P23681-B16: Diese Arbeit wurde vom österreichischen Wissenschaftsfonds (FWF) finanziert.

1. Herstellung von Pflanzenmaterialien <ol><li> Verwenden Sie entweder Blätter oder ganze Pflanzen unterliegen Pflanzenmaterial (Hosta spp. Oder Phaseolus vulgaris). </li></ol> 2. Herstellung von Wasserlösungen, die Eisnukleation Aktiv (INA) Bacteria <ol><li> Kultur der INA Bakterium Pseudomonas syringae (Stamm Cit-7) in Petrischalen bei 25 ° C auf Pseudomonas Agar F mit 10 g / l 100% Glycerin gemäß den Hersteller Richtung vorbereitet. </li><li> Nach Kulturen ausreichend, bei 4 ° C gezüchtet, bis sie benötigt aber halten Sie bei 4 ° C für zwei Tage vor der ein hohes Maß an Eiskeimbildungsaktivität gewährleisten. </li><li> Schaben Bakterien aus einer einzigen Platte von der Oberfläche des Agars mit einem Kunststoff, wegwerfbar oder wiederverwendbar Spatel zum Zeitpunkt der Verwendung und in 10-15 ml entionisiertem Wasser in einem 25 ml-Einwegküvette. Die Konzentration sollte im Bereich von 1 × 10 7 bis 1 x 10 9 be · </sup&gt; Ml -1. Die Lösung wird trübe erscheinen. Es besteht keine Notwendigkeit, um die Konzentration unter Verwendung eines Hämozytometers oder Spektralphotometer bestätigen, wie Konzentration muss nur ungefähr sein. </li><li> Vortexen Küvette für ein Minimum von 10 Sekunden, um die Bakterien zu verteilen. Hinweis: Die spezifische Konzentration des erhaltenen INA Gemisch ist nicht wichtig, und die beschriebenen Protokoll wird mehr als ein angemessenes Niveau der Eiskeimbildungsaktivität. Diese Mischung von INA-Bakterien und Wasser wird dann in den Keim Experimente eingesetzt werden. </li></ol> 3. Einrichten eines Freezing Experiment <ol><li> Legen Sie die hochauflösende Infrarotkamera (SC-660) im Inneren des Schutz Acryl-Box, so dass die Linse durch die Öffnung in der Vorderseite der Box und die Drähte der Kamera an einen Laptop oder Aufnahmegerät Ausgang Verbindung durch die hintere Öffnung der Box . Sichern Sie den Deckel der Box und legen Sie das Feld in der Klimakammer oder Gefrierschrank an einem Ort, werden alleow Gegenstand Pflanzenmaterials abzuwarten. <ol><li> Geben Sie einen dunklen Hintergrund rund um die Pflanze Material von den Wänden der Kammer mit schwarzem Papier, um Störungen durch reflektierte Infrarotenergie zu verhindern. </li><li> Setzen Sie die Kammer mit LED-Beleuchtung, um Heizung von der Lichtquelle zu minimieren, wenn die Aufzeichnung von Bildern in sichtbaren Wellenlängen erforderlich ist. Nur ein Minimum an Licht, wie beispielsweise eine batteriebetriebene Schrank Licht oder andere kleine LED-Vorrichtung wird benötigt für die Pflanzen durch die Kamera sichtbar sind. <ol><li> Sobald sichtbare Bilder des Themas Pflanzenmaterial aufgenommen werden, schalten Sie die LED-Beleuchtung. Alle von externen Kabelverbindungen (FireWire-Anschluss zum Computer, Netzkabel, etc.), um die Kamera über einen Port oder eine andere Öffnung in der Kammer. </li></ol></li><li> Füllen Sie zusätzlichen Platz im Hafen oder die Öffnung mit isolierendem Schaummaterial zu vermeiden oder zu reduzieren Temperaturgradienten innerhalb der Kammer. Stellen Sie die Anfangstemperatur der Kammer bei 1 ° C. </li></ol></li><li> Ausrichten Pflanzen oder Pflanzenteile, so dass das Pflanzenmaterial in der Gesichtsfeldansicht der Kamera und das Pflanzenmaterial sichtbar ist auf der Fernbetrachtungsschirm oder innerhalb des gewählten Software. </li><li> Erlauben Pflanzen auf 1 ° C für 30 min bis 1 Stunde vor dem Einleiten eines gesteuerten Gefrier Experiment äquilibrieren, abhängig von der Größe des Pflanzenmaterials. Dies stellt sicher, dass die Temperatur der Anlage nicht hinter Lufttemperatur liegen sie um viele Grade sobald das Einfrieren Experiment begonnen wird. Äquilibrierung wird erreicht, wenn die Temperatur des Pflanzenmaterials innerhalb von 0,5 ° C Lufttemperatur. <ol><li> Eine Schicht Styropor-Isolierung auf dem Boden von Topfpflanzen, wenn Topfpflanzen eingesetzt werden. Sobald die Pflanzen äquilibriert beginnen Abkühlen der Kammer. Anmerkung: Die Isolationsschicht auf der Bodenoberfläche des Topfes reduziert den Wärmeverlust weiter aus dem Topf zur Luft die Pflanze umgibt, und verhindert, dass die Wurzeln aus freezing, da dies in der Regel nicht während einer Frostereignis in der Natur vorkommen würde aufgrund der massiven Reservoir im Boden vorhandene Restwärme. </li></ol></li><li> Stellen Sie die gewünschten Kameraparameter (Farbpalette, Temperaturbereich, bestimmte Bereiche von Interesse, etc.), wie in 3.4.1-3.4.4 diskutiert. <ol><li> Wählen Sie den Regenbogen-Palette, um die Temperaturschwankungen angezeigt, während Sie das Live-Bild. </li><li> Den Temperaturspanne auf 5 ° C durch Einstellen der Temperatur Leiste unter dem Bild in der Software liegt. </li><li> Wählen Sie die lineare Skala (Algorithmus) zur Umwandlung der Infrarotdaten in das Falschfarbenbild, wie durch die ausgewählte Palette (rainbow) definiert und stellen Sie den Temperaturbereich bis 5 ° C und automatisch zu verfolgen auf der Basis des Bildes. Alternativ können die eingestellten Bereich manuell einzustellen, während Durchführung des Experiments. <ol><li> Verwenden Sie die Temperatur einen bestimmten Punkt oder eine durchschnittliche Temperatur innerhalb des definierten interessierenden Bereich von der Softw bereitgestelltsind. Abrufen der Temperaturdaten aller Pixel aus der aufgezeichneten Videosequenz oder aus den Informationen in der Bilddatei eingebettet sind, Fig. 3 zeigt eine typische Bildschirm aus ResearchIR Software. </li></ol></li><li> Platzieren Sie einen Cursor auf einem Standort auf dem Pflanzengewebe, die einen bestimmten Punkt von Interesse darstellt. Definieