth ratio (6.70) compared with the B-AP diet group, but this ration was higher that the AH or OH diet groups. Lower levels of soluble fiber tended (P = 0.074) to increase the cellular immune response (CD8+ lymphocytes). Diet affected IL-2 production (CD25+, P = 0.029; CD5+CD25+, P = 0.057), with no clear relationship between soluble fiber and IL-2. The intestinal microbiota biodiversity was not affected by diets (P ≥ 0.38). Animals fed B-AP and AH diets had a reduced cecal frequency of detection compatible with Campylobacter spp. (20.3 vs. 37.8, P = 0.074), and Clostridium perfringens (4.3 vs. 17.6%, P = 0.047), compared with the OH diet group. Moreover, the mortality rates decreased from 14.4 (OH diet) to 5.1% (B-AP diet) with the increased presence of soluble fiber in the diet. Between 32 and 35 days of age, faecal apparent digestibility of dry matter (DM), gross energy (GE), crude protein (CP), neutral detergent fiber (NDF), acid detergent fiber (ADF) and starch was determined (14/diet). This group, plus another nine rabbits/diet were used to determine weight of stomach and caecum and their contents, cecal fermentation traits and similarity rate (SR) of intestinal microbiota. Besides, growth performance parameters (35 rabbits/diet) were studied during the whole fattening period, in animals consuming the experimental feed after the weaning up to 35 days of age and later on a commercial diet up animals reached 60 days of age. Increasing levels of Neutral Detergent Soluble Fiber improved faecal dry matter and energy digestibility (P<0.001). NDSF inclusion improved linearly weight of the caecal content (P=0.001) and the total gastrointestinal tract (P=0.008), and in the previous days to slaughter a linear decrease of daily feed intake in diet with highest level of soluble fiber was also observed. Stomach pH decreased linearly with increasing levels of NDFS (P≤0.041). No relation between NDSF level on pH, concentration and molar proportion of VFA was found. Treatments appeared to influence the similarity rate of microbiota, even higher to mother effect. These effects were higher in ileum than in caecum. A linear positive effect of feed efficiency was observed, which increased around 12% in the two weeks post-weaning (25-39d) and 3% in the whole fattening period between extreme diets with highest levels of soluble fiber. Average daily feed intake during the two weeks after weaning and in the whole fattening period, tended (P≤0.079) to increase with highest levels of NDSF; although there were no effect on daily weight gain (≥0.15). In conclusion, an increase of soluble fiber in the feed seems to be beneficial for animal health, due to improve mucose integrity and reduce detection frequency of those poten tial pathogens like C. perfringens and Campylobacter spp. According to these results, level of soluble fiber should be taking care in feed rabbit formulation in the post-weaning period. The objective of the second experiment was to determine the effect of source of starch on digestion, intestinal microbiota and growth performance in twenty-five-day old weaned rabbits. To accomplish with this aim three iso-nutritive diets were formulated with different source of starch: raw wheat, boiled wheat and a combination of boiled wheat and boiled rice. Two groups of 99 and 193 rabbits were weaned at 25 days of age. The first group was used for growth performance determination following the same protocol than in previous experiment. The second group was used to determine faecal digestibility from 32 to 35 d, apparent ileal digestibility (AID) at 35 d, jejunal mucosa morphology, caecal fermentation traits and characterization of intestinal microbiota. For mortality, two additional groups of 384 (medicated) and 177 (not medicated) were used in order to study the effect of antibiotic water supply supplementation. Heat processing of starch slightly improved ileal digestibility of starch (P=0.020) but did not modify the flow of starch to the caecum. An increase in frequency of detection of Campylobacter spp. y Ruminococcus spp. was observed in the caecum (P≤0.023), with no changes at ileal level. Heat processing of wheat did not modify growth performance, mortality, ileal or faecal digestibility and mucosa morphology. Partial substitution of boiled wheat for boiled rice in the diet impaired ileal starch digestibility (P=0.020) and increased the ileal flow of this nutrient to the caecum (P=0.007). However, it did not affect mortality rate, although changes in the ileal and caecal intestinal microbiota were detected, decreasing the frequency of detection of Campylobacter spp. (both ileum and caecum), Helicobacter spp. (at ileum) and Ruminococcus spp (at caecum) and increasing the Bacteroides spp. (at caecum) (P≤0.046). The effect of boiled rice supplementation did not alter growth performance, mortality, ileal or faecal digestibility of other nutrients than starch, and mucosa morphology. Medication of rabbits reduced the ileal frequency of detection of most bacteria studied (P≤0.048), especially for Campylobacter spp., Clostridium perfringens y Propionibacterium spp. (P≤0.048), resulting the effect higher at ileal than caecal level and relating it with a strong reduction of mortality rate (P<0.001). In conclusion, the results of this experiment make think that the source of starch affects the intestinal microbiota but they do not seem to influence animal health. In relation to the effect of heat processed the use of cooked wheat or cooked rice it does not seem to im prove the results obtained with hard wheat, but there would be necessary more experiments that were confirming this point. The last experiment focused on a methodological aspect. Considering that, weaned rabbits have a different digestive pattern than older animals due to their digestive immaturity; the fixed objective was to determine the best procedure for faecal digestibility determination in young rabbits in the post-weaning period. Fifteen rabbits from 5 different litters were weaned at 25 days of age and fed with a commercial feed. Feed intake and faeces excretion were recorded daily from 25 to 40 days of age for dry matter digestibility (DMd) determination. Litter affected daily DM intake and excretion (P=0.013 y 0.014, respectively) and tended to affect DMd (P=0.061). Age affected all these factors (P<0.001), but ingestion increased slowly than dry matter excretion during the first week buth they evolved similarly in the second week. The correlation between daily feed intakes was higher with the faeces excretion of the day than with faeces excretion of the next day, and the first values were used to determine daily DMd. The DMd decreased linearly from weaning to 32 d of age (2.17±0.25 percentage units per day), whereas from 32 to 40 d remained constant (69.4±0.47%). On the other hand, average standard deviation of DMd decreased by 54% when the length of collection period increased from 2 to 6d. Consequently to the obtained results, it could be concluded that it would not be advisable to start digestibility trials before the 32 days of age and that the number of animals required to detect a significant difference among means would depend on the collection period.

