The authors would like to acknowledge funding from the National Science Foundation (award # 1335550), Utah Water Research Laboratory, Professor Joan McLean and Tessa Guy for their help during the metal/metalloids analysis. The authors also thank Laura Birkhold for her support with the data collection and Danna Olbright.

<ol>
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The authors declare that they have no competing financial interests.

Saline Mikroalgen N. salina erfolgreich im Wachstums gestaltet System reproduzierbare Ergebnisse und hohe Biomasseerträge angebaut werden. Airlift Mischen für einen gut gemischten suspendierten Kultur mit minimaler Absetzen oder Biofouling auf der 7-tägigen Wachstumsperioden erlaubt. Die minimale Licht Variabilität über die Fluoreszenzlicht Bank wird auch gezeigt, dass deutliche Unterschiede in Wachstum nicht zu produzieren. Die Studie zeigt, Schwermetall verschmutzten Medien i...

Im Vergleich zu herkömmlichen terrestrischen Kulturpflanzen Mikroalgen sind gezeigt worden, um höhere Biomasse und Lipid Ausbeuten aufgrund inhärenter höherer Solarumwandlungseffizienzen 1,2 erzielen. Kultivierung von Mikroalgen bei hohen Produktivitätsraten erfordert die Versorgung mit verschiedenen Nährstoffen, einschließlich einer externen Kohlenstoffquelle. Es wird erwartet, dass großflächige Wachstum Einrichtungen werden mit industriellen Abfallströme wie industriellen Abgas um die Produktionskosten zu minimieren integrieren und gleichzeitig bieten Umweltsanierung. Industrieabfälle Kohlenstoff ist in der Regel in Form von gasförmigem Kohlendioxid und Schadstoffe, die das Potenzial, sich negativ auf die Produktion von Mikroalgen haben, enthalten. Konkret wird Rauchgas aus Kohle eine Vielzahl von Verunreinigungen aufweisen, einschließlich aber nicht beschränkt auf Verbrennungsprodukte Wasser und Kohlendioxid, sowie Oxide von Schwefel und Stickstoff, Feinstaub, organische Verunreinigungen, wie Dioxine und Furane sowie anorganische con begrenztSchadstoffe wie Schwermetalle. Die Auswirkungen der meisten dieser Verunreinigungen, einschließlich anorganische Stoffe mit einigen von ihnen, wie Schwermetalle auf Mikroalgen Produktivität bekannt wurden nicht untersucht worden. Einige dieser Elemente können Nährstoffe in geeigneten Konzentrationen sein, aber bei höheren Konzentrationen sie Zelldysfunktion und sogar Tod 3 produzieren kann. Die Integration von Mikroalgen mit industriellen Abgas hat das Potential, anorganischen Verunreinigungen in das Wachstumsmedium direkt einzuführen. Kohlebasis Abgas hat eine Vielzahl von anorganischen Elementen (beispielsweise As, Cd, Co, Cr, Cu, Hg, Mn, Ni, Pb, Sb, Se, Sn, V und Zn) in verschiedenen Konzentrationen von denen einige, in niedrigen Konzentration, stellen Nährstoffe für Mikroalgenwachstum. Anorganischen Verunreinigungen haben eine hohe Affinität zu den Mikroalgen zu binden und weiter intern durch Nährstofftransport sorbiert werden. Einige anorganische Verunreinigungen (dh, Co, Cu, Zn und Mn) sind Nährstoffe, die Bestandteil von Enzymen bilden beinhaltend in der Photosynthese, Atmung und andere Funktionen 3,4. Jedoch im Überschuß von Metallen und Metalloiden können toxisch sein. Andere Elemente, wie Pb, Cd, Sn, Sb, Se, As und Hg, sind nicht bekannt, die Zellfunktion in jeder Konzentration unterstützt und stellen nicht-Nährstoff Metalle, die sich negativ auswirken könnte Kulturwachstum 3,5,6. Das Vorhandensein von jeder dieser Verunreinigungen hat das Potenzial, negative Auswirkungen auf die Mikroalgenzellfunktion zu erzeugen. Weiterhin ist die Wechselwirkung von mehreren Metallen mit Mikroalgen verkompliziert Wachstumsdynamik und das Potential hat, das Wachstum auswirken. Großökonomie wurden direkt auf die Produktivität des Kultivierungssystem 7-19 verbunden. Außerdem ist Medium Rückführung in der Mikroalge Wachstumssystem für entweder offen Laufbahn Teichen (ORP) oder Photobioreaktoren (PBR) kritisch, da sie repräsentiert 99,9 und 99,4% der Masse, bzw. 20. Die Anwesenheit von anorganischen Verunreinigungen in den Medien könnte letztlich begrenzen microalgae Produktivität und das Recycling von Medien durch Verunreinigung Aufbau. Diese Studie experimentell bestimmt die Auswirkungen 14 anorganischen Verunreinigungen (As, Cd, Co, Cr, Cu, Hg, Mn, Ni, Pb, Sb, Se, Sn, V und Zn) in Konzentrationen von der Integration von Mikroalgen Kultivierungssysteme