This work was funded, in part, by the Great Lakes Fishery Commission and the Annis Water Resources Institute. Use of trade, product, or firm names does not imply endorsement by the U. S. Government. This article is Contribution 1867 of the U. S. Geological Survey Great Lakes Science Center.

1. Experimento Laboratorio <ol><li> Obtener los peces presa para la alimentación de los peces depredadores durante el experimento. Preferiblemente estos peces presa deben ser capturados en el campo, congelados, y se almacenaron a aproximadamente -30 ° C. Considere la posibilidad de operaciones de pesca comercial como una fuente potencial para los peces presa. </li><li> Dar a conocer a los peces depredadores en los tanques de laboratorio que se utilizarán para el experimento. Hasta 15 peces depredadores se han introducido en cada uno de los tanques 870-L, y hasta 30 peces depredadores se han introducido en cada uno de los tanques de 2380 L en estudios previos 16,18. </li><li> Aclimatar el pez depredador a una dieta de los peces presa seleccionada. Una vez aclimatados, el pez depredador debe permanecer en esta dieta durante 1-3 meses antes de comenzar el experimento. </li><li> Ponga a un lado las muestras de peces presa con selección aleatoria de 10 a 20 muestras compuestas del lote de peces presa. Número de peces presa en una muestra compuesta puede variar desde 3 a 100, dependiendo del tamaño de la presapeces. Cada muestra compuesta debe ser doble bolsa, congelado, y se almacenan a -30 ° C. </li><li> Iniciar el experimento al sacrificar 30 a 50% de los peces en cada uno de los tanques. <ol><li> Para practicar la eutanasia a los peces, mezclar 8 g de Finquel con 45 litros de agua en un recipiente grande de plástico y luego coloque el pescado en el contenedor con la solución Finquel. </li><li> Una vez sacrificados, coloque todos los peces sacrificados de un tanque en una bolsa, la bolsa luego de matrimonio, y se almacenan a -30 ° C hasta el momento de la tramitación. </li><li> Pesar cada uno de los peces restantes en cada uno de los tanques, y registrar los pesos; es probable que se necesita un anestésico para llevar a cabo el pesaje. </li><li> Para anestesiar a los peces, mezclar 4,6 g de Finquel con 45 litros de agua en un recipiente grande de plástico, y luego coloque el pescado en el contenedor con la solución Finquel. </li><li> Espere unos minutos para que la anestesia surta efecto antes de pesar el pescado. </li></ol></li><li> En cada día del experimento, descongelar uncantidad apropiada de peces presa, y cortar el pescado presa en pedazos que pesan aproximadamente 1 a 5 g. Pesar la cantidad de peces presa que se colocará en cada uno de los tanques, a continuación, colocar los trozos de pescado presa en cada tanque y permitir que el pez depredador alrededor de 1 hora para alimentar. A continuación, retire todos los alimentos no consumidos, permitir que los alimentos se sequen al aire durante aproximadamente 20 minutos, y luego pesar el alimento no consumido para cada uno de los tanques. Registrar la cantidad de alimento colocado en el tanque y la cantidad de alimento no consumido para cada uno de los tanques de cada día. NOTA: Para el experimento representativo, se alimentaron de la trucha del lago tanta comida como ellos consumirían durante un período de alimentación cada día 18. Sin embargo, los peces depredadores también podría ser colocado en raciones fijas 16,19. </li><li> Terminar el experimento al sacrificar todos los peces depredador que queda en cada uno de los tanques. Para practicar la eutanasia a los peces, mezclar 8 g de Finquel con 45 litros de agua en un recipiente grande de plástico y luego coloque el pescado en el contenedor con la solución Finquel. Record tque el peso de cada uno de los peces sacrificado. Para obtener resultados fiables, el experimento debe funcionar durante al menos 100 días, preferiblemente durante al menos 130 días. Coloque todos los peces de un tanque en una bolsa, la bolsa luego de matrimonio, y se almacenan a -30 ° C hasta el momento de su procesamiento. </li></ol> Homogeneización 2. Pescado <ol><li> Seleccione un conjunto de peces depredadores y / o compuestos de peces de presa para la descongelación. Permita que los compuestos que se descongele parcialmente. Cada material compuesto puede requerir de 0,5 a 1 h para homogeneizar. </li><li> El uso de los mezcladores de tamaño apropiado, homogeneizar cada uno de los compuestos. Para cada compuesto, colocar una muestra (de 50 a 100 g) del homogeneizado en un frasco de limpiado, se enjuagó-acetona, y etiquetados. Luego tapar el frasco y almacenar el frasco a aproximadamente -30 ° C hasta el momento del procesamiento. </li><li> Lave todo el equipo utilizado para homogeneizar los peces, y luego enjuague bien con agua destilada y metanol, entre las muestras. </li></ol> 3. Extracción <ol><li> Pesar 20,0 gde descongelado tejido de los peces se homogeneizaron en un vaso de 200 ml. </li><li> Añadir aproximadamente 40 g de sulfato de sodio y mezclar bien con una espátula. </li><li> Añadir la solución sustituta que contiene pico congéneres 30, 61, 161, y 166 de Spike a una concentración que produce una concentración final de 20 ng / ml en el extracto. </li><li> Deje que la muestra se seque a temperatura ambiente mientras se mezcla cada 20 minutos. </li><li> Permitir que la muestra alcance una consistencia de arena seca, momento en el que la muestra está lista para la extracción. </li><li> La instalación del equipo de extracción Soxhlet con un frasco de 500 ml que contiene chips de hervir teflón, Soxhlet, y del condensador. </li><li> Añadir la mezcla de pescado seco a un dedal de vidrio con una parte inferior del disco grueso vitrificado o dedal de papel. </li><li> Añadir 