Los autores agradecen la financiación del Bosque EE.UU. Monitoreo Evaluación de Protección de la Salud del Servicio Forestal y programas especiales de desarrollo tecnológico que han apoyado el desarrollo, modificación y aplicación del protocolo descrito aquí para identificar los árboles que sean resistentes y susceptibles a la cochinilla de haya. También damos las gracias a la Holden Arboretum, Kirtland, Ohio, por su constante colaboración y apoyo de nuestro programa de detección de resistencia haya americana.

1 Planta Material:. Campo árboles maduros, en maceta plántulas o maceta Injertos <ol><li> Para las pruebas de campo, seleccione hayas maduros sanos estadounidenses que no muestran signos de infestación escala o enfermedad para la prueba de resistencia posible. Visiblemente también tendrán que ser identificados para ser utilizado como un control (Figura 1) árboles susceptibles. </li><li> Para la prueba de plántulas en maceta o injertos, recoger y germinar hayucos como se describe en Koch y Carey, 2004 o descendiente del injerto como se describe en Carey et al, 2013. </li><li> Crezca las plantas de semillero o injertos en macetas mezcla suelo enmendado con 47 g de micronutrientes, 477 g fertilizante de liberación lenta 15N-3.9P-9.9K, 700 g de perlita gruesa y 75 g de sulfato de aluminio por 2.8 cu. bolsa de pies. Si es necesario más adelante en la temporada de cultivo, fertilizar las plantas semanalmente con solubles 17N-1.3P-14.1K en 200 ppm de nitrógeno. </li><li> Durante la temporada de crecimiento, mantener las plantas en una casa de sombra. Permita que las plantas en estado latente fuera en el otoño antes de que se muevana una instalación de almacenamiento de temperatura controlada (~ 4 ° C), de noviembre a abril. </li></ol> 2. Colección de Beech Escala Huevos <ol><li> En un stand infestado de BBD, inspeccione los árboles muy infestados (fácilmente identificados por su &quot;blanqueado&quot; la apariencia, la Figura 1A) con una lupa de mano para confirmar la presencia de huevos, normalmente abundantes desde mediados de julio hasta mediados de agosto. </li><li> Utilice una brocha para cepillar suavemente los grumos de cera blanca de cochinillas adultas, huevos y otros restos, en una bolsa sellable colección un galón de plástico (fig. 1B). Reunir a partir de un mínimo de tres árboles diferentes, al menos, 12 m de distancia. </li><li> Si es necesario, guarde los huevos en la bolsa de recolección sellado por hasta dos semanas a 4 ° C. Pegue un pedazo pequeño (2,5 cm cuadrados) de espuma de polietileno húmedo en el interior de la bolsa para evitar que los huevos se sequen. </li><li> Para separar los huevos de escala (0,15 x 0,25 mm) de los adultos (0,60 mm) y debris, la construcción de un tamiz utilizando una pieza corta de 2 &quot;cañería de PVC y un acoplamiento para soportar una pieza cuadrada de 250 micras de malla de nylon (Figura 2). </li><li> Vaciar la mezcla de los adultos, cera, huevos, y los residuos de la bolsa de recogida en el tamiz y utilizar un pequeño pincel para fomentar suavemente los huevos para pasar a través de la malla en una placa de Petri de vidrio a continuación. Platos de plástico Petri deben evitarse porque tienen más electricidad estática, por lo que es difícil de mover los huevos. Los huevos antes y después de tamizado se muestran en la Figura 3. </li><li> Huevos purificadas se pueden almacenar en la placa de Petri a 4 ° C durante al menos una semana. Para evitar que los huevos se seque, pegue un pedazo húmedo de espuma para la tapa y el sello con Parafilm. </li></ol> 3. Huevo de viabilidad Ensayos <ol><li> Para evaluar la viabilidad del huevo, usar una jeringa de 10 cc a aplicar un delgado anillo de vaselina alrededor de la circunferencia de la parte inferior de una placa de Petri de 60 mm de vidrio (Figura 4A </strong&gt;). </li><li> Transferir unos 100 huevos al centro del ring, coloque la tapa en la caja de Petri y selle con Parafilm. Dejar que la cápsula Petri sellada a permanecer en reposo a temperatura ambiente durante 2 semanas, 3 semanas o si los huevos se almacenaron a 4 ° C antes de comenzar el ensayo. </li><li> Ninfas eclosionadas se atascarse en la vaselina y pueden ser contados fácilmente, y los huevos vacíos se distinguen fácilmente de los huevos sin eclosionar por su color y brillo (Figura 4B). Calcular la viabilidad