erwartet Kohle abgeleiteten Rauchgas, auf die Produktivität der N. salina in Luftbrücke PBRs gewachsen. Die in dieser Studie verwendeten Verunreinigungen haben gezeigt, dass nicht nur in Steinkohle gewonnene Rauchgas aber Siedlungsabfall-basierte Abgas, biosolids-basierte Abgas, kommunales Abwasser, erzeugt Wasser, beeinträchtigt das Grundwasser und Meerwasser 21-23 vorliegen. Die in dieser Studie verwendeten Konzentrationen sind auf erwarten wäre, wenn Mikroalgenwachstumssysteme wurden mit einem Kohlebasis CO 2 -Quelle mit einer Aufnahmeeffizienz in kommerziellen PBR Systeme 20 gezeigt, integriert werden kann. Detaillierte Berechnungen unterstützen die Konzentration der Schwermetalle und anorganische Verunreinigungen werden in Napan vorgestelltet al. 24 Analytische Techniken wurden verwendet, um die Verteilung der meisten Metalle in den Biomasse, Medien und Umwelt zu verstehen. Die vorgestellten Methoden ermöglichte die Beurteilung der Produktivitätspotenzial von Mikroalgen unter anorganische Kontaminanten Stress und Quantifizierung ihrer Ende Schicksal.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td colspan="4">Chemicals</td>
		</tr>
		<tr>
			<td>Sodium chloride</td>
			<td>Fisher Scientific</td>
			<td>S271-3</td>
			<td></td>
		</tr>
		<tr>
			<td>Calcium chloride dihydrate</td>
			<td>Fisher Scientific</td>
			<td>C79-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium chloride</td>
			<td>Fisher Scientific</td>
			<td>P217-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium meta silicate nonahydrate&#160;</td>
			<td>Fisher Scientific</td>
			<td>S408-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Magnesium sulfate heptahydrate&#160;</td>
			<td>Fisher Scientific</td>
			<td>M63-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium nitrate</td>
			<td>EMD Chemical</td>
			<td>PX1520-5</td>
			<td></td>
		</tr>
		<tr>
			<td>Potassium phosphate monobasic&#160;</td>
			<td>Fisher Scientific</td>
			<td>P285-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Ammonium ferric citrate</td>
			<td>Fisher Scientific</td>
			<td>I72-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Boric acid</td>
			<td>Fisher Scientific</td>
			<td>A73-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium molybdate, dihydrate</td>
			<td>EMD Chemical</td>
			<td>SX0650-2</td>
			<td></td>
		</tr>
		<tr>
			<td>Manganese chloride tetrahydrate</td>
			<td>Fisher Scientific</td>
			<td>M87-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Zinc sulfate heptahydrate</td>
			<td>Fisher Scientific</td>
			<td>Z68-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Cupric sulfate pentahydrate</td>
			<td>Fisher Scientific</td>
			<td>C489-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Biotin&#160;</td>
			<td>Acros Organics</td>
			<td>230090010</td>
			<td></td>
		</tr>
		<tr>
			<td>Thiamine&#160;</td>
			<td>Acros Organics</td>
			<td>148990100</td>
			<td></td>
		</tr>
		<tr>
			<td>Vitamin B12&#160;</td>
			<td>Acros Organics</td>
			<td>405920010</td>
			<td></td>
		</tr>
		<tr>
			<td>Copper (II) chloride dihydrate&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>221783-100G</td>
			<td>Irritant, Dangerous to the Environment</td>
		</tr>
		<tr>
			<td>Lead (II) chloride&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>268690-250G</td>
			<td>Toxic, Dangerous to the Environment</td>
		</tr>
		<tr>
			<td>Sodium dichromate dihydrate&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>398063-100G</td>
			<td>Oxidizing, Highly Toxic, Dangerous to the Environment</td>
		</tr>
		<tr>
			<td>Cobalt (II) chloride hexahydrate&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>255599-100G</td>
			<td>Toxic, Dangerous to the Environment</td>
		</tr>
		<tr>
			<td>Nickel (II) chloride hexahydrate&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>223387-500G</td>
			<td>Toxic, Dangerous to the Environment</td>
		</tr>
		<tr>
			<td>Sodium (meta) arsenite&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>71287</td>
			<td>Toxic, Dangerous to the Environment</td>
		</tr>
		<tr>
			<td>Cadmium chloride&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>202908-10G</td>
			<td>Highly Toxic, Dangerous to the Environment</td>
		</tr>
		<tr>
			<td>Mercury (II) chloride&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>215465-100G</td>