150 ml de hexano al 50% y 50% de diclorometano en el vaso utilizado para la muestra y se agita mientras raspar las paredes del vaso de precipitados con una espátula. </li><li> Transfiera el disolvente a la parte superior de la Soxhlet con el frasco adjunto y permitir que el ciclo a través de la Soxhlet y enal matraz. </li><li> Repetir una segunda vez con 150 ml de nuevo. </li><li> Colocar el matraz de Soxhlet con el adjunto en el elemento de calentamiento y conecte el condensador. </li><li> Encienda el elemento de calefacción y llevar el disolvente a ebullición suave, a continuación, extraer durante un mínimo de 16 horas asegurándose de que el agua fría se suministra a los condensadores. </li><li> Después de dejar que el disolvente se enfríe, revise para ver si alguno de los frascos de muestra contienen agua. Para los matraces que contienen agua, añadir sulfato de sodio y agitar hasta que el agua es absorbida por el sulfato de sodio. </li><li> Concentrar la muestra utilizando un concentrador de muestra de nitrógeno o un Kaderna danesa (KD) de instalación de vidrio con un baño de agua caliente. </li><li> Permitir que la muestra se evapore hasta un volumen de menos de 2 ml, y luego llevar a un volumen final de 5 ml mediante el uso de pequeños lavados de hexano para transferir la muestra del material de vidrio utilizado para un matraz volumétrico de 5 ml. </li><li> Pasar a un vial de 10 ml y la etiqueta con información de la muestra. </li></ol><ol><li> Preparar gel de sílice se acidificó mediante la adición de 44 g de ácido sulfúrico concentrado a 100 g de gel de sílice activado. </li><li> Añadir 10 g de gel de sílice acidificado en una columna de cromatografía pequeño que contiene un pequeño tapón de lana de vidrio en la parte inferior. </li><li> Añadir 1 ml de extracto de la muestra a la columna después de la pre-limpieza de la columna con 10 ml de hexano. </li><li> Se eluye la columna con 20 ml de hexano y recoger en un tubo de vidrio de 20 ml cónico. </li><li> Colocar el tubo de vidrio en el aparato evaporador de nitrógeno (N-Vap) bajo una corriente de nitrógeno y se sumerge en agua caliente. </li><li> Evaporar a menos de 1 ml pero no a sequedad. </li><li> Retirar de la fuente N-Vap y traslado al matraz aforado de 1 ml con pequeños lavados de hexano. </li><li> Pasar a un 1,8 ml vial automuestreador etiquetado con información de la muestra. </li><li> Pico 4 l de estándar interno en el vial. La muestra está lista para su análisis. </li></ol> 5. Analanalysis por cromatografía de gases - espectrometría de masas de ionización química negativa Uso <ol><li> Utilice estándares para calibrar el instrumento: Las normas están disponibles en las mezclas que consisten en grupos de congéneres bien separados. Mezclas 1-5 consisten en casi todos los congéneres que se encuentran en arocloros 1016, 1221, 1232, 1242, 1248, 1254, y 1260 Mix 1 se utiliza como una mezcla de calibración multi-nivel, y la linealidad del sistema se confirma mediante la preparación de al menos cinco niveles de calibración en concentraciones entre 2 y 100 ng / ml. Mezclas 2-5 se utilizan como calibraciones de un solo punto para cada congénere. </li><li> Configurar el cromatografía - sistema de espectrometría de masas en el modo de ionización química negativa con hidrógeno como gas portador (1 ml / min) y metano como gas reactivo. </li><li> Utilice una de sílice fundida, columna capilar (60 m × 0,25 mm de diámetro interior) recubiertas con DB-XLB al espesor de la película de 0,25 micras para la separación. Programa de temperatura del horno 60 a 212 ° C a 25 ° C / min, luego a 260 ° C a 1 ° C / min, y luego a 280 ° C a 4 ° C / min, con un tiempo de retención final de 4 min. Las temperaturas del inyector y línea de transferencia deben establecerse a 280 ° C. Inyectar 1 a 2 l de la muestra usando el modo de inyección sin división. </li><li> Analizar todos los estándares y las muestras por el método estándar interno utilizando 13 decaclorobifenilo C-etiquetados. </li><li> Lleve a cabo una comprobación de la calibración inicial mediante la ejecución de un segundo estándar de código y Aroclors 1242 y 1260, y luego comparar los valores previstos para los congéneres Aroclor con las cantidades observadas de este procedimiento de revisión. </li><li> Una vez que el procedimiento de calibración inicial se ha realizado con éxito, el análisis completo de todas las muestras. Ejecute una comprobación de una calibración cada diez muestras, utilizando cualquiera de las mezclas de calibración de la calibración inicial. </li></ol> 6. Cálculo del Patrimonio trófico Eficiencia de Transferencia <ol><li> Calcular la eficiencia neta de transferencia trófica, γ, para cada combinación of tanque y congénere PCB utilizando la siguiente ecuación: <img alt="Ecuación 1" fo:content-width="2in" src="/files/ftp_upload/51496/51496eq1.jpg" /> , Donde [PCB f] es la concentración promedio de los congéneres de PCB de los peces depredadores en el tanque al final del experimento, f W es el peso medio de los peces depredadores en el tanque al final del experimento, [PCB i] es la concentración media de los congéneres de PCB de los peces depredadores en el recipiente al inicio del experimento, W i es el peso medio de los peces depredador al comienzo del experimento, y la cantidad de congénere PCB ingerido se refiere al peso de la congénere PCB ingiere, en promedio, por cada una trucha de lago en el tanque durante el curso del experimento. </li><li> Calcular el denominador en la ecuación anterior, multiplicando la concentración media del congénere PCB en los materiales compuestos de peces presa por la cantidad promedio (peso) de los peces presa comido por pez depredador en el dur tanqueing todo el curso del experimento. </li></ol>