Evaluation of cover crops based on characteristics of nitrogen use, and aerial and root growth

La caracterización de los cultivos cubierta (cover crops) puede permitir comparar la idoneidad de diferentes especies para proporcionar servicios ecológicos como el control de la erosión, el reciclado de nutrientes o la producción de forrajes. En este trabajo se estudiaron bajo condiciones de campo diferentes técnicas para caracterizar el dosel vegetal con objeto de establecer una metodología para medir y comparar las arquitecturas de los cultivos cubierta más comunes. Se estableció un ensayo de campo en Madrid (España central) para determinar la relación entre el índice de área foliar (LAI) y la cobertura del suelo (GC) para un cultivo de gramínea, uno de leguminosa y uno de crucífera. Para ello se sembraron doce parcelas con cebada (Hordeum vulgare L.), veza (Vicia sativa L.), y colza (Brassica napus L.). En 10 fechas de muestreo se midieron el LAI (con estimaciones directas y del LAI-2000), la fracción interceptada de la radiación fotosintéticamente activa (FIPAR) y la GC. Un experimento de campo de dos años (Octubre-Abril) se estableció en la misma localización para evaluar diferentes especies (Hordeum vulgare L., Secale cereale L., x Triticosecale Whim, Sinapis alba L., Vicia sativa L.) y cultivares (20) en relación con su idoneidad para ser usadas como cultivos cubierta. La GC se monitorizó mediante análisis de imágenes digitales con 21 y 22 muestreos, y la biomasa se midió 8 y 10 veces, respectivamente para cada año. Un modelo de Gompertz caracterizó la cobertura del suelo hasta el decaimiento observado tras las heladas, mientras que la biomasa se ajustó a ecuaciones de Gompertz, logísticas y lineales-exponenciales. Al final del experimento se determinaron el C, el N y el contenido en fibra (neutrodetergente, ácidodetergente y lignina), así como el N fijado por las leguminosas. Se aplicó el análisis de decisión multicriterio (MCDA) con objeto de obtener un ranking de especies y cultivares de acuerdo con su idoneidad para actuar como cultivos cubierta en cuatro modalidades diferentes: cultivo de cobertura, cultivo captura, abono verde y forraje. Las asociaciones de cultivos leguminosas con no leguminosas pueden afectar al crecimiento radicular y a la absorción de N de ambos componentes de la mezcla. El conocimiento de cómo los sistemas radiculares específicos afectan al crecimiento individual de las especies es útil para entender las interacciones en las asociaciones, así como para planificar estrategias de cultivos cubierta. En un tercer ensayo se combinaron estudios en rhizotrones con extracción de raíces e identificación de especies por microscopía, así como con estudios de crecimiento, absorción de N y 15N en capas profundas del suelo. Las interacciones entre raíces en su crecimiento y en el aprovisionamiento de N se estudiaron para dos de los cultivares mejor valorados en el estudio previo: uno de cebada (Hordeum vulgare L. cv. Hispanic) y otro de veza (Vicia sativa L. cv. Aitana). Se añadió N en dosis de 0 (N0), 50 (N1) y 150 (N2) kg N ha-1. Como resultados del primer estudio, se ajustaron correctamente modelos lineales y cuadráticos a la relación entre la GC y el LAI para todos los cultivos, pero en la gramínea alcanzaron una meseta para un LAI>4. Antes de alcanzar la cobertura total, la pendiente de la relación lineal entre ambas variables se situó en un rango entre 0.025 y 0.030. Las lecturas del LAI-2000 estuvieron correlacionadas linealmente con el LAI, aunque con tendencia a la sobreestimación. Las correcciones basadas en el efecto de aglutinación redujeron el error cuadrático medio del LAI estimado por el LAI-2000 desde 1.2 hasta 0.5 para la crucífera y la leguminosa, no siendo efectivas para la cebada. Esto determinó que para los siguientes estudios se midieran únicamente la GC y la biomasa. En el segundo experimento, las gramíneas alcanzaron la mayor cobertura del suelo (83-99%) y la mayor biomasa (1226-1928 g m-2) al final del mismo. Con la mayor relación C/N (27-39) y contenido en fibra digestible (53-60%) y la menor calidad de residuo (~68%). La mostaza presentó elevadas GC, biomasa