dividiendo el número de ninfas por la suma de los huevos vacíos más los huevos completos restantes. Buena viabilidad debe estar en el rango de los huevos 75% a 90% eclosionado. </li></ol> 4. Resistencia Escala Proyección de grande maduro Hayas en el campo <ol><li> Para una prueba cuantitativa, contar en 500 huevos utilizando un microscopio de disección y el uso de una pequeña espátula espolvorear suavemente a través del centro de un predampened, de celda abierta de 10 x 15 x 1,3 cm rectángulo de espuma de polietileno. Aamortiguar espuma, mojarlo y luego exprima el agua tanto como sea posible. Para una prueba cualitativa, 500 huevos se pueden contar y se coloca en un pequeño frasco de vidrio y una línea de &quot;relleno&quot; dibujado que se puede utilizar para medir los lotes adicionales de aproximadamente 500 huevos. </li><li> Coloque la almohadilla de espuma en el árbol de prueba con el lado del huevo frente contra la corteza. Sostenga la almohadilla en su lugar con una cuerda, cuerda, bramante o plástico recubierto de alambre del hardware. Plástico o material metálico a base deben utilizarse en lugar de los materiales de fibra naturales que son más fácilmente barridos por la fauna silvestre. </li><li> Línea de la parte superior y los lados de un cm pedazo de vapor permeable casa de abrigo impermeable con acetato adhesivo a base de silicona precortadas 23 x 30 y colocarlo sobre la almohadilla de espuma del ensayo. Presione los bordes de la envoltura de la casa en el árbol para crear un sello impermeable. Coloque la guita de nylon o alambre recubierto de plástico de hardware alrededor del árbol y la casa de abrigo, para mantenerlo en su lugar mientras que el adhesivo fragüe (Figura 5). </li><li> Coloque un mínimo de dosalmohadillas de análisis de cada árbol de prueba, preferiblemente en lados opuestos de la fuste. En cada sitio, coloque almohadillas de prueba en al menos 2 árboles susceptibles (con obvia infestación escala natural), como control. Antes de colocar los bloques de ensayo en los árboles de control, retire los insectos o huevos de escala que tienen lugar de forma natural con un cepillo de cerdas firmes. Nota: Las almohadillas de espuma con los huevos también se pueden colocar en los árboles con el propósito de huevos escala de cría, que puede ser especialmente útil en áreas donde el nivel de infestación es baja. </li></ol> 5. Resistencia escala de cribado de plántulas en maceta o injertos <ol><li> Para las pruebas, seleccionar árboles en macetas con un diámetro mínimo de 1 cm (pinza 5 cm por encima de la línea del suelo) que son lo suficientemente alto como para tener por lo menos 2 almohadillas de análisis separadas puestas en ellos. Pode las ramas pequeñas laterales cuando sea necesario para dar cabida a las almohadillas de análisis. </li><li> Utilice un microscopio de disección para contar 150 huevos y derrama sobre un precorte y humedecido 2,5 x 7,6 x 1,3 cm bolsas d de células abiertasalmohadilla de espuma lene. </li><li> Coloque la almohadilla de espuma de plántulas con el lado del huevo contra la corteza, con alambre recubierto de plástico. Envuelva un pequeño cuadrado de envoltura de la casa alrededor de la plántula justo por encima de la espuma y sellarla con silicona a base de acetato (ver Figura 6 A). </li><li> Incluir conocido familias plántulas susceptibles o injertos como controles. </li></ol> 6. Recogida de datos Nota: Aproximadamente 52 a 57 semanas después de la colocación de las almohadillas de espuma y huevos en los árboles de las pruebas, los datos pueden ser recolectados. Es importante que esto no se realiza hasta después de adultos han empezado a poner huevos, para que su capacidad de reproducción se puede determinar. En algunos árboles resistentes, no es raro ver a un pequeño número de adultos a establecer, pero sin la reproducción se produzca. <ol><li> Retire con cuidado la almohadilla de espuma y contar el número de cochinillas adultas establecidos en la corteza usando una lupa o lente de aumento (10X). </li><li> Cuando se retira la almohadilla de espuma, no es uncommon para algunos adultos y la mayoría de las masas de huevos para ser tirados fuera del árbol con la espuma (ver Figura 7). Usando un microscopio de disección, contar los grupos de huevo y adultos que han permanecido unido a la almohadilla de espuma. </li></ol>