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C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Weighted+least-squares+approach+to+calculating+limits+of+detection+and+quantification+by+modeling+variability+as+a+function+of+concentration.">Weighted least-squares approach to calculating limits of detection and quantification by modeling variability as a function of concentration.</a> Analytical Chemistry. 69 (15), 3069-3075 (1997).</li></ol>

Para las estimaciones más exactas de γ, el experimentador debe ser capaz de rastrear con precisión tanto la cantidad de alimento colocado en cada uno de los tanques y la cantidad de alimento no consumido en cada uno de los tanques durante el curso del experimento. Para lograr esto, el experimentador debe ser capaz de eliminar todos los restos de comida de los tanques y determinar con precisión su peso. Además de un seguimiento preciso de la comida en realidad comido por el pez depredador, estimación precisa de γ ...

Of all of the factors affecting the rate at which fish accumulate contaminants, the efficiency with which fish retain contaminants from the food that they eat is one of the most important1-3. Risk assessment models have been developed to predict future risks to both people and wildlife eating contaminated fish under various scenarios of environmental contamination, and the reliability of these predictions critically depends on the accuracy of the estimates of the efficiency at which fish retain contaminants from their food4.
The efficiency with which the contaminant in the food ingested by the predator is transported through the gut wall is known as gross trophic transfer efficiency5. A portion of the quantity of the contaminant transported through the gut wall of the predator may eventually be lost through depuration and/or metabolic transformation. The efficiency with which the contaminant in the food ingested by the predator is retained by the predator, including any losses due to elimination and metabolic transformation, is known as net trophic transfer efficiency6.
Gross trophic transfer efficiency of organic contaminants to fish from their prey appears to vary with the contaminant&#8217;s chemical properties, including lipid affiliation as measured by the octanol-water partition coefficient, Kow3,7. According to an empirical relationship developed by Thomann3, gross trophic transfer efficiency is relatively high when log Kow is equal to a value between 5 and 6. Gross trophic transfer efficiency declines exponentially at a rate of 50% per unit of log Kow as log Kow increases from 6 to 10, according to the Thomann3 relationship.
Nevertheless, the gross and net trophic transfer efficiencies of polychlorinated biphenyl (PCB) congeners to fish from their prey do not appear to follow the Thomann3 relationship in most cases. Although the trophic transfer efficiencies of PCB congeners to lake whitefish (Coregonus clupeaformis) from its food followed the relationship proposed by Thomann8, trophic transfer efficiencies of PCB congeners were either just weakly related or not related at all to log Kow for Atlantic salmon (Salmo salar)9, rainbow trout (Oncorhynchus mykiss)10, coho salmon (Oncorhynchus kisutch)11, and northern pike (Esox lucius)11.
The overall goal of this study was to present a laboratory technique for estimating the net trophic transfer efficiencies of PCB congeners to a piscivorous fish from its prey. Lake trout (Salvelinus namaycush) was chosen as the piscivorous fish for our experiment because lake trout are relatively easy to maintain in laboratory tanks. Bloater (Coregonus hoyi) was selected as the prey fish to be fed to the lake trout because bloater is eaten by lake trout in its natural setting12. In addition, we determined whether the net trophic transfer efficiencies for lake trout estimated from our laboratory experiment followed the Thomann3 relationship. We also determined whether the degree of activity by the lake trout had a significant effect on net trophic transfer efficiency (&#947;) of the PCB congeners. Activity by lake trout in the Laurentian Great Lakes is believed to have recently increased because changes in the food webs have caused lake trout to allocate more energy toward searching for food13. Lake trout were forced to exercise in one set of tanks by subjecting these lake trout to relatively high flow rates, whereas the other lake trout were permitted to remain relatively inactive by subjecting them to relatively low flow rates. Finally, the specific details of our laboratory procedure that need to be carefully followed to ensure the highest degree of accuracy in the &#947; estimates and to make the laboratory results applicable to the field are discussed, as well as future directions for research building on our laboratory technique. Net trophic transfer efficiency can be estimated both in the laboratory and in the field, and advantages and disadvantages are associated with both approaches. Accuracy in the estimate of &#947; depends on the accuracy of the estimate of food consumption. The amount of food eaten by fish in the laboratory can be accurately determined when proper protocols are followed, whereas the amount of food eaten by fish in the field is typically estimated via bioenergetics modeling. Use of bioenergetics modeling to derive the amount of food eaten has the potential to introduce a substantial amount of uncertainty into the estimates of food consumption. Fish bioenergetics models have been shown to estimate food consumption with no detectable bias for the case of lake trout14,15, but considerable bias in bioenergetics model estimates of food consumption has been detected for the case of lake whitefish15,16. On the other hand, estimates of net trophic transfer efficiency estimated in the laboratory may not be applicable to the field due to a difference in feeding rates between the laboratory and the field17. Evidence from both the laboratory and the field suggest that feeding rate can influence &#947;14,17.
The methodology used in the present study for estimating &#947; in the laboratory is applicable to situations where the predator fish is fed prey fish, and the amount of prey fish eaten by the predator can be accurately tracked. To accomplish this, the experimenter must weigh all of the food before placement in the tank; and the experimenter must be able to remove all of the uneaten food from the tank, and then weigh the uneaten food. In addition, an adequate suite of mixers and blenders should be available to obtain a sufficient degree of homogenization of the samples of both predator and prey fish. Finally, the gas chromatography &#8211; mass spectrometry instrumentation used to determine the PCB congener concentrations must be capable of detecting and quantifying individual PCB congeners at relatively low concentrations.