y absorción de N en el año más templado en similitud con las gramíneas, aunque escasa calidad como forraje en ambos años. La veza presentó la menor absorción de N (2.4-0.7 g N m-2) debido a la fijación de N (9.8-1.6 g N m-2) y escasa acumulación de N. El tiempo térmico hasta alcanzar el 30% de GC constituyó un buen indicador de especies de rápida cubrición. La cuantificación de las variables permitió hallar variabilidad entre las especies y proporcionó información para posteriores decisiones sobre la selección y manejo de los cultivos cubierta. La agregación de dichas variables a través de funciones de utilidad permitió confeccionar rankings de especies y cultivares para cada uso. Las gramíneas fueron las más indicadas para los usos de cultivo de cobertura, cultivo captura y forraje, mientras que las vezas fueron las mejor como abono verde. La mostaza alcanzó altos valores como cultivo de cobertura y captura en el primer año, pero el segundo decayó debido a su pobre actuación en los inviernos fríos. Hispanic fue el mejor cultivar de cebada como cultivo de cobertura y captura, mientras que Albacete como forraje. El triticale Titania alcanzó la posición más alta como cultiva de cobertura, captura y forraje. Las vezas Aitana y BGE014897 mostraron buenas aptitudes como abono verde y cultivo captura. El MCDA permitió la comparación entre especies y cultivares proporcionando información relevante para la selección y manejo de cultivos cubierta. En el estudio en rhizotrones tanto la mezcla de especies como la cebada alcanzaron mayor intensidad de raíces (RI) y profundidad (RD) que la veza, con valores alrededor de 150 cruces m-1 y 1.4 m respectivamente, comparados con 50 cruces m-1 y 0.9 m para la veza. En las capas más profundas del suelo, la asociación de cultivos mostró valores de RI ligeramente mayores que la cebada en monocultivo. La cebada y la asociación obtuvieron mayores valores de densidad de raíces (RLD) (200-600 m m-3) que la veza (25-130) entre 0.8 y 1.2 m de profundidad. Los niveles de N no mostraron efectos claros en RI, RD ó RLD, sin embargo, el incremento de N favoreció la proliferación de raíces de veza en la asociación en capas profundas del suelo, con un ratio cebada/veza situado entre 25 a N0 y 5 a N2. La absorción de N de la cebada se incrementó en la asociación a expensas de la veza (de ~100 a 200 mg planta-1). Las raíces de cebada en la asociación absorbieron también más nitrógeno marcado de las capas profundas del suelo (0.6 mg 15N planta-1) que en el monocultivo (0.3 mg 15N planta-1). ABSTRACT Cover crop characterization may allow comparing the suitability of different species to provide ecological services such as erosion control, nutrient recycling or fodder production. Different techniques to characterize plant canopy were studied under field conditions in order to establish a methodology for measuring and comparing cover crops canopies. A field trial was established in Madrid (central Spain) to determine the relationship between leaf area index (LAI) and ground cover (GC) in a grass, a legume and a crucifer crop. Twelve plots were sown with either barley (Hordeum vulgare L.), vetch (Vicia sativa L.), or rape (Brassica napus L.). On 10 sampling dates the LAI (both direct and LAI-2000 estimations), fraction intercepted of photosynthetically active radiation (FIPAR) and GC were measured. A two-year field experiment (October-April) was established in the same location to evaluate different species (Hordeum vulgare L., Secale cereale L., x Triticosecale Whim, Sinapis alba L., Vicia sativa L.) and cultivars (20) according to their suitability to be used as cover crops. GC was monitored through digital image analysis with 21 and 22 samples, and biomass measured 8 and 10 times, respectively for each season. A Gompertz model characterized ground cover until the decay observed after frosts, while biomass was fitted to Gompertz, logistic and linear-exponential equations. At the end of the experiment C, N, and fiber (neutral detergent, acid and lignin) contents, and the N fixed by the legumes were determined. Multicriteria decision analysis (MCDA) was applied in order to rank the species