			<td>Toxic, Dangerous to the Environment</td>
		</tr>
		<tr>
			<td>Tin (II) chloride dihydrate</td>
			<td>Fisher Scientific</td>
			<td>T142-500</td>
			<td>Corrosive. Suitable for Hg analysis. Very hazardous.</td>
		</tr>
		<tr>
			<td>Manganese chloride tetrahydrate</td>
			<td>Fisher Scientific</td>
			<td>M87-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Vanadium (V) oxide</td>
			<td>Acros Organics</td>
			<td>206422500</td>
			<td>Dangerous to the Environment</td>
		</tr>
		<tr>
			<td>Carbon dioxide&#160;</td>
			<td>Air Liquide</td>
			<td>I2301S-1</td>
			<td>Compressed</td>
		</tr>
		<tr>
			<td>Hydrogen peroxide</td>
			<td>H325-500</td>
			<td>Fisher Scientific</td>
			<td>30% in water</td>
		</tr>
		<tr>
			<td>ICP-MS standard</td>
			<td>ICP-MS-6020</td>
			<td>High Purity Standards</td>
			<td></td>
		</tr>
		<tr>
			<td>Mercury standard</td>
			<td>CGHG1-1</td>
			<td>Inorganic Ventures</td>
			<td>1000&#177;6 &#181;g/mL in 5% nitric acid</td>
		</tr>
		<tr>
			<td>Argon</td>
			<td>Air Liquide</td>
			<td></td>
			<td>Compressed</td>
		</tr>
		<tr>
			<td>Helium</td>
			<td>Air Liquide</td>
			<td></td>
			<td>Compressed, ultra high purity</td>
		</tr>
		<tr>
			<td>Hydrogen</td>
			<td>Air Liquide</td>
			<td></td>
			<td>Compressed, ultra high purity</td>
		</tr>
		<tr>
			<td>Nitric acid</td>
			<td>Fisher Scientific</td>
			<td>A509-P212</td>
			<td>67-70% nitric acid, trace metal grade. Caution: manipulate under fume hood.</td>
		</tr>
		<tr>
			<td>Hydrochloric acid</td>
			<td>Fisher Scientific</td>
			<td>A508-P212</td>
			<td>35% hydrochloric acid, trace metal grade. Caution: manipulate under fume hood.</td>
		</tr>
		<tr>
			<td colspan="4">Equipment</td>
		</tr>
		<tr>
			<td>Scientific prevacuum sterilizer</td>
			<td>Steris</td>
			<td>31626A</td>
			<td>SV-120</td>
		</tr>
		<tr>
			<td>Centrifuge</td>
			<td>Thermo Fisher</td>
			<td>46910</td>
			<td>RC-6 Plus</td>
		</tr>
		<tr>
			<td>Spectrophotometer</td>
			<td>Shimadzu</td>
			<td>1867</td>
			<td>UV-1800</td>
		</tr>
		<tr>
			<td>pH controller</td>
			<td>Hanna</td>
			<td>BL981411</td>
			<td>X4</td>
		</tr>
		<tr>
			<td>Rotometer, X5</td>
			<td>Dwyer</td>
			<td>RMA-151-SSV</td>
			<td>T31Y</td>
		</tr>
		<tr>
			<td>Rotometer, X5</td>
			<td>Dwyer</td>
			<td>RMA-26-SSV</td>
			<td>T35Y</td>
		</tr>
		<tr>
			<td>Water bath circulator</td>
			<td>Fisher Scientific</td>
			<td>13-873-45A</td>
			<td></td>
		</tr>
		<tr>
			<td>Compact chiller</td>
			<td>VWR</td>
			<td>13270-120</td>
			<td></td>
		</tr>
		<tr>
			<td>Freeze dryer</td>
			<td>Labconco</td>
			<td>7752020</td>
			<td></td>
		</tr>
		<tr>
			<td>Stir plate</td>
			<td>Fisher Scientific</td>
			<td>11-100-49S</td>
			<td></td>
		</tr>
		<tr>
			<td>pH lab electrode</td>
			<td>Phidgets Inc</td>
			<td>3550</td>
			<td></td>
		</tr>
		<tr>
			<td>Inductively coupled plasma mass spectrometer</td>
			<td>Agilent Technologies</td>
			<td>7700 Series ICP-MS</td>
			<td>Attached to autosampler CETAC ASX-520</td>
		</tr>
		<tr>
			<td>FIAS 100</td>
			<td>Perkin Elmer Instruments</td>
			<td>B0506520</td>
			<td></td>
		</tr>
		<tr>
			<td>Atomic absorption spectrometer</td>
			<td>Perkin Elmer Instruments</td>
			<td>AAnalyst 800</td>
			<td></td>
		</tr>
		<tr>
			<td>Cell heater (quartz)</td>
			<td>Perkin Elmer Instruments</td>
			<td>B3120397</td>
			<td></td>
		</tr>
		<tr>
			<td>Microwave</td>
			<td>Milestone</td>
			<td></td>
			<td>Programmable, maximum power 1200 W</td>
		</tr>
		<tr>
			<td>Microwave rotor</td>
			<td>Milestone</td>
			<td></td>
			<td>Rotor with 24 75 mL Teflon vessels for closed-vessel microwave assisted digestion.</td>
		</tr>
		<tr>
			<td colspan="4">Materials</td>
		</tr>
		<tr>
			<td>0.2 micron syringe filter</td>
			<td>Whatman</td>
			<td>6713-0425</td>
			<td></td>
		</tr>
		<tr>
			<td>0.2 micron syringe filter</td>
			<td>Whatman</td>
			<td>6713-1650</td>
			<td></td>
		</tr>
		<tr>
			<td>0.45 micron syringe filter</td>
			<td>Thermo Fisher</td>
			<td>F2500-3</td>
			<td></td>
		</tr>
		<tr>
			<td>Polystyrene tubes</td>
			<td>Evergreen</td>
			<td>222-2094-050</td>
			<td>17x100 mm w/cap, 16 mL, polysteryne</td>
		</tr>
		<tr>
			<td>Octogonal magnetic&#160;stir&#160;bars</td>
			<td>Fisher scientific</td>
			<td>14-513-60</td>
			<td>Magnets encased in PTFE fluoropolymer</td>
		</tr>
	</tbody>
</table>