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			<td style="width:167px;">Comments</td>
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			<td height="21" style="height:21px;width:259px;">870-L fiberglass tanks</td>
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			<td height="68" style="height:68px;width:259px;">Tricaine methanesulfonate (Finquel)</td>
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			<td style="width:167px;">Eugenol could also be used as an anesthetic.</td>
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			<td height="21" style="height:21px;width:259px;">Sodium sulfate, anhydrous - granular</td>
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			<td style="width:87px;">Wilmad-Lab Glass</td>
			<td style="width:119px;">LG-7070-114</td>
			<td></td>
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			<td height="21" style="height:21px;width:259px;">Teflon boiling chips</td>
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			<td style="width:119px;">919120</td>
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			<td height="21" style="height:21px;width:259px;">Rapid Vap nitrogen sample concentrator</td>
			<td style="width:87px;">Labconco</td>
			<td style="width:119px;">7910000</td>
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			<td height="21" style="height:21px;width:259px;">N-Vap nitrogen concentrator</td>
			<td style="width:87px;">Organomation</td>
			<td style="width:119px;">112</td>
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			<td height="35" style="height:35px;width:259px;">Soxhlet extraction glassware (500 mL)</td>
			<td style="width:87px;">Wilmad-Lab Glass</td>
			<td style="width:119px;">&#160;LG-6900-104</td>
			<td></td>
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		<tr height="35">
			<td height="35" style="height:35px;width:259px;">Hexane</td>
			<td style="width:87px;">Burdick &#38; Jackson&#160;</td>
			<td style="width:119px;">Cat. 211-4</td>
			<td></td>
		</tr>
		<tr height="35">
			<td height="35" style="height:35px;width:259px;">Dichloromethane</td>
			<td style="width:87px;">Burdick &#38; Jackson&#160;</td>
			<td style="width:119px;">Cat. 300-4</td>
			<td></td>
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			<td height="21" style="height:21px;width:259px;">Silica gel</td>
			<td style="width:87px;">BDH</td>
			<td style="width:119px;">Cat. BDH9004-1KG</td>
			<td></td>
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		<tr height="35">
			<td height="35" style="height:35px;width:259px;">Labl Line 5000 mult-unit extraction heater</td>
			<td style="width:87px;">Lab Line Instruments</td>
			<td style="width:119px;"></td>
			<td></td>
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			<td height="35" style="height:35px;width:259px;">Agilent 5973 GC/MS with chemical ionization</td>
			<td style="width:87px;">Agilent</td>
			<td style="width:119px;">5973N</td>
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		<tr height="52">
			<td height="52" style="height:52px;width:259px;">Internal standard solution&#160;</td>
			<td style="width:87px;">Cambridge Isotope Laboratories</td>
			<td style="width:119px;">EC-1410-1.2</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:259px;">PCB congener calibration standards</td>
			<td style="width:87px;">Accustandard</td>
			<td style="width:119px;">C-CSQ-SET</td>
			<td></td>
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			<td height="21" style="height:21px;width:259px;">DB-XLB column (60m x 0.25mm, 0.25 micron)</td>
			<td style="width:87px;">Agilent/ J&#38;W</td>
			<td style="width:119px;">122-1262</td>
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laboratory estimation of net trophic transfer efficiencies of pcb congeners to lake trout (salvelinus namaycush) from its prey

Una técnica para la estimación de la eficiencia de laboratorio red trófica de transferencia (γ) de bifenilos policlorados (PCB) congéneres para los peces piscívoros de sus presas se describe en este documento. Durante un experimento de laboratorio de 135 días, nos alimentamos bloater (Coregonus hoyi) que habían sido capturados en el Lago Michigan para la trucha de lago (Salvelinus namaycush) mantenido en ocho tanques de laboratorio. Bloater es una presa natural para la trucha de lago. En cuatro de los tanques, se utilizó una velocidad de flujo relativamente alta para asegurar una actividad relativamente alta por la trucha de lago, mientras que se utilizó un caudal bajo en los otros cuatro tanques, lo que permite una baja actividad trucha de lago. Sobre una base del tanque por tanque, se registró la cantidad de alimento ingerido por la trucha de lago en cada día del experimento. Cada trucha de lago se pesó al comienzo y al final del experimento. Cuatro a nueve trucha de lago de cada uno de los ocho tanques fueron sacrificados al inicio del experimento, y todo trucha 10 lago que queda en cada uno de los tanques fueron euthanzado al final del experimento. Se determinó la concentración de 75 congéneres de PCB en la trucha de lago en el inicio del experimento, en la trucha de lago, al final del experimento, y en bloaters alimentamos a la trucha de lago durante el experimento. En base a estas mediciones, γ se calculó para cada uno de 75 congéneres de PCB en cada uno de los ocho tanques. Significan γ se calculó para cada uno de los 75 congéneres de PCB para la trucha de lago activa e inactiva. Debido a que el experimento se repitió en ocho tanques, el error estándar de decir γ podría estimarse. Los resultados de este tipo de experimentos son útiles en los modelos de evaluación de riesgo para predecir el riesgo futuro de los seres humanos y la vida silvestre que comen peces contaminados en diversos escenarios de la contaminación ambiental.