<ol>
	<li>Houston, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+technique+to+artificially+infest+beech+bark+with+the+scale,+Cryptococcus+fagisuga+(Lindinger).">A technique to artificially infest beech bark with the scale, Cryptococcus fagisuga (Lindinger).</a> U.S. Forest Service. , (1982).</li><li>Houston, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=American+beech+resistance+to+Cryptococcus+fagisuga.">American beech resistance to Cryptococcus fagisuga.</a> Proceeding, IUFRO beech bark disease working party conference. , 38-42 (1983).</li><li>Ehrlich, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+beech+bark+disease:+a+nectria+disease+of+Fagus,+following+Cryptococcus+fagi.">The beech bark disease: a nectria disease of Fagus, following Cryptococcus fagi.</a> 10, 593-692 (1934).</li><li>Papaik, M. J., Canham, C. D., Latty, E. F., Woods, K. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+an+introduced+pathogen+on+resistance+to+natural+disturbance:+beech+bark+disease+and+withdrow.">Effects of an introduced pathogen on resistance to natural disturbance: beech bark disease and withdrow.</a> 35, 1832-1843 (2005).</li><li>Miller-Weeks, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Current+status+of+beech+bark+disease+in+New+England+and+New+York.">Current status of beech bark disease in New England and New York.</a> Proceeding, IUFRO beech bark disease working party conference. , 21-23 (1983).</li><li>Houston, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Beech+bark+disease:+The+aftermath+forests+are+structured+for+a+new+outbreak.">Beech bark disease: The aftermath forests are structured for a new outbreak.</a> J. For. 73, 660-663 (1975).</li><li>Twery, M. J., Patterson, W. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Variations+in+beech+bark+disease+and+its+effects+on+species+composition+and+structure+of+northern+hardwood+stands+in+central+New.">Variations in beech bark disease and its effects on species composition and structure of northern hardwood stands in central New.</a> 14, 565-574 (1984).</li><li>Jakubus, W. J., McLaughlin, C. R., Jensen, P. G., McNulty, S. A., Evans, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alternate+year+beechnut+production+and+its+influence+on+bear+and+marten+populations.">Alternate year beechnut production and its influence on bear and marten populations.</a> Beech Bark Disease: Proceedings of the Beech Bark Disease Symposium. , 79-87 (2005).</li><li>Koch, J. L., Carey, D. W., Mason, M. E., Nelson, C. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+beech+scale+resistance+in+full-+and+half-sib+families.">Assessment of beech scale resistance in full- and half-sib families.</a> Can. J. For. Res. 40, 265-272 (2010).</li><li>Koch, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Beech+bark+disease:+The+oldest+&#8220;new&#8221;+threat+to+American+beech+in+the+United+States.">Beech bark disease: The oldest &#8220;new&#8221; threat to American beech in the United States.</a> Outlooks Pest Manag. 21, 64-68 (2010).</li><li>Koch, J. L., Mason, M. E., Carey, D. W., Palmieri, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Screening+for+resistance+to+beech+bark+disease:+improvements+and+results+from+seedlings+and+grafted+field+selections.">Screening for resistance to beech bark disease: improvements and results from seedlings and grafted field selections.</a> Proceedings of the 4th International Workshop on Genetics of Host-Parasite Interactions in Forestry. , (2012).</li><li>Leak, W. 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Los autores declaran que no tienen intereses financieros en competencia.