and cultivars according to their suitability to perform as cover crops in four different modalities: cover crop, catch crop, green manure and fodder. Intercropping legumes and non-legumes may affect the root growth and N uptake of both components in the mixture. The knowledge of how specific root systems affect the growth of the individual species is useful for understanding the interactions in intercrops as well as for planning cover cropping strategies. In a third trial rhizotron studies were combined with root extraction and species identification by microscopy and with studies of growth, N uptake and 15N uptake from deeper soil layers. The root interactions of root growth and N foraging were studied for two of the best ranked cultivars in the previous study: a barley (Hordeum vulgare L. cv. Hispanic) and a vetch (Vicia sativa L. cv. Aitana). N was added at 0 (N0), 50 (N1) and 150 (N2) kg N ha-1. As a result, linear and quadratic models fitted to the relationship between the GC and LAI for all of the crops, but they reached a plateau in the grass when the LAI > 4. Before reaching full cover, the slope of the linear relationship between both variables was within the range of 0.025 to 0.030. The LAI-2000 readings were linearly correlated with the LAI but they tended to overestimation. Corrections based on the clumping effect reduced the root mean square error of the estimated LAI from the LAI-2000 readings from 1.2 to less than 0.50 for the crucifer and the legume, but were not effective for barley. This determined that in the following studies only the GC and biomass were measured. In the second experiment, the grasses reached the highest ground cover (83- 99%) and biomass (1226-1928 g/m2) at the end of the experiment. The grasses had the highest C/N ratio (27-39) and dietary fiber (53-60%) and the lowest residue quality (~68%). The mustard presented high GC, biomass and N uptake in the warmer year with similarity to grasses, but low fodder capability in both years. The vetch presented the lowest N uptake (2.4-0.7 g N/m2) due to N fixation (9.8-1.6 g N/m2) and low biomass accumulation. The thermal time until reaching 30% ground cover was a good indicator of early coverage species. Variable quantification allowed finding variability among the species and provided information for further decisions involving cover crops selection and management. Aggregation of these variables through utility functions allowed ranking species and cultivars for each usage. Grasses were the most suitable for the cover crop, catch crop and fodder uses, while the vetches were the best as green manures. The mustard attained high ranks as cover and catch crop the first season, but the second decayed due to low performance in cold winters. Hispanic was the most suitable barley cultivar as cover and catch crop, and Albacete as fodder. The triticale Titania attained the highest rank as cover and catch crop and fodder. Vetches Aitana and BGE014897 showed good aptitudes as green manures and catch crops. MCDA allowed comparison among species and cultivars and might provide relevant information for cover crops selection and management. In the rhizotron study the intercrop and the barley attained slightly higher root intensity (RI) and root depth (RD) than the vetch, with values around 150 crosses m-1 and 1.4 m respectively, compared to 50 crosses m-1 and 0.9 m for the vetch. At deep soil layers, intercropping showed slightly larger RI values compared to the sole cropped barley. The barley and the intercropping had larger root length density (RLD) values (200-600 m m-3) than the vetch (25-130) at 0.8-1.2 m depth. The topsoil N supply did not show a clear effect on the RI, RD or RLD; however increasing topsoil N favored the proliferation of vetch roots in the intercropping at deep soil layers, with the barley/vetch root ratio ranging from 25 at N0 to 5 at N2. The N uptake of the barley was enhanced in the intercropping at the expense of the vetch (from ~100 mg plant-1 to 200). The intercropped barley roots took up more labeled nitrogen (0.6 mg 15N plant-1) than the sole-cropped barley roots (0.3 mg 15N plant-1) from deep layers.