quantification of heavy metals and other inorganic contaminants on the productivity of microalgae

Die zunehmende Nachfrage nach erneuerbaren Kraftstoffen hat Forscher die Machbarkeit alternativer Einsatzmaterialien, wie beispielsweise Mikroalgen. Inhärenten Vorteile sind die hohe Ertragspotenzial, Verwendung von Nicht-Ackerland und die Integration mit Abfallströmen. Die Nährstoffbedarf eines großen Mikroalgen-Produktionssystem wird die Kopplung von Anbausystemen mit Industrieabfällen Ressourcen wie Kohlendioxid aus Rauchgas und Nährstoffe aus dem Abwasser erforderlich. In diesen Abfällen vorhanden anorganischen Verunreinigungen kann potenziell zu Bioakkumulation in Mikroalgen-Biomasse führen, sich negativ auf die Produktivität und die Begrenzung der Endanwendung. Diese Studie konzentriert sich auf die experimentelle Evaluierung der Auswirkungen und das Schicksal von 14 anorganischen Verunreinigungen (As, Cd, Co, Cr, Cu, Hg, Mn, Ni, Pb, Sb, Se, Sn, V und Zn) auf Nannochloropsis salina Wachstum . Mikroalgen in Photobioreaktoren bei 984 &amp; mgr; mol m -2 s belichtet bei pH 7 in einem Wachstums m -1 kultiviert und gepflegtEdia verschmutzt mit anorganischer Verunreinigungen in Konzentrationen erwartet, basierend auf der Zusammensetzung in kommerziellen Kohlenabgasanlagen gefunden. In der Biomasse und dem Medium am Ende einer 7-tägigen Wachstumsperiode vorhandenen Verunreinigungen wurden analytisch durch Kaltdampf-Atomabsorptionsspektrometrie für Hg und durch induktiv gekoppelte Plasma-Massenspektrometrie für As, Cd, Co, Cr, Cu, Mn, Ni quantifiziert, Pb, Sb, Se, Sn, V und Zn. Die Ergebnisse zeigen, N. salina ist ein empfindlicher Stamm, der zur Mehrmetall-Umgebung mit einer statistischen Abnahme der Biomasse yieldwith der Einführung dieser Verunreinigungen. Die hier vorgestellten Techniken sind ausreichend für die Quantifizierung von Algenwachstum und Bestimmung des Schicksals von anorganischen Verunreinigungen.

Biomasseerträge Herstellung von N. salina in der in dieser Studie verwendeten PBR Systems stieg von 1 g / l -1 auf 8,5 ± 0,19 g / L -1 (N = 12) für die Steuerung Reaktoren und 4,0 ± 0,3 g / L -1 (N = 12) für den in 7 Tagen Multimetall kontaminiert. Die Experimente erzeugt wiederholbare Daten in dreifacher Ausfertigung Reaktoren und mehrere Stapel auf. 2A zeigt die durchschnittlichen Kulturdichte mit sehr geringen Standardfehler ...

Watch this Scientific Journal Video about Quantification of Heavy Metals and Other Inorganic Contaminants on the Productivity of Microalgae at JoVE.com

Quantification of Heavy Metals and Other Inorganic Contaminants on the Productivity of Microalgae

Integration of microalgal cultivation with industrial flue gas will ultimately introduce heavy metals and other inorganic compounds into the growth media. This study presents a procedure used to determine the end fate and impact of heavy metals and inorganic contaminants on the growth of Nannochloropsis salina grown in photobioreactors.

Quantifizierung von Schwermetallen und anderen anorganischen Kontaminanten auf die Produktivität von Mikroalgen

Increasing demand for renewable fuels has researchers investigating the feasibility of alternative feedstocks, such as microalgae. Inherent advantages ...

quantification-heavy-metals-other-inorganic-contaminants-on

Research

JoVE Journal

Environment

15.8K Aufrufe.  Utah State University. Integration of microalgal cultivation with industrial flue gas will ultimately introduce heavy metals and other inorganic compounds into the growth media. This study presents a procedure used to determine the end fate and impact of heavy metals and inorganic contaminants on the growth of Nannochloropsis salina grown in photobioreactors.