Los pasos críticos necesarios para el éxito de este ensayo incluyen la realización de una prueba de viabilidad sobre los huevos y el uso de los controles susceptibles en cada sitio de prueba, y con materiales de macetas. También es importante la utilización de más de una almohadilla de análisis por árbol o planta de semillero. Hemos encontrado que, tanto en el campo como en las pruebas más controladas en macetas con plantas, una fuente frecuente de error puede ser el fracaso de la almohadilla. Por ejemplo, en e...

Enfermedad de la corteza de la haya (BBD) ha tenido un impacto negativo en haya americana en América del Norte desde la introducción de la cochinilla de haya invasivo, Cryptococcus fagisuga, en la provincia canadiense de Nueva Escocia a finales del decenio de 1890 3. Este complejo de insectos de la enfermedad se inicia cuando el insecto de la escala de haya inserta su estilete de alimentación en la corteza de la creación de pequeñas fisuras que proporcionan entrada para la infección por una de las especies Neonectria de hongo (ditissima Neonectria o Neonectria faginata). A medida que crecen los micelios de hongos, grandes áreas de tejido puede morir, con el tiempo que rodea completamente el árbol. El daño causado por la enfermedad debilita el árbol, por lo que es propenso a romperse con vientos fuertes 4. Los niveles de mortalidad en la primera ola de la enfermedad se han notificado a ser tan alta como 50% 5. Los árboles supervivientes a menudo son severamente deformados como chancros forman la reducción de valor del árbol como un producto de madera.0; Estos árboles tienen una propensión a la raíz-brote que conduce a la formación de &quot;matorrales de haya&quot; que impiden a otras especies más deseables desde el establecimiento, lo que reduce el valor económico y ecológico de la posición 6. Aunque la enfermedad de la corteza del haya, no es probable que conduzca a la extinción de la haya americana, que altera la composición de pie y la salud que lleva a una disminución de alimento y hábitat para la vida silvestre 7,8. En las masas afectadas por BBD en muchos años, se ha informado de árboles que permanecen libres de cualquier síntoma de la enfermedad. Ensayos de inoculación artificial han demostrado que estos árboles son resistentes al insecto escala 2. Sin infestación de escala, hay pocas oportunidades para la infección Neonectria, minimizando el impacto del hongo. La mortalidad a gran escala en madera de haya americana debido a la infección Neonectria en ausencia de infestación escala antes nunca tieneha informado, por lo que la resistencia a los resultados de insectos escala de haya en la resistencia a la BBD. Las investigaciones recientes sobre la gestión de BBD se ha centrado en la identificación, la propagación, la cría y la conservación de los árboles de haya americana con resistencia al insecto escala de haya. Los estudios genéticos han demostrado que la resistencia al insecto escala es heredable y cuidadosa selección y mejora genética de los árboles resistentes puede resultar en una mejora significativa en una sola generación 9. Este hallazgo ha impulsado los esfuerzos de los administradores estatales y nacionales forestales en los Estados Unidos para establecer huertos semilleros regionales de haya americana resistente para proporcionar una fuente de semilla BBD resistentes genéticamente diversas para las plantaciones de restauración 10,11. La investigación también ha indicado que la manipulación genética silvícola de soporte por la remoción de los árboles susceptibles y retención de árboles resistentes puede resultar en soporte 9,12 mejorar. &lt;/ P&gt; Gestión de BBD a través de actividades de mejoramiento de árboles o por medio de la realización de prescripciones silvícolas requiere la capacidad de seleccionar y distinguir entre árboles de hayas escala resistentes y susceptibles. Los métodos que aquí se presentan han sido adaptados de un método introducido por primera vez por Dave Houston para inocular artificialmente plántulas con huevos escala de haya 1. El método se puede utilizar como una herramienta de detección para identificar los loci de rasgos cuantitativos (QTL) asociados con la resistencia o para distinguir entre las plantas de semillero en macetas resistentes y susceptibles o rametos injertados en los estudios genéticos. Alternativamente, puede ser utilizado para el cribado de árboles maduros en el campo para identificar árboles resistentes para el desarrollo huerto de semillas, o la retención en el campo. Árboles susceptibles pueden ser identificados y eliminados para minimizar el impacto de la enfermedad.

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a technique to screen american beech for resistance to the beech scale insect (cryptococcus fagisuga lind.)

Enfermedad de la corteza de la haya (BBD) se traduce en altos niveles de mortalidad inicial, dejando detrás de los árboles sobrevivientes que se debilitan en gran medida y deformadas. La enfermedad se inicia por la alimentación de las actividades de la insecto de la escala de haya invasiva, Cryptococcus fagisuga, que crea puntos de entrada para la infección por una de las especies Neonectria de hongo. Sin infestación de escala, hay pocas oportunidades para la infección por hongos. El uso de huevos de escala para infestar artificialmente árboles sanos en rodales muy BBD impactadas demostró que estos árboles fueron resistentes a la porción de insectos escala del complejo de la enfermedad 1. Aquí se presenta un protocolo que hemos desarrollado, basado en la técnica de la infestación artificial por Houston 2, que se puede utilizar para la detección de árboles escala resistentes en el campo y en las plantas de semillero y los injertos en macetas pequeñas. La identificación de los árboles escala resistente es un componente importante de la gestión de BBD través del árbolprogramas de mejora y manipulación silvicultural.

La figura 6 muestra una planta de semillero resistentes (C) y dos plantas de semillero susceptibles (D, E) que exhibe diferentes grados de susceptibilidad. Un ejemplo de lo que un árbol maduro susceptible parece a 57 semanas después de la prueba de infestación artificial fue creado se muestra en la Figura 5B. Cuando la espuma se desprendió del árbol, no es raro que las cochinillas y sus racimos de huevos para permanecer pegado a la espuma como se m...

Watch this Scientific Journal Video about A Technique to Screen American Beech for Resistance to the Beech Scale Insect Cryptococcus fagisuga Lind. at JoVE.com

A Technique to Screen American Beech for Resistance to the Beech Scale Insect Cryptococcus fagisuga Lind.

Enfermedad de la corteza de la haya es iniciada por la alimentación de las actividades de la cochinilla de haya que crear puntos de entrada de hongos en la corteza. Los árboles que son resistentes a la cochinilla son también resistentes a las enfermedades. Aquí presentamos el protocolo que hemos desarrollado para detectar árboles individuales de haya para la resistencia a escala de haya.

Una técnica a la pantalla haya americana para la resistencia al insecto Haya Scale (<em&gt; Fagisuga Cryptococcus</em&gt; Lind.)

Beech bark disease (BBD) results in high levels of initial mortality, leaving behind survivor trees that are greatly weakened and deformed.&#160;The ...

a-technique-to-screen-american-beech-for-resistance-to-beech-scale

Research

JoVE Journal

Environment

A Technique to Screen American Beech for Resistance to the Beech Scale Insect (Cryptococcus fagisuga Lind.)