Advances on MBE selective area growth of III-nitride nanostructures: from nanoLEDs to pseudo substrates

The aim of this work is to provide an overview on the recent advances in the selective area growth (SAG) of (In)GaN nanostructures by plasma assisted molecular beam epitaxy, focusing on their potential as building blocks for next generation LEDs. The first three sections deal with the basic growth mechanisms of GaN SAG and the emission control in the entire ultraviolet to infrared range, including approaches for white light emission, using InGaN disks and thick segments on axial nanocolumns. SAG of axial nanostructures is eveloped on both GaN/sapphire templates and GaN-buffered Si(111). As an alternative to axial nanocolumns, section 4 reports on the growth and characterization of InGaN/GaN core-shell structures on an ordered array of top-down patterned GaN microrods. Finally, section 5 reports on the SAG of GaN, with and without InGaN insertion, on semi-polar (11-22) and non-polar (11-20) templates. Upon SAG the high defect density present in the templates is strongly reduced as indicated by a dramatic improvement of the optical properties. In the case of SAG on nonpolar (11-22) templates, the formation of nanostructures with a low aspect ratio took place allowing for the fabrication of high-quality, non-polar GaN pseudo-templates by coalescence of these nanostructures.

Influence of source and micronization of soya bean meal on growth performance, nutrient digestibility and ileal mucosal morphology of Iberian piglets

The effects of inclusion in the diet of different sources of soya bean meal (SBM) on growth performance, total tract apparent digestibility (TTAD) and apparent ileal digestibility (AID) of major dietary components and mucosal ileum morphology were studied in Iberian pigs weaned at 30 days of age. From 30 to 51 days of age (phase I), there was a control diet based on regular soya bean meal (R-SBM; 44% CP) of Argentina (ARG) origin and five extra diets in which a high-protein soya bean meal (HP-SBM; 49% CP) of the USA or ARG origin, either ground (990 μm) or micronized (60 μm), or a soya protein concentrate (SPC; 65% CP) substituted the R-SBM. From 51 to 61 days of age (phase II), all pigs were fed a common commercial diet in mash form. The following pre-planned orthogonal contrasts were conducted: (1) R-SBM v. all the other diets, (2) SPC v. all the HP-SBM diets, (3) micronized HP-SBM v. ground HP-SBM, (4) HP-SBM of ARG origin v. HP-SBM of US origin and (5) interaction between source and the degree of grinding of the HP-SBM. Dietary treatment did not affect growth performance of the pigs at any age but from 30 to 51 days of age, post weaning diarrhoea (PWD) was higher (P<0.001) and the TTAD and AID of all nutrients were lower for pigs fed the R-SBM diet than for pigs fed the HP-SBM or the SPC diets. However, no differences between the HP-SBM and the SPC containing diets were detected for any trait. The TTAD of organic matter (P=0.07) and gross energy (GE) (P=0.05) tended to be higher for the micronized HP-SBM than for the ground HP-SBM and that of GE was higher (P<0.05) for US meal than for the ARG meal. Pigs fed R-SBM had lower villus height (P<0.01) than pigs fed HP-SBM or SPC but no differences in ileal mucosal morphology were detected between SPC and HP-SBM containing diets. It is concluded that feeding the HP-SBM or SPC-reduced PWD and improved nutrient digestibility and ileal morphology as compared with feeding the R-SBM, but had no effect on pig performance. The inclusion in the diet of added value soya products (micronized SBM or SPC) in substitution of the R-SBM increased the TTAD of all nutrients and reduced PWD but had no advantage in terms of growth performance over the use of ground HP-SBM.