Serie: Quantification of Heavy Metals and Other Inorganic Contaminants on the Productivity of Microalgae

Analysis of Fatty Acid Content and Composition in Microalgae

A method to determine the content and composition of total fatty acids present in microalgae is described. Fatty acids are a major constituent of microalgal biomass. These fatty acids can be present in different acyl-lipid classes. Especially the fatty acids present in triacylglycerol (TAG) are of commercial interest, because they can be used for production of transportation fuels, bulk chemicals, nutraceuticals (&omega;-3 fatty acids), and food commodities. To develop commercial applications, reliable analytical methods for quantification of fatty acid content and composition are needed. Microalgae are single cells surrounded by a rigid cell wall. A fatty acid analysis method should provide sufficient cell disruption to liberate all acyl lipids and the extraction procedure used should be able to extract all acyl lipid classes.
With the method presented here all fatty acids present in microalgae can be accurately and reproducibly identified and quantified using small amounts of sample (5 mg) independent of their chain length, degree of unsaturation, or the lipid class they are part of.
This method does not provide information about the relative abundance of different lipid classes, but can be extended to separate lipid classes from each other.
The method is based on a sequence of mechanical cell disruption, solvent based lipid extraction, transesterification of fatty acids to fatty acid methyl esters (FAMEs), and quantification and identification of FAMEs using gas chromatography (GC-FID). A TAG internal standard (tripentadecanoin) is added prior to the analytical procedure to correct for losses during extraction and incomplete transesterification.

A method for the determination of fatty acid content and composition in microalgae based on mechanical cell disruption, solvent based lipid extraction, transesterification, and quantification and identification of fatty acids using gas chromatography is described. A tripentadecanoin internal standard is used to compensate for the possible losses during extraction and incomplete transesterification.

Die Analyse der Fettsäuregehalt und Zusammensetzung in Mikroalgen

A method to determine the content and composition of total fatty acids present in microalgae is described. Fatty acids are a major constituent of ...

Forschung

Umwelt

A Whole Cell Bioreporter Approach to Assess Transport and Bioavailability of Organic Contaminants in Water Unsaturated Systems

Bioavailability of contaminants is a prerequisite for their effective biodegradation in soil. The average bulk concentration of a contaminant, however, is not an appropriate measure for its availability; bioavailability rather depends on the dynamic interplay of potential mass transfer (flux) of a compound to a microbial cell and the capacity of the latter to degrade the compound. In water-unsaturated parts of the soil, mycelia have been shown to overcome bioavailability limitations by actively transporting and mobilizing organic compounds over the range of centimeters. Whereas the extent of mycelia-based transport can be quantified easily by chemical means, verification of the contaminant-bioavailability to bacterial cells requires a biological method. Addressing this constraint, we chose the PAH fluorene (FLU) as a model compound and developed a water unsaturated model microcosm linking a spatially separated FLU point source and the FLU degrading bioreporter bacterium Burkholderia sartisoli RP037-mChe by a mycelial network of Pythium ultimum. Since the bioreporter expresses eGFP in response of the PAH flux to the cell, bacterial FLU exposure and degradation could be monitored directly in the microcosms via confocal laser scanning microscopy (CLSM). CLSM and image analyses revealed a significant increase of the eGFP expression in the presence of P. ultimum compared to controls without mycelia or FLU thus indicating FLU bioavailability to bacteria after mycelia-mediated transport. CLSM results were supported by chemical analyses in identical microcosms. The developed microcosm proved suitable to investigate contaminant bioavailability and to concomitantly visualize the involved bacteria-mycelial interactions.

A whole cell bioreporter assay with Burkholderia sartisoli RP037-mChe was developed to detect fractions of an organic contaminant (i.e., fluorene) available for bacterial degradation after active transport by mycelia bridging air-filled pores in a water unsaturated model system.

Eine ganze Zelle Bioreporter Ansatz für Transport und Bioverfügbarkeit von organischen Schadstoffen in Wasser Ungesättigte Systeme Bewerten

Bioavailability of contaminants is a prerequisite for their effective biodegradation in soil. The average bulk concentration of a contaminant, however, is ...