9.5K vistas.  US Forest Service.  Enfermedad de la corteza de la haya es iniciada por la alimentación de las actividades de la cochinilla de haya que crear puntos de entrada de hongos en la corteza. Los árboles que son resistentes a la cochinilla son también resistentes a las enfermedades. Aquí presentamos el protocolo que hemos desarrollado para detectar árboles individuales de haya para la resistencia a escala de haya. 

Video: A Technique to Screen American Beech for Resistance to the Beech Scale Insect Cryptococcus fagisuga Lind.

A Noninvasive Method For In situ Determination of Mating Success in Female American Lobsters (Homarus americanus)

Despite being one of the most productive fisheries in the Northwest Atlantic, much remains unknown about the natural reproductive dynamics of American lobsters. Recent work in exploited crustacean populations (crabs and lobsters) suggests that there are circumstances where mature females are unable to achieve their full reproductive potential due to sperm limitation. To examine this possibility in different regions of the American lobster fishery, a reliable and noninvasive method was developed for sampling large numbers of female lobsters at sea. This method involves inserting a blunt-tipped needle into the female&#39;s seminal receptacle to determine the presence or absence of a sperm plug and to withdraw a sample that can be examined for the presence of sperm. A series of control studies were conducted at the dock and in the laboratory to test the reliability of this technique. These efforts entailed sampling 294 female lobsters to confirm that the presence of a sperm plug was a reliable indicator of sperm within the receptacle and thus, mating. This paper details the methodology and the results obtained from a subset of the total females sampled. Of the 230 female lobsters sampled from George&#39;s Bank and Cape Ann, MA (size range = 71-145 mm in carapace length), 90.3% were positive for sperm. Potential explanations for the absence of sperm in some females include: immaturity (lack of physiological maturity), breakdown of the sperm plug after being used to fertilize a clutch of eggs, and lack of mating activity. The surveys indicate that this technique for examining the mating success of female lobsters is a reliable proxy that can be used in the field to document reproductive activity in natural populations.

A Noninvasive Method For In situ Determination of Mating Success in Female American Lobsters Homarus americanus

Fishery-induced changes to exploited crustacean fisheries, such as the American lobster fishery, could potentially influence their reproductive dynamics, leading to a reduction in mating success. This study&#39;s goal was to develop a noninvasive method for ascertaining the mating success of female lobsters that may be physiologically or functionally mature.

Un método no invasivo para<em&gt; In situ</em&gt; Determinación de Éxito de apareamiento en americano femenino Langostas (<em&gt; Homarus americanus</em&gt;)

Despite being one of the most productive fisheries in the Northwest Atlantic, much remains unknown about the natural reproductive dynamics of American ...

Investigación

Medio ambiente

Video Tracking Protocol to Screen Deterrent Chemistries for Honey Bees

The European honey bee, Apis mellifera L., is an economically and agriculturally important pollinator that generates billions of dollars annually. Honey bee colony numbers have been declining in the United States and many European countries since 1947. A number of factors play a role in this decline, including the unintentional exposure of honey bees to pesticides. The development of new methods and regulations are warranted to reduce pesticide exposures to these pollinators. One approach is the use of repellent chemistries that deter honey bees from a recently pesticide-treated crop. Here, we describe a protocol to discern the deterrence of honey bees exposed to select repellent chemistries. Honey bee foragers are collected and starved overnight in an incubator 15 h prior to testing. Individual honey bees are placed into Petri dishes that have either a sugar-agarose cube (control treatment) or sugar-agarose-compound cube (repellent treatment) placed into the middle of the dish. The Petri dish serves as the arena that is placed under a camera in a light box to record the honey bee locomotor activities using video tracking software. A total of 8 control and 8 repellent treatments were analyzed for a 10 min period with each treatment was duplicated with new honey bees. Here, we demonstrate that honey bees are deterred from the sugar-agarose cubes with a compound treatment whereas honey bees are attracted to the sugar-agarose cubes without an added compound.

The loss of honey bee colonies presents a challenge to crop pollination services. Current pollinator protection practices warrant an alternative approach to minimize the contact of honey bees to harmful pesticides using repellent chemistries. Here, we provide detailed methods for a visual tracking protocol to screen deterrents for bees.

Protocolo de seguimiento de vídeo a la pantalla Químicos de disuasión para las abejas de miel

The European honey bee, Apis mellifera L., is an economically and agriculturally important pollinator that generates billions of dollars annually. Honey ...