Inclusion of fiber in diets for brown-egg laying pullets: Effects on growth performance and digestive tract traits from hatching to 17 weeks of age

We investigated the effects of fiber inclusion in the diet on growth performance and digestive traits in pullets from hatching to 17 wk of age. The control diets of the 3 feeding periods (0 to 5 wk, 5 to 10 wk, and 10 to 17 wk) were based on corn and soybean meal and did not include any additional fiber source. The experimental diets included 2 or 4% of cereal straw or sugar beet pulp (SBP) at the expense (wt:wt) of the control diet. From 0 to 5 wk of age, fiber inclusion did not affect pullet performance. From hatch to 17 wk of age, the inclusion of straw had little effect on pullet performance but the inclusion of 4% SBP reduced (ADG) (P < 0.05) and reduced feed conversion ratio (FCR; P < 0.001). Pullets fed straw had greater ADG (P < 0.05) and better energy conversion ratio (P < 0.01) than pullets fed SBP. An increase in fiber from 2 to 4% reduced FCR (P < 0.05). Body weight uniformity was not affected by diet. Fiber inclusion increased the relative weight (% BW) of the gizzard at 5 wk (P = 0.056) and 10 wk (P < 0.01) of age, but no differences were detected between fiber sources. At same ages, the relative length (cm/kg BW) of the pullets (P = 0.058 and P < 0.01, respectively) and tarsus (P = 0.079 and P < 0.05, respectively) was higher in pullets fed SBP than in pullets fed straw. Fiber inclusion, however, did not affect any of these traits at 17 wk of age. In summary, the inclusion of 2% straw at the expense (wt:wt) of the whole diet did not affect pullet performance at 17 wk of age. An increase in the level of straw from 2 to 4% reduced FCR but did not affect ADG. The inclusion of SBP, however, reduced pullet growth, with effects being more pronounced at the higher level.

Feed form and energy concentration of the diet affect growth performance and digestive tract traits of brown-egg laying pullets from hatching to 17 weeks of age

The influence of feed form and energy concentration of the diet on growth performance and the development of the gastrointestinal tract (GIT) was studied in brown-egg laying pullets. Diets formed a 2 x 5 factorial with 2 feed forms (mash vs. crumbles) and 5 levels of energy differing in 50 kcal AMEn/kg. For the entire study (0 to 17 wk of age) feeding crumbles increased ADFI (52.9 vs. 49.7 g; P < 0.001) and ADG (12.7 vs. 11.6 g; P < 0.001) and improved feed conversion ratio (FCR; 4.18 vs. 4.27; P < 0.001). An increase in the energy content of the diet decreased ADFI linearly (P < 0.001) and improved FCR quadratically (P < 0.01) but energy intake (kcal AMEn/d) was not affected. BW uniformity was higher (P < 0.05) in pullets fed crumbles than in those fed mash but was not affected (P > 0.05) by energy content of the diet. At 5, 10, and 17 wk of age, the relative weight (RW, % BW) of the GIT and the gizzard, and gizzard digesta content were lower (P < 0.05 to P < 0.001) and gizzard pH was higher (P < 0.05 to P < 0.001) in pullets fed crumbles than in pullets fed mash. Energy concentration of the diet did not affect any of the GIT variables studied. In summary, feeding crumbles improved pullet performance and reduced the RW of the GIT and gizzard, and increased gizzard pH at all ages. An increase in the energy content of the diet improved FCR from 0 to 17 wk of age. The use of crumbles and the increase in the AMEn content of the diet might be used adventageously when the objetive is to increase the BW of the pullets. However, crumbles affected the development and weight of the organs of the GIT, which might have negative effects on feed intake and egg production at the beginning of the egg laying cycle.

A yeast sterol auxotroph (erg25) is rescued by addition of azole antifungals and reduced levels of heme

Genetic disruption of the Saccharomyces cerevisiae C-4 sterol methyl oxidase ERG25 gene leads to sterol auxotrophy. We have characterized a suppression system that requires two mutations to restore viability to this disrupted strain. One suppressor mutation is erg11, which is blocked in 14α-demethylation of lanosterol and is itself an auxotroph. The second suppressor mutation required is either slu1 or slu2 (suppressor of lanosterol utilization). These mutations are leaky versions of HEM2 and HEM4, respectively; addition of exogenous hemin reverses the suppressing effects of slu1 and slu2. Suppression of erg25 by erg11 slu1 (or erg11 slu2) results in a slow-growing strain in which lanosterol, the first sterol in the pathway, accumulates. This result indicates that endogenously synthesized lanosterol can substitute for ergosterol and support growth. In the triple mutants, all but 1 (ERG6) of the 13 subsequent reactions of the ergosterol pathway are inactive. Azole antibiotics (clotrimazole, ketoconazole, and itraconazole) widely used to combat fungal infections are known to do so by inhibiting the ERG11 gene product, the 14α-demethylase. In this investigation, we demonstrate that treatment of the sterol auxotrophs erg25 slu1 or erg25 slu2 with azole antibiotics paradoxically restores viability to these strains in the absence of sterol supplementation via the suppression system we have described.