Assaying Predatory Feeding Behaviors in Pristionchus and Other Nematodes

This protocol provides multiple methods for the analysis and quantification of predatory feeding behaviors in nematodes. Many nematode species including Pristionchus pacificus display complex behaviors, the most striking of which is the predation of other nematode larvae. However, as these behaviors are absent in the model organism Caenorhabditis elegans, they have thus far only recently been described in detail along with the development of reliable behavioral assays 1. These predatory behaviors are dependent upon phenotypically plastic but fixed mouth morphs making the correct identification and categorization of these animals essential. In P. pacificus there are two mouth types, the stenostomatous and eurystomatous morphs 2, with only the wide mouthed eurystomatous containing an extra tooth and being capable of killing other nematode larvae. Through the isolation of an abundance of size matched prey larvae and subsequent exposure to predatory nematodes, assays including both "corpse assays" and "bite assays" on correctly identified mouth morph nematodes are possible. These assays provide a means to rapidly quantify predation success rates and provide a detailed behavioral analysis of individual nematodes engaged in predatory feeding activities. In addition, with the use of a high-speed camera, visualization of changes in pharyngeal activity including tooth and pumping dynamics are also possible.

Assaying Predatory Feeding Behaviors in Pristionchus and Other Nematodes

Behavioral assays provide powerful tools for understanding neuronal function. Here we present several protocols for quantifying predatory feeding behavior found in the model nematode Pristionchus pacificus and its relatives. Additionally, we provide methods for analyzing predatory feeding adaptations including mouth structures and teeth.

Die Untersuchung Räuberische Fütterung Behaviors in<em&gt; Pristionchus</em&gt; Und andere Nematoden

This protocol provides multiple methods for the analysis and quantification of predatory feeding behaviors in nematodes. Many nematode species including ...

Ecotoxicological Method with Marine Bacteria Vibrio anguillarum to Evaluate the Acute Toxicity of Environmental Contaminants

Bacteria are an important component of the ecosystem, and microbial community alterations can have a significant effect on biogeochemical cycling and food webs. Toxicity testing based on microorganisms are widely used because they are relatively quick, reproducible, cheap, and are not associated with ethical issues. Here, we describe an ecotoxicological method to evaluate the biological response of the marine bacterium Vibrio anguillarum. This method assesses the acute toxicity of chemical compounds, including new contaminants such as nanoparticles, as well as environmental samples. The endpoint is the reduction of bacterial culturability (i.e., the capability to replicate and form colonies) due to exposure to a toxicant. This reduction can be generally referred to as mortality. The test allows for the determination of the LC50, the concentration that causes a 50% decrease of bacteria actively replicating and forming colonies, after a 6 h exposure. The culturable bacteria are counted in terms of colony forming units (CFU), and the &#34;mortality&#34; is evaluated and compared to the control. In this work, the toxicity of copper sulphate (CuSO4) was evaluated. A clear dose-response relationship was observed, with a mean LC50 of 1.13 mg/L, after three independent tests. This protocol, compared to existing methods with microorganisms, is applicable in a wider range of salinity and has no limitations for colored/turbid samples. It uses saline solution as the exposure medium, avoiding any possible interferences of growth medium with the investigated contaminants. The LC50 calculation facilitates comparisons with other bioassays commonly applied to ecotoxicological assessments of the marine environment.

Ecotoxicological Method with Marine Bacteria Vibrio anguillarum to Evaluate the Acute Toxicity of Environmental Contaminants

This work describes a new protocol to assess the ecotoxicity of pollutants, including emerging contaminants such as nanomaterials, using the marine bacterium Vibrio anguillarum. This method allows for the determination of the LC50, or mortality, the concentration that causes a 50% decrease in bacteria culturability, after a 6 h exposure.

Ökotoxikologische Methode mit Meeresbakterien<em&gt; Vibrio anguillarum</em&gt; Zur Beurteilung der akuten Toxizität von Umweltschadstoffen

Bacteria are an important component of the ecosystem, and microbial community alterations can have a significant effect on biogeochemical cycling and food ...

Preparation of DMMTAV and DMDTAV Using DMAV for Environmental Applications: Synthesis, Purification, and Confirmation

Dimethylated thioarsenicals such as dimethylmonothioarsinic acid (DMMTAV) and dimethyldithioarsinic acid (DMDTAV), which are produced by the metabolic pathway of dimethylarsinic acid (DMAV) thiolation, have been recently found in the environment as well as human organs. DMMTAV and DMDTAV can be quantified to determine the ecological effects of dimethylated thioarsenicals and their stability in environmental media. The synthesis method for these compounds is unstandardized, making replicating previous studies challenging. Furthermore, there is a lack of information about storage techniques, including storage of compounds without species transformation. Moreover, because only limited information about synthesis methods is available, there may be experimental difficulties in synthesizing standard chemicals and performing quantitative analysis. The protocol presented herein provides a practically modified synthesis method for the dimethylated thioarsenicals, DMMTAV and DMDTAV, and will help in the quantification of species separation analysis using high performance liquid chromatography in conjunction with inductively coupled plasma mass spectrometry (HPLC-ICP-MS). The experimental steps of this procedure were modified by focusing on the preparation of chemical reagents, filtration methods, and storage.

Preparation of DMMTAV and DMDTAV Using DMAV for Environmental Applications: Synthesis, Purification, and Confirmation

This article presents modified experimental protocols for dimethylmonothioarsinic acid (DMMTAV) and dimethyldithioarsinic acid (DMDTAV) synthesis, inducing dimethylarsinic acid (DMAV) thiolation through mixing of DMAV, Na2S, and H2SO4. The modified protocol provides an experimental guideline, thereby overcoming limitations of the synthesis steps that could have caused experimental failures in quantitative analysis.