Lab-Scale Model to Evaluate Odor and Gas Concentrations Emitted by Deep Bedded Pack Manure

A lab-scaled simulated bedded pack model was developed to study air quality and nutrient composition of deep-bedded packs used in cattle mono-slope facilities. This protocol has been used to effectively evaluate many different bedding materials, environmental variables (temperature, humidity), and potential mitigation treatments that can improve air quality in commercial deep-bedded mono-slope facilities. The model is dynamic and allows researchers to easily collect many chemical and physical measurements from the bedded pack. Weekly measurements, collected over the course of six to seven weeks, allows sufficient time to see changes in air quality measurements over time as the bedded pack matures. The data collected from the simulated bedded packs is within the range of concentrations previously measured in commercial deep-bedded mono-slope facilities. Past studies have demonstrated that 8 - 10 experimental units per treatment are sufficient to detect statistical differences among the simulated bedded packs. The bedded packs are easy to maintain, requiring less than 10 minutes of labor per bedded packs per week to add urine, feces, and bedding. Sample collection using the gas sampling system requires 20 - 30 minutes per bedded pack, depending on the measurements that are being collected. The use of lab-scaled bedded packs allows the researcher to control variables such as temperature, humidity, and bedding source that are difficult or impossible to control in a research or commercial facility. While not a perfect simulation of &#34;real-world&#34; conditions, the simulated bedded packs serve as a good model for researchers to use to examine treatment differences among bedded packs. Several lab-scale studies can be conducted to eliminate possible treatments before trying them in a research or commercial-sized facility.

A protocol has been developed to measure gases, odors, and nutrient composition in lab-scaled bedded manure packs, which can be used to study ways to improve air quality in commercial cattle facilities using deep-bedded manure packs.

Modelo de escala de laboratorio para evaluar olor y concentraciones de Gas emitidas por profundo camas Pack abono

A lab-scaled simulated bedded pack model was developed to study air quality and nutrient composition of deep-bedded packs used in cattle mono-slope ...

Double-stranded RNA Oral Delivery Methods to Induce RNA Interference in Phloem and Plant-sap-feeding Hemipteran Insects

Phloem and plant sap feeding insects invade the integrity of crops and fruits to retrieve nutrients, in the process damaging food crops. Hemipteran insects account for a number of economically substantial pests of plants that cause damage to crops by feeding on phloem sap. The brown marmorated stink bug (BMSB), Halyomorpha halys (Heteroptera: Pentatomidae) and the Asian citrus psyllid (ACP), Diaphorina citri Kuwayama (Hemiptera: Liviidae) are hemipteran insect pests introduced in North America, where they are an invasive agricultural pest of high-value specialty, row, and staple crops and citrus fruits, as well as a nuisance pest when they aggregate indoors. Insecticide resistance in many species has led to the development of alternate methods of pest management strategies. Double-stranded RNA (dsRNA)-mediated RNA interference (RNAi) is a gene silencing mechanism for functional genomic studies that has potential applications as a tool for the management of insect pests. Exogenously synthesized dsRNA or small interfering RNA (siRNA) can trigger highly efficient gene silencing through the degradation of endogenous RNA, which is homologous to that presented. Effective and environmental use of RNAi as molecular biopesticides for biocontrol of hemipteran insects requires the in vivo delivery of dsRNAs through feeding. Here we demonstrate methods for delivery of dsRNA to insects: loading of dsRNA into green beans by immersion, and absorbing of gene-specific dsRNA with oral delivery through ingestion. We have also outlined non-transgenic plant delivery approaches using foliar sprays, root drench, trunk injections as well as clay granules, all of which may be essential for sustained release of dsRNA. Efficient delivery by orally ingested dsRNA was confirmed as an effective dosage to induce a significant decrease in expression of targeted genes, such as juvenile hormone acid O-methyltransferase (JHAMT) and vitellogenin (Vg). These innovative methods represent strategies for delivery of dsRNA to use in crop protection and overcome environmental challenges for pest management.

This article demonstrates novel techniques developed for oral delivery of double-stranded RNA (dsRNA) through the vascular tissues of plants for RNA interference (RNAi) in phloem sap feeding insects.

Double-stranded RNA Oral entrega métodos para inducir la interferencia de ARN en floema e insectos Hemipteran alimentación planta de sap

Phloem and plant sap feeding insects invade the integrity of crops and fruits to retrieve nutrients, in the process damaging food crops. Hemipteran ...

The Barnacle Balanus improvisus as a Marine Model - Culturing and Gene Expression

Barnacles are marine crustaceans with a sessile adult and free-swimming, planktonic larvae. The barnacle Balanus (Amphibalanus) improvisus is particularly relevant as a model for the studies of osmoregulatory mechanisms because of its extreme tolerance to low salinity. It is also widely used as a model of settling biology, in particular in relation to antifouling research. However, natural seasonal spawning yields an unpredictable supply of cyprid larvae for studies. A protocol for the all-year-round culturing of B. improvisus has been developed and a detailed description of all steps in the production line is outlined (i.e., the establishment of adult cultures on panels, the collection and rearing of barnacle larvae, and the administration of feed for adults and larvae). The description also provides guidance on troubleshooting and discusses critical parameters (e.g., the removal of contamination, the production of high-quality feed, the manpower needed, and the importance of high-quality seawater). Each batch from the culturing system maximally yields roughly 12,000 nauplii and can deliver four batches in a week, so up to almost 50,000 larvae per week can be produced. The method used to culture B. improvisus is, probably, to a large extent also applicable to other marine invertebrates with free-swimminglarvae. Protocols are presented for the dissection of various tissues from adults as well as the production of high-quality RNA for studies on gene expression. It is also described how cultured adults and reared cyprids can be utilized in a wide array of experimental designs for examining gene expression in relation to external factors. The use of cultured barnacles in gene expression is illustrated with studies of possible osmoregulatory roles of Na+/K+ ATPase and aquaporins.