Insulin and insulin-like growth factor-I acutely inhibit surface translocation of growth hormone receptors in osteoblasts: A novel mechanism of growth hormone receptor regulation

We previously have demonstrated that insulin and insulin-like growth factor-I (IGF-I) down-regulate growth hormone (GH) binding in osteoblasts by reducing the number of surface GH receptors (GHRs). The present study was undertaken to investigate the mechanism of GHR down-regulation. Treatment with 5 nM insulin or IGF-I for 18 hr significantly decreased surface GH binding to 26.4 ± 2.9% and 23.0 ± 2.7% of control (mean ± SE; P < 0.05), respectively. No corresponding reductions in the mRNA level and total cellular content of GHR were found, nor was the rate of receptor internalization affected. The effects on GHR translocation were assessed by measuring the reappearance of GH binding of whole cells after trypsinization to remove the surface receptors. GH binding of control cultures significantly increased (P < 0.05) over 2 hr after trypsinization, whereas no recovery of binding activity was detected in insulin and IGF-I-treated cultures, indicating that GHR translocation was impaired. Studies on the time course of GHR down-regulation revealed that surface GH binding was reduced significantly by 3-hr treatment (P ≤ 0.0005), whereas GHR translocation was completely abolished by 75–90 min with insulin and IGF-I. The inhibition of receptor translocation by insulin, but not IGF-I, was attenuated by wortmannin. In conclusion, insulin and IGF-I down-regulated GH binding in osteoblasts by acutely impairing GHR translocation, with their effects exerted through distinct postreceptor signaling pathways.

A single in vivo administration of bombesin antagonist RC-3095 reduces the levels and mRNA expression of epidermal growth factor receptors in MXT mouse mammary cancers

Epidermal growth factor (EGF) and its receptors (EGFR) play important roles in tumorigenesis. In various experimental cancers, treatment with antagonists of bombesin/gastrin-releasing peptide (BN/GRP) produces a reduction in EGFRs, concomitant to inhibition of tumor growth. To investigate the mechanisms involved, we monitored concentrations of BN/GRP antagonist RC-3095 in serum of mice, rats, and hamsters given a single subcutaneous or intravenous injection of this analog. In parallel studies, we measured levels and mRNA expression of EGFRs in estrogen-dependent and independent MXT mouse mammary cancers, following a single subcutaneous administration of RC-3095 to tumor-bearing mice. Peak values of RC-3095 in serum were detected 2 min after intravenous or 15 min after subcutaneous injection. The levels of RC-3095 declined rapidly and became undetectable after 3–5 hr. In the estrogen-dependent MXT tumors, the concentration of EGF receptors was reduced by about 60% 6 hr following injection and returned to original level after 24 hr. Levels of mRNA for EGFR fell parallel with the receptor number and were nearly normal after 24 hr. In the hormone-independent MXT cancers, the number of EGFRs decreased progressively, becoming undetectable 6 hr after injection of RC-3095, and returned to normal values at 24 hr, but EGFR mRNA levels remained lower for 48 hr. Thus, in spite of rapid elimination from serum, BN/GRP antagonist RC-3095 can induce a prolonged decrease in levels and mRNA expression of EGFRs. These findings may explain how single daily injections of BN/GRP antagonists can maintain tumor growth inhibition.

Growth and viability of macrophages continuously stimulated to produce nitric oxide