Trucha de lago mostró una cantidad sustancial de crecimiento durante el experimento, como la trucha iniciales lago significan pesos oscilaron desde 694 hasta 907 g, mientras que las truchas finales lago significan pesos variaron de 853 a 1.566 g (Tabla 1). El importe medio de los alimentos consumidos por una trucha de lago durante el curso del experimento de 135 días osciló entre 641 a 2.649 g. La media de las concentraciones de congéneres de PCB en la trucha de lago aumentó durante el experimento,...

Watch this Scientific Journal Video about Laboratory Estimation of Net Trophic Transfer Efficiencies of PCB Congeners to Lake Trout Salvelinus namaycush from Its Prey at JoVE.com

Laboratory Estimation of Net Trophic Transfer Efficiencies of PCB Congeners to Lake Trout Salvelinus namaycush from Its Prey

Se presenta una técnica para la estimación de laboratorio de eficiencia de transferencia trófica neto de bifenilos policlorados (PCB) congéneres para los peces piscívoros de sus presas. Para maximizar la aplicabilidad de los resultados de laboratorio al campo, los peces piscívoros se debe alimentar peces presa que normalmente se come en el campo.

Estimación Laboratorio de red trófica eficiencias de transferencia de los congéneres de PCB a la trucha de lago (<em&gt; Salvelinus namaycush</em&gt;) Desde Su Presa

A technique for laboratory estimation of net trophic transfer efficiency (&#947;) of polychlorinated biphenyl (PCB) congeners to piscivorous fish from ...

laboratory-estimation-net-trophic-transfer-efficiencies-pcb-congeners

Research

JoVE Journal

Environment

Laboratory Estimation of Net Trophic Transfer Efficiencies of PCB Congeners to Lake Trout (Salvelinus namaycush) from Its Prey

10.9K vistas.  U. S. Geological Survey.  Se presenta una técnica para la estimación de laboratorio de eficiencia de transferencia trófica neto de bifenilos policlorados (PCB) congéneres para los peces piscívoros de sus presas. Para maximizar la aplicabilidad de los resultados de laboratorio al campo, los peces piscívoros se debe alimentar peces presa que normalmente se come en el campo. 

Video: Laboratory Estimation of Net Trophic Transfer Efficiencies of PCB Congeners to Lake Trout Salvelinus namaycush from Its Prey

Laboratory-determined Phosphorus Flux from Lake Sediments as a Measure of Internal Phosphorus Loading

Eutrophication is a water quality issue in lakes worldwide, and there is a critical need to identify and control nutrient sources. Internal phosphorus (P) loading from lake sediments can account for a substantial portion of the total P load in eutrophic, and some mesotrophic, lakes. Laboratory determination of P release rates from sediment cores is one approach for determining the role of internal P loading and guiding management decisions. Two principal alternatives to experimental determination of sediment P release exist for estimating internal load: in situ measurements of changes in hypolimnetic P over time and P mass balance. The experimental approach using laboratory-based sediment incubations to quantify internal P load is a direct method, making it a valuable tool for lake management and restoration.
Laboratory incubations of sediment cores can help determine the relative importance of internal vs. external P loads, as well as be used to answer a variety of lake management and research questions. We illustrate the use of sediment core incubations to assess the effectiveness of an aluminum sulfate (alum) treatment for reducing sediment P release. Other research questions that can be investigated using this approach include the effects of sediment resuspension and bioturbation on P release.
The approach also has limitations. Assumptions must be made with respect to: extrapolating results from sediment cores to the entire lake; deciding over what time periods to measure nutrient release; and addressing possible core tube artifacts. A comprehensive dissolved oxygen monitoring strategy to assess temporal and spatial redox status in the lake provides greater confidence in annual P loads estimated from sediment core incubations.

Lake eutrophication is a water quality issue worldwide, making the need to identify and control nutrient sources critical. Laboratory determination of phosphorus release rates from sediment cores is a valuable approach for determining the role of internal phosphorus loading and guiding management decisions.

Laboratorio-determinado Fósforo Flujo de sedimentos lacustres como medida de fósforo Interna Loading

Eutrophication is a water quality issue in lakes worldwide, and there is a critical need to identify and control nutrient sources. Internal phosphorus (P) ...

Investigación

Medio ambiente

A Fish-feeding Laboratory Bioassay to Assess the Antipredatory Activity of Secondary Metabolites from the Tissues of Marine Organisms

Marine chemical ecology is a young discipline, having emerged from the collaboration of natural products chemists and marine ecologists in the 1980s with the goal of examining the ecological functions of secondary metabolites from the tissues of marine organisms. The result has been a progression of protocols that have increasingly refined the ecological relevance of the experimental approach. Here we present the most up-to-date version of a fish-feeding laboratory bioassay that enables investigators to assess the antipredatory activity of secondary metabolites from the tissues of marine organisms. Organic metabolites of all polarities are exhaustively extracted from the tissue of the target organism and reconstituted at natural concentrations in a nutritionally appropriate food matrix. Experimental food pellets are presented to a generalist predator in laboratory feeding assays to assess the antipredatory activity of the extract. The procedure described herein uses the bluehead, Thalassoma bifasciatum, to test the palatability of Caribbean marine invertebrates; however, the design may be readily adapted to other systems. Results obtained using this laboratory assay are an important prelude to field experiments that rely on the feeding responses of a full complement of potential predators. Additionally, this bioassay can be used to direct the isolation of feeding-deterrent metabolites through bioassay-guided fractionation. This feeding bioassay has advanced our understanding of the factors that control the distribution and abundance of marine invertebrates on Caribbean coral reefs and may inform investigations in diverse fields of inquiry, including pharmacology, biotechnology, and evolutionary ecology.