The Barnacle Balanus improvisus as a Marine Model - Culturing and Gene Expression

The barnacle Balanus (Amphibalanus) improvisus is a model for studying osmoregulation and antifouling. However, natural seasonal spawning yields an unpredictable supply of cyprid larvae. Here, a protocol for the all-year-round culturing of B. improvisus is described, including the production of larvae. The use of cultured barnacles in gene expression studies is illustrated.

El percebe Balanus improvisus como modelo Marina - cultivo y expresión del gen

Barnacles are marine crustaceans with a sessile adult and free-swimming, planktonic larvae. The barnacle Balanus (Amphibalanus) improvisus is particularly ...

Experimental Protocol for Examining Behavioral Response Profiles in Larval Fish: Application to the Neuro-stimulant Caffeine

Fish models and behaviors are increasingly used in the biomedical sciences; however, fish have long been the subject of ecological, physiological and toxicological studies. Using automated digital tracking platforms, recent efforts in neuropharmacology are leveraging larval fish locomotor behaviors to identify potential therapeutic targets for novel small molecules. Similar to these efforts, research in the environmental sciences and comparative pharmacology and toxicology is examining various behaviors of fish models as diagnostic tools in tiered evaluation of contaminants and real-time monitoring of surface waters for contaminant threats. Whereas the zebrafish is a popular larval fish model in the biomedical sciences, the fathead minnow is a common larval fish model in ecotoxicology. Unfortunately, fathead minnow larvae have received considerably less attention in behavioral studies. Here, we develop and demonstrate a behavioral profile protocol using caffeine as a model neurostimulant. Though photomotor responses of fathead minnows were occasionally affected by caffeine, zebrafish&#160;were markedly more sensitive for photomotor and locomotor endpoints, which responded at environmentally relevant levels. Future studies are needed to understand comparative behavioral sensitivity differences among fish with age and time of day, and to determine whether similar behavioral effects would occur in nature and be indicative of adverse outcomes at the individual or population levels of biological organization.

Here, we present a protocol to examine larval zebrafish and fathead minnow locomotor activities and photomotor responses (PMR) using an automated tracking software. When incorporated in common toxicology bioassays, analyses of these behaviors provide a diagnostic tool to examine chemical bioactivity. This protocol is described using caffeine, a model neurostimulant.

Protocolo experimental para examinar los perfiles de respuesta conductual en peces larvales: aplicación a la Neuro-estimulantes cafeína

Fish models and behaviors are increasingly used in the biomedical sciences; however, fish have long been the subject of ecological, physiological and ...

A Precise and Autonomous System for the Detection of Insect Emergence Patterns

Existing systems to measure insect emergence patterns have limitations; they are only partially automated and are limited in the maximum number of emerging insects they can detect. In order to obtain precise measurement of insect emergence, it is necessary for systems to be semi-automated and able to measure large numbers of emerging insects. We addressed these issues by designing and building a system that is automated and can measure emergence of up to 1200 insects. We modified the existing &#34;falling-ball&#34; system using Arduino microcontrollers to automate data collection and expand the sample size through multiple data channels. Multiple data channels enable the user to not only increase their sample size, but also allows for multiple treatments to be run simultaneously in a single experiment. Furthermore, we created an R script to automatically visualize the data as a bubble plot, while also calculating the median day and time of emergence. The current system was designed using 3D printing so the user can modify the system to be adjusted for different species of insects. The goal of this protocol is to investigate important questions in chronobiology and stress physiology, using this precise and automated system to measure insect emergence patterns.

Measurement of insect emergence patterns requires precision. Existing systems are only semi-automated and sample size is limited. We addressed these issues by designing a system using microcontrollers to precisely measure the time of emergence of large numbers of emerging insects.

Un sistema preciso y autónomo para la detección de patrones de aparición de insectos

Existing systems to measure insect emergence patterns have limitations; they are only partially automated and are limited in the maximum number of ...