Deregulated production of nitric oxide (NO) has been implicated in the development of certain human diseases, including cancer. We sought to assess the damaging potential of NO produced under long-term conditions through the development of a suitable model cell culture system. In this study, we report that when murine macrophage-like RAW264.7 cells were exposed continuously to bacterial lipopolysaccharide (LPS) or mouse recombinant interferon-γ (IFN-γ) over periods of 21–23 days, they continued to grow, but with doubling times 2 to 4 times, respectively, longer than the doubling time of unstimulated cells. Stimulated cells produced NO at rates of 30 to 70 nmol per million cells per day throughout the stimulation period. Within 24 hr after removal of stimulant, cells resumed exponential growth. Simultaneous exposure to LPS and IFN-γ resulted in decreased cell number, which persisted for 2 days after removal of the stimulants. Exponential growth was attained only after an additional 4 days. Addition of N-methyl-l-arginine (NMA), an NO synthase inhibitor, to the medium inhibited NO production by 90% of all stimulated cells, partially reduced doubling time of cells stimulated with LPS or IFN-γ, and partially increased viability and growth rates in those exposed to both LPS and IFN-γ. However, when incubated with LPS and IFN-γ at low densities both in the presence and in the absence of NMA, cells grew at a rate slower than that of unstimulated cells, with no cell death, and they resumed exponential growth 24 hr after removal of stimulants. Results from cell density experiments suggest that macrophages are protected from intracellularly generated NO; much of the NO damaging activity occurred outside of the producer cells. Collectively, results presented in this study suggest that the type of cellular toxicity observed in macrophages is markedly influenced by rate of exposure to NO: at low rates of exposure, cells exhibit slower growth; at higher rates, cells begin to die; at even higher rates, cells undergo growth arrest or die. The ability of RAW264.7 cells to produce NO over many cell generations makes the cell line a useful system for the study of other aspects of cellular damage, including genotoxicity, resulting from exposure to NO under long-term conditions.

Identification of eukaryotic mRNAs that are translated at reduced cap binding complex eIF4F concentrations using a cDNA microarray

Although most eukaryotic mRNAs need a functional cap binding complex eIF4F for efficient 5′ end- dependent scanning to initiate translation, picornaviral, hepatitis C viral, and a few cellular RNAs have been shown to be translated by internal ribosome entry, a mechanism that can operate in the presence of low levels of functional eIF4F. To identify cellular mRNAs that can be translated when eIF4F is depleted or in low abundance and that, therefore, may contain internal ribosome entry sites, mRNAs that remained associated with polysomes were isolated from human cells after infection with poliovirus and were identified by using a cDNA microarray. Approximately 200 of the 7000 mRNAs analyzed remained associated with polysomes under these conditions. Among the gene products encoded by these polysome-associated mRNAs were immediate-early transcription factors, kinases, and phosphatases of the mitogen-activated protein kinase pathways and several protooncogenes, including c-myc and Pim-1. In addition, the mRNA encoding Cyr61, a secreted factor that can promote angiogenesis and tumor growth, was selectively mobilized into polysomes when eIF4F concentrations were reduced, although its overall abundance changed only slightly. Subsequent tests confirmed the presence of internal ribosome entry sites in the 5′ noncoding regions of both Cyr61 and Pim-1 mRNAs. Overall, this study suggests that diverse mRNAs whose gene products have been implicated in a variety of stress responses, including inflammation, angiogenesis, and the response to serum, can use translational initiation mechanisms that require little or no intact cap binding protein complex eIF4F.

WISP genes are members of the connective tissue growth factor family that are up-regulated in Wnt-1-transformed cells and aberrantly expressed in human colon tumors

Wnt family members are critical to many developmental processes, and components of the Wnt signaling pathway have been linked to tumorigenesis in familial and sporadic colon carcinomas. Here we report the identification of two genes, WISP-1 and WISP-2, that are up-regulated in the mouse mammary epithelial cell line C57MG transformed by Wnt-1, but not by Wnt-4. Together with a third related gene, WISP-3, these proteins define a subfamily of the connective tissue growth factor family. Two distinct systems demonstrated WISP induction to be associated with the expression of Wnt-1. These included (i) C57MG cells infected with a Wnt-1 retroviral vector or expressing Wnt-1 under the control of a tetracyline repressible promoter, and (ii) Wnt-1 transgenic mice. The WISP-1 gene was localized to human chromosome 8q24.1–8q24.3. WISP-1 genomic DNA was amplified in colon cancer cell lines and in human colon tumors and its RNA overexpressed (2- to >30-fold) in 84% of the tumors examined compared with patient-matched normal mucosa. WISP-3 mapped to chromosome 6q22–6q23 and also was overexpressed (4- to >40-fold) in 63% of the colon tumors analyzed. In contrast, WISP-2 mapped to human chromosome 20q12–20q13 and its DNA was amplified, but RNA expression was reduced (2- to >30-fold) in 79% of the tumors. These results suggest that the WISP genes may be downstream of Wnt-1 signaling and that aberrant levels of WISP expression in colon cancer may play a role in colon tumorigenesis.