This bioassay employs a model predatory fish to assess the presence of feeding-deterrent metabolites from organic extracts of the tissues of marine organisms at natural concentrations using a nutritionally comparable food matrix.

Un pez de amamantar Laboratorio Bioensayo para evaluar la actividad antidepredación de metabolitos secundarios de los tejidos de organismos marinos

Marine chemical ecology is a young discipline, having emerged from the collaboration of natural products chemists and marine ecologists in the 1980s with ...

Automated Gel Size Selection to Improve the Quality of Next-generation Sequencing Libraries Prepared from Environmental Water Samples

Next-generation sequencing of environmental samples can be challenging because of the variable DNA quantity and quality in these samples. High quality DNA libraries are needed for optimal results from next-generation sequencing. Environmental samples such as water may have low quality and quantities of DNA as well as contaminants that co-precipitate with DNA. The mechanical and enzymatic processes involved in extraction and library preparation may further damage the DNA. Gel size selection enables purification and recovery of DNA fragments of a defined size for sequencing applications. Nevertheless, this task is one of the most time-consuming steps in the DNA library preparation workflow. The protocol described here enables complete automation of agarose gel loading, electrophoretic analysis, and recovery of targeted DNA fragments. 
In this study, we describe a high-throughput approach to prepare high quality DNA libraries from freshwater samples that can be applied also to other environmental samples. We used an indirect approach to concentrate bacterial cells from environmental freshwater samples; DNA was extracted using a commercially available DNA extraction kit, and DNA libraries were prepared using a commercial transposon-based protocol. DNA fragments of 500 to 800 bp were gel size selected using Ranger Technology, an automated electrophoresis workstation. Sequencing of the size-selected DNA libraries demonstrated significant improvements to read length and quality of the sequencing reads.

This manuscript describes an automated gel size selection approach for purifying DNA fragments for next-generation sequencing. The Ranger Technology provides complete automation of the entire process of agarose gel loading, electrophoretic analysis, and recovery of targeted DNA fragments allowing for high-throughput and high quality next-generation sequencing libraries.

Automatizado Gel Tamaño Selección para mejorar la calidad de las bibliotecas de secuenciación de próxima generación Preparado a partir de muestras de agua del medio ambiente

Next-generation sequencing of environmental samples can be challenging because of the variable DNA quantity and quality in these samples. High quality DNA ...

Removal of Trace Elements by Cupric Oxide Nanoparticles from Uranium In Situ Recovery Bleed Water and Its Effect on Cell Viability

In situ recovery (ISR) is the predominant method of uranium extraction in the United States. During ISR, uranium is leached from an ore body and extracted through ion exchange. The resultant production bleed water (PBW) contains contaminants such as arsenic and other heavy metals. Samples of PBW from an active ISR uranium facility were treated with cupric oxide nanoparticles (CuO-NPs). CuO-NP treatment of PBW reduced priority contaminants, including arsenic, selenium, uranium, and vanadium. Untreated and CuO-NP treated PBW was used as the liquid component of the cell growth media and changes in viability were determined by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay in human embryonic kidney (HEK 293) and human hepatocellular carcinoma (Hep G2) cells. CuO-NP treatment was associated with improved HEK and HEP cell viability. Limitations of this method include dilution of the PBW by growth media components and during osmolality adjustment as well as necessary pH adjustment. This method is limited in its wider context due to dilution effects and changes in the pH of the PBW which is traditionally slightly acidic&#160;however; this method could have a broader use assessing CuO-NP treatment in more neutral waters.

Removal of Trace Elements by Cupric Oxide Nanoparticles from Uranium In Situ Recovery Bleed Water and Its Effect on Cell Viability

Production bleed water (PBW) was treated with cupric oxide nanoparticles (CuO-NPs) and cellular toxicity was assessed in cultured human cells. The goal of this protocol was to integrate the native environmental sample into a cell culture format assessing the changes in toxicity due to CuO-NP treatment.

La eliminación de oligoelementos por cúprico Óxido nanopartículas de uranio<em&gt; In Situ</em&gt; Recuperación de purga de agua y su efecto sobre la viabilidad celular

In situ recovery (ISR) is the predominant method of uranium extraction in the United States. During ISR, uranium is leached from an ore body and extracted ...

Laboratory Simulation of an Iron(II)-rich Precambrian Marine Upwelling System to Explore the Growth of Photosynthetic Bacteria

A conventional concept for the deposition of some Precambrian Banded Iron Formations (BIF) proceeds on the assumption that ferrous iron [Fe(II)] upwelling from hydrothermal sources in the Precambrian ocean was oxidized by molecular oxygen [O2] produced by cyanobacteria. The oldest BIFs, deposited prior to the Great Oxidation Event (GOE) at about 2.4 billion years (Gy) ago, could have formed by direct oxidation of Fe(II) by anoxygenic photoferrotrophs under anoxic conditions. As a method for testing the geochemical and mineralogical patterns that develop under different biological scenarios, we designed a 40 cm long vertical flow-through column to simulate an anoxic Fe(II)-rich marine upwelling system representative of an ancient ocean on a lab scale. The cylinder was packed with a porous glass bead matrix to stabilize the geochemical gradients, and liquid samples for iron quantification could be taken throughout the water column. Dissolved oxygen was detected non-invasively via optodes from the outside. Results from biotic experiments that involved upwelling fluxes of Fe(II) from the bottom, a distinct light gradient from top, and cyanobacteria present in the water column, show clear evidence for the formation of Fe(III) mineral precipitates and development of a chemocline between Fe(II) and O2. This column allows us to test hypotheses for the formation of the BIFs by culturing cyanobacteria (and in the future photoferrotrophs) under simulated marine Precambrian conditions. Furthermore we hypothesize that our column concept allows for the simulation of various chemical and physical environments &#8212; including shallow marine or lacustrine sediments.

Laboratory Simulation of an IronII-rich Precambrian Marine Upwelling System to Explore the Growth of Photosynthetic Bacteria

We simulated a Precambrian ferruginous marine upwelling system in a lab-scale vertical flow-through column. The goal was to understand how geochemical profiles of O2 and Fe(II) evolve as cyanobacteria produce O2. The results show the establishment of a chemocline due to Fe(II) oxidation by photosynthetically produced O2.

Simulación de un laboratorio de hierro (II) ricos sistema de afloramiento Precámbrico marina para explorar el crecimiento de bacterias fotosintéticas

A conventional concept for the deposition of some Precambrian Banded Iron Formations (BIF) proceeds on the assumption that ferrous iron [Fe(II)] upwelling ...

Development of Metarhizium anisopliae as a Mycoinsecticide: From Isolation to Field Performance

A major concern when developing commercial mycoinsecticides is the kill speed compared to that of chemical insecticides. Therefore, isolation and screening for the selection of a fast-acting, highly virulent entomopathogenic fungus are important steps. Entomopathogenic fungi, such as Metarhizium, Beauveria, and Nomurea, which act by contact, are better suited than Bacillus thuringiensis or nucleopolyhedrosis virus (NPV), which must be ingested by the insect pest. In the present work, we isolated 68 Metarhizium strains from infected insects using a soil dilution and bait method. The isolates were identified by the amplification and sequencing of the ITS1-5.8S-ITS2 and 26S rDNA region. The most virulent strain of Metarhizium anisopliae was selected based on the median lethal concentration (LC50) and time (LT50) obtained in insect bioassays against III-instar larvae of Helicoverpa armigera. The mass production of spores by the selected strain was carried out with solid-state fermentation (SSF) using rice as a substrate for 14 days. Spores were extracted from the sporulated biomass using 0.1% tween-80, and different formulations of the spores were prepared. Field trials of the formulations for the control of an H. armigera infestation in pigeon peas were carried out by randomized block design. The infestation control levels obtained with oil and aqueous formulations (78.0% and 70.9%, respectively) were better than the 63.4% obtained with chemical pesticide.

Development of Metarhizium anisopliae as a Mycoinsecticide: From Isolation to Field Performance

Here, we report the different stages involved in the knowledge-based development of an effective mycoinsecticide, including the isolation, identification, screening, and selection of the "best-fit" entomopathogenic fungus, Metarhizium anisopliae, for the control of insect pests in agriculture.

Desarrollo de<em&gt; Metarhizium anisopliae</em&gt; Como Mycoinsecticide: Del aislamiento al funcionamiento del campo

A major concern when developing commercial mycoinsecticides is the kill speed compared to that of chemical insecticides. Therefore, isolation and ...

Procedures of Laboratory Fumigation for Pest Control with Nitric Oxide Gas

Nitric oxide (NO) is a newly discovered fumigant for postharvest pest control. This paper provides detailed protocols for conducting NO fumigation on fresh products and procedures for residue analysis and product quality evaluation. An airtight fumigation chamber containing fresh fruit and vegetables is first flushed with nitrogen (N2) to establish an ultralow oxygen (ULO) environment followed by injection of NO. The fumigation chamber is then kept at a low temperature of 2 - 5 &#176;C for a specified time period necessary to kill a target pest to complete a fumigation treatment. At the end of a fumigation treatment, the fumigation chamber is flushed with N2 to dilute NO prior to opening the chamber to ambient air to prevent the reaction between NO and O2, which produces NO2 and may damage delicate fresh products. At different times after NO fumigation, NO2 in headspace and nitrate and nitrite in liquid samples were measured as residues. Product quality was evaluated after 2 weeks of post-treatment cold storage to determine effects of NO fumigation on product quality. Keeping&#160;O2 from reacting with NO is critical to NO fumigation and is an important part of the protocols. Measuring NO levels is challenging and a practical solution is provided. Possible protocol modifications are also suggested for measuring NO levels in the fumigation chambers as well as residues. NO fumigation has the potential to be a practical alternative to methyl bromide fumigation for postharvest pest control on fresh and stored products. This publication is intended to assist other researchers in conducting NO fumigation research for postharvest pest control and accelerating the development of NO fumigation for practical applications.

This paper describes nitric oxide (NO) fumigation protocols for postharvest pest control. Fumigation chambers are flushed with nitrogen (N2) to establish ultralow oxygen conditions before NO is injected. At the end, chambers are flushed with N2 to dilute NO before exposing products to ambient air to prevent exposure to NO2.