Composition and Distribution Analysis of Bioaerosols Under Different Environmental Conditions

Variable microorganisms in particulate matter (PM) under different environmental conditions may have significant impacts on human health. In this study, we described a protocol for multiple analyses of the biological compositions in environmental PM. Five experiments are presented: (1) PM number monitoring by using a laser particle counter; (2) PM collection by using a cyclonic aerosol sampler; (3) PM collection by using a high-volume air sampler with filters; (4) culturable microbes collection by the Andersen six-stage sampler; and (5) detection of biological composition of environmental PM by bacterial 16SrDNA and fungal ITS region sequencing. We selected hazy days and a livestock farm as two typical examples of the application in this protocol. In this study, these two sampling methods, cyclonic aerosol sampler and filter sampler, showed different sampling efficiency. The cyclonic aerosol sampler performed much better in terms of collecting bacteria, while these two methods showed the same efficiency in collecting fungi. Filter samplers can work under low temperature conditions while cyclonic aerosol samplers have a sampling limitation for temperature. A solid impacting sampler, such as an Andersen six-stage sampler, can be used to sample bioaerosols directly into the culture medium, which increases the survival rate of culturable microorganisms. However, this method mainly relies on culture while more than 99% of microbes cannot be cultured. DNA extracted from the culturable bacteria collected by the Andersen six-stage sampler and samples collected by cyclonic aerosol sampler and filter sampler were detected by bacterial 16S rDNA and fungal ITS region sequencing.All the methods above may have wide application in many fields of study, such as environmental monitoring and airborne pathogen detection. From these results, we can conclude that these methods can be used under different conditions and may help other researchers further explore the health impacts of environmental bioaerosols.

Understanding the biological composition of environmental particulate matter is important for the study of its significant impacts on human health and disease spread. Here, we used three types of bioaerosol sampling methods and a biological analysis of airborne microbes to better explore airborne microbial communities under different environmental conditions.

Composición y análisis de la distribución de bioaerosoles bajo diferentes condiciones ambientales

Variable microorganisms in particulate matter (PM) under different environmental conditions may have significant impacts on human health. In this study, ...

Detached Leaf Assays to Simplify Gene Expression Studies in Potato During Infestation by Chewing Insect Manduca sexta

The multitrophic nature of gene expression studies of insect herbivory demands large numbers of biological replicates, creating the need for simpler, more streamlined herbivory protocols. Perturbations of chewing insects are usually studied in whole plant systems. While this whole organism strategy is popular, it is not necessary if similar observations can be replicated in a single detached leaf. The assumption is that basic elements required for signal transduction are present within the leaf itself. In the case of early events in signal transduction, cells need only to receive the signal from the perturbation and transmit that signal to neighboring cells which are assayed for gene expression.
The proposed method simply changes the timing of the detachment. In whole plant experiments, larvae are confined to a single leaf which is eventually detached from the plant and assayed for gene expression. If the order of excision is reversed, from last in whole plant studies, to first in the detached study, the feeding experiment is simplified.
Solanum tuberosum var. Kennebec is propagated by nodal transfer in a simple tissue culture medium and transferred to soil for further growth if desired. Leaves are excised from the parent plant and relocated to Petri dishes where the feeding assay is conducted with the larval stages of M. sexta. Damaged leaf tissue is assayed for the expression of relatively early events in signal transduction. Gene expression analysis identified infestation-specific Cys2-His2 (C2H2) transcription factors, confirming the success of using detached leaves in early response studies. The method is easier to perform than whole plant infestations and uses less space.

The presented method creates natural herbivore damaged plant tissue through the application of Manduca sexta larvae to detached leaves of potato. The plant tissue is assayed for expression of six transcription factor homologs involved in early responses to insect herbivory.

Ensayos de hojas independientes para simplificar los estudios de expresión génica en papa durante la infestación por masticar insectos Manduca sexta

The multitrophic nature of gene expression studies of insect herbivory demands large numbers of biological replicates, creating the need for simpler, more ...

Implantation of a New Micro Acoustic Tag in Juvenile Pacific Lamprey and American Eel

Juvenile Pacific Lamprey and American eels were used for laboratory evaluations to determine potential effects from tag implantation. Telemetry technology has been identified as a way to obtain more detailed information on movement and behavior across a broader spatial scale than is possible with other known technology. The purpose of this method is to provide a detailed step by step instruction on tag implantation for both lampreys and eel. For laboratory studies using actively migrating juvenile Pacific Lamprey (120-160 mm), we determined that the presence of the tag did not alter the swimming ability between tagged and untagged Individuals or have any significant tag loss (&lt;3%). Similar results were determined during laboratory testing of Yellow phase American Eels (113-175 mm). No mortality occurred during a 38-day holding period and there was minimal tag loss (3.8%). The presence of the tag did not have any significant effect on the swimming ability or survival of tagged eels compared to untagged controls and there was minimal tag loss.

This article illustrates the procedure for implanting a micro acoustic tag in juvenile Pacific Lamprey and American Eel. Our application is based on laboratory trials, which indicate that acoustic tags can be successfully implanted with no apparent effects on swimming ability or survival and minimal tag loss.

Implantación de una nueva etiqueta acústica Micro en juvenil Pacífico lamprea y anguila americana

Juvenile Pacific Lamprey and American eels were used for laboratory evaluations to determine potential effects from tag implantation. Telemetry technology ...