Vorbereitung des DMMTAV und DMDTAV mit DMAV für Umweltanwendungen: Synthese, Reinigung und Bestätigung

Dimethylated thioarsenicals such as dimethylmonothioarsinic acid (DMMTAV) and dimethyldithioarsinic acid (DMDTAV), which are produced by the metabolic ...

Harvesting Venom Toxins from Assassin Bugs and Other Heteropteran Insects

Heteropteran insects such as assassin bugs (Reduviidae) and giant water bugs (Belostomatidae) descended from a common predaceous and venomous ancestor, and the majority of extant heteropterans retain this trophic strategy. Some heteropterans have transitioned to feeding on vertebrate blood (such as the kissing bugs, Triatominae; and bed bugs, Cimicidae) while others have reverted to feeding on plants (most Pentatomomorpha). However, with the exception of saliva used by kissing bugs to facilitate blood-feeding, little is known about heteropteran venoms compared to the venoms of spiders, scorpions and snakes.
One obstacle to the characterization of heteropteran venom toxins is the structure and function of the venom/labial glands, which are both morphologically complex and perform multiple biological roles (defense, prey capture, and extra-oral digestion). In this article, we describe three methods we have successfully used to collect heteropteran venoms. First, we present electrostimulation as a convenient way to collect venom that is often lethal when injected into prey animals, and which obviates contamination by glandular tissue. Second, we show that gentle harassment of animals is sufficient to produce venom extrusion from the proboscis and/or venom spitting in some groups of heteropterans. Third, we describe methods to harvest venom toxins by dissection of anaesthetized animals to obtain the venom glands. This method is complementary to other methods, as it may allow harvesting of toxins from taxa in which electrostimulation and harassment are ineffective. These protocols will enable researchers to harvest toxins from heteropteran insects for structure-function characterization and possible applications in medicine and agriculture.

Although many insects in the suborder Heteroptera (Insecta: Hemiptera) are venomous, their venom composition and the functions of their venom toxins are mostly unknown. This protocol describes methods to harvest heteropteran venoms for further characterization, using electrostimulation, harassment, and gland dissection.

Ernte Venom Giftstoffe aus Assassin Bugs und andere Heteropteran Insekten

Heteropteran insects such as assassin bugs (Reduviidae) and giant water bugs (Belostomatidae) descended from a common predaceous and venomous ancestor, ...

Evaluation of the Productivity of Social Wasp Colonies (Vespinae) and an Introduction to the Traditional Japanese Vespula Wasp Hunting Technique

For vespine wasps, colony productivity is typically estimated by counting the number of larval cells. This paper presents an improved method that enables researchers to estimate more accurately the number of adults produced, counting the number of meconia (the stools left in the cells by wasp larvae when pupating into adults, per 100 cells) in each comb. This method can be applied before or after colony collapse (i.e., in active or inactive nests). The paper also describes how to locate wild Vespula wasp colonies by &#34;flagging&#34; wasp baits and chasing the wasp collecting them, using a method traditionally performed by local people in central Japan (as illustrated in the associated video). The Vespula chasing method described has several advantages: it is easy to reinitiate the chase from a point where the forager flying back to the nest was lost, and it is easy to pinpoint the nest location as marked wasps often lose their flag at the nest entrance. These methods for estimating colony productivity and collecting nests can be valuable for researchers studying social wasps.

Evaluation of the Productivity of Social Wasp Colonies Vespinae and an Introduction to the Traditional Japanese Vespula Wasp Hunting Technique

This methodological paper evaluates the productivity of a social wasp colony by examining the number of meconia per 100 cells of comb, to estimate the total number of adults the wasps produced. The associated video describes how to search for Vespula wasp nests, a method developed by amateur wasp chasers.

Bewertung der Produktivität sozialer Wespenkolonien (Vespinae) und Einführung in die traditionelle japanische Vespula-Wesp-Jagdtechnik

For vespine wasps, colony productivity is typically estimated by counting the number of larval cells. This paper presents an improved method that enables ...

Cultivation of Green Microalgae in Bubble Column Photobioreactors and an Assay for Neutral Lipids

There is significant interest in the study of microalgae for engineering applications such as the production of biofuels, high value products, and for the treatment of wastes. As most new research efforts begin at laboratory scale, there is a need for cost-effective methods for culturing microalgae in a reproducible manner. Here, we communicate an effective approach to culture microalgae in laboratory-scale photobioreactors, and to measure the growth and neutral lipid content of that algae. Instructions are also included on how to set up the photobioreactor system. Although the example organisms are species of Chlorella and Auxenochlorella, this system can be adapted to cultivate a wide range of microalgae, including co-cultures of algae with non-algae species. Stock cultures are first grown in bottles to produce inoculum for the photobioreactor system. Algae inoculum is concentrated and transferred to photobioreactors for cultivation in batch mode. Samples are collected daily for the optical density readings. At the end of the batch culture, cells are harvested by centrifuge, washed, and freeze dried to obtain a final dry weight concentration. The final dry weight concentration is used to create a correlation between the optical density and the dry weight concentration. A modified Folch method is subsequently used to extract total lipids from the freeze-dried biomass and the extract is assayed for its neutral lipid content using a microplate assay. This assay has been published previously but protocol steps were included here to highlight critical steps in the procedure where errors frequently occur. The bioreactor system described here fills a niche between simple flask cultivation and fully-controlled commercial bioreactors. Even with only 3-4 biological replicates per treatment, our approach to culturing algae leads to tight standard deviations in the growth and lipid assays.

Here, we present a protocol to construct lab-scale bubble column photobioreactors and use them to culture microalgae. It also provides a method for the determination of the culture growth rate and neutral lipid content.

Anbau von grünen Mikroalgen in Bubble Spalte Photobioreaktoren und ein Test für neutrale Lipide

There is significant interest in the study of microalgae for engineering applications such as the production of biofuels, high value products, and for the ...

Microalgae Cultivation and Biomass Quantification in a Bench-Scale Photobioreactor with Corrosive Flue Gases

Photobioreactors are illuminated cultivation systems for experiments on phototrophic microorganisms. These systems provide a sterile environment for microalgal cultivation with temperature, pH, and gas composition and flow rate control. At bench-scale, photobioreactors are advantageous to researchers studying microalgal properties, productivity, and growth optimization. At industrial scales, photobioreactors can maintain product purity and improve production efficiency. The video describes the preparation and use of a bench-scale photobioreactor for microalgal cultivation, including the safe use of corrosive gas inputs, and details relevant biomass measurements and biomass productivity calculations. Specifically, the video illustrates microalgal culture storage and preparation for inoculation, photobioreactor assembly and sterilization, biomass concentration measurements, and a logistic model for microalgal biomass productivity with rate calculations including maximum and overall biomass productivities. Additionally, since there is growing interest in experiments to cultivate microalgae using simulated or real waste gas emissions, the video will cover the photobioreactor equipment adaptations necessary to work with corrosive gases and discuss safe sampling in such scenarios.

Bench-scale, axenic cultivation facilitates microalgal characterization and productivity optimization before subsequent process scale-up. Photobioreactors provide the necessary control for reliable and reproducible microalgal experiments and can be adapted to safely cultivate microalgae with the corrosive gases (CO2, SO2, NO2) from municipal or industrial combustion emissions.

Mikroalgen-Anbau und Biomasse-Quantifizierung in einem Bench-Scale Photobioreaktor mit ätzenden Rauchgasen

Photobioreactors are illuminated cultivation systems for experiments on phototrophic microorganisms. These systems provide a sterile environment for ...

Two-Dimensional Visualization and Quantification of Labile, Inorganic Plant Nutrients and Contaminants in Soil

We describe a method for two-dimensional (2D) visualization and quantification of the distribution of labile (i.e., reversibly adsorbed) inorganic nutrient (e.g., P, Fe, Mn) and contaminant (e.g., As, Cd, Pb) solute species in the soil adjacent to plant roots (the &#8216;rhizosphere&#8217;) at sub-millimeter (~100 &#181;m) spatial resolution. The method combines sink-based solute sampling by the diffusive gradients in thin films (DGT) technique with spatially resolved chemical analysis by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS). The DGT technique is based on thin hydrogels with homogeneously distributed analyte-selective binding phases. The variety of available binding phases allows for the preparation of different DGT gel types following simple gel fabrication procedures. For DGT gel deployment in the rhizosphere, plants are grown in flat, transparent growth containers (rhizotrons), which enable minimal invasive access to a soil-grown root system. After a pre-growth period, DGT gels are applied to selected regions of interest for in situ solute sampling in the rhizosphere. Afterwards, DGT gels are retrieved and prepared for subsequent chemical analysis of the bound solutes using LA-ICP-MS line-scan imaging. Application of internal normalization using 13C and external calibration using matrix-matched gel standards further allows for the quantification of the 2D solute fluxes. This method is unique in its capability to generate quantitative, sub-mm scale 2D images of multi-element solute fluxes in soil-plant environments, exceeding the achievable spatial resolution of other methods for measuring solute gradients in the rhizosphere substantially. We present the application and evaluation of the method for imaging multiple cationic and anionic solute species in the rhizosphere of terrestrial plants and highlight the possibility of combining this method with complementary solute imaging techniques.

This protocol presents a workflow for sub-mm 2D visualization of multiple labile inorganic nutrient and contaminant solute species using diffusive gradients in thin films (DGT) combined with mass spectrometry imaging. Solute sampling and high-resolution chemical analysis are described in detail for quantitative mapping of solutes in the rhizosphere of terrestrial plants.