Procedimientos de laboratorio fumigación Control de plagas con Gas de óxido nítrico

Nitric oxide (NO) is a newly discovered fumigant for postharvest pest control. This paper provides detailed protocols for conducting NO fumigation on ...

Adherence of Bacteria to Plant Surfaces Measured in the Laboratory

This manuscript describes a method to measure bacterial binding to axenic plant surfaces in the light microscope and through the use of viable cell counts. Plant materials used include roots, sprouts, leaves, and cut fruits. The methods described are inexpensive, easy, and suitable for small sample sizes. Binding is measured in the laboratory and a variety of incubation media and conditions can be used. The effect of inhibitors can be determined. Situations that promote and inhibit binding can also be assessed. In some cases it is possible to distinguish whether various conditions alter binding primarily due to their effects on the plant or on the bacteria.

An easy method for measuring and characterizing bacterial adhesion to plants, particularly roots and sprouts, is described in this article.

Adherencia de bacterias a las superficies de midieron en el laboratorio de la planta

This manuscript describes a method to measure bacterial binding to axenic plant surfaces in the light microscope and through the use of viable cell ...

Ecosystem Fabrication (EcoFAB) Protocols for The Construction of Laboratory Ecosystems Designed to Study Plant-microbe Interactions

Beneficial plant-microbe interactions offer a sustainable biological solution with the potential to boost low-input food and bioenergy production. A better mechanistic understanding of these complex plant-microbe interactions will be crucial to improving plant production as well as performing basic ecological studies investigating plant-soil-microbe interactions. Here, a detailed description for ecosystem fabrication is presented, using widely available 3D printing technologies, to create controlled laboratory habitats (EcoFABs) for mechanistic studies of plant-microbe interactions within specific environmental conditions. Two sizes of EcoFABs are described that are suited for the investigation of microbial interactions with various plant species, including Arabidopsis thaliana, Brachypodium distachyon, and Panicum virgatum. These flow-through devices allow for controlled manipulation and sampling of root microbiomes, root chemistry as well as imaging of root morphology and microbial localization. This protocol includes the details for maintaining sterile conditions inside EcoFABs and mounting independent LED light systems onto EcoFABs. Detailed methods for addition of different forms of media, including soils, sand, and liquid growth media coupled to the characterization of these systems using imaging and metabolomics are described. Together, these systems enable dynamic and detailed investigation of plant and plant-microbial consortia including the manipulation of microbiome composition (including mutants), the monitoring of plant growth, root morphology, exudate composition, and microbial localization under controlled environmental conditions. We anticipate that these detailed protocols will serve as an important starting point for other researchers, ideally helping create standardized experimental systems for investigating plant-microbe interactions.

Ecosystem Fabrication EcoFAB Protocols for The Construction of Laboratory Ecosystems Designed to Study Plant-microbe Interactions

This article describes detailed protocols for ecosystem fabrication of devices (EcoFABs) that enable the studies of plants and plant-microbe interactions in highly controlled laboratory conditions.

Protocolos de fabricación (EcoFAB) del ecosistema para la construcción de ecosistemas de laboratorio diseñado para estudiar las interacciones planta-microorganismo

Beneficial plant-microbe interactions offer a sustainable biological solution with the potential to boost low-input food and bioenergy production. A ...

Laboratory and Field Protocol for Estimating Sheet Erosion Rates from Dendrogeomorphology

Sheet erosion is among the crucial drivers of soil degradation. Erosion is controlled by environmental factors and human activities, which often lead to severe environmental impacts. The understanding of sheet erosion is, consequently, a worldwide issue with implications for both environment and economies. However, the knowledge on how erosion evolves in space and time is still limited, as well as its effects on the environment. Below, we explain a new dendrogeomorphological protocol for deriving eroded soil thickness (Ex) by acquiring accurate microtopographic data using both terrestrial laser scanning (TLS) and microtopographic profile gauges. Additionally, standard dendrogeomorphic procedures, dependent on anatomical variations in root rings, are utilized to establish the timing of exposure. Both TLS and microtopographic profile gauges are used to obtain ground surface pro&#64257;les, from which Ex is estimated after the threshold distance (TD) is determined, i.e., the distance between the root and the sediment knickpoint, which allows de&#64257;ning the lowering of the ground surface caused by sheet erosion. For each profile, we measured the height between the topside of the root and a virtual plane tangential to the ground surface. In this way, we intended to avoid small-scale impacts of soil deformation, which may be due to pressures exerted by the root system, or by the arrangement of exposed roots. This may provoke small amounts of soil sedimentation or erosion depending on how they physically affect the surface runoff. We demonstrate that an adequate microtopographic characterization of exposed roots and their associated ground surface is very valuable to obtain accurate erosion rates. This finding could be utilized to develop the best management practices designed to eventually halt or perhaps, at least, lessen soil erosion, so that more sustainable management policies can be put into practice.

Characterizing erosion from dendrogeomorphology has usually focused on accurately finding the starting time of root exposure, by examining macroscopic or cell level changes caused by exposure. Here, we offer a detailed description of different novel techniques to obtain more precise erosion rates from highly accurate microtopographic data.

Laboratorio y el protocolo de campo para estimar tasas de erosión de la hoja de Dendrogeomorphology

Sheet erosion is among the crucial drivers of soil degradation. Erosion is controlled by environmental factors and human activities, which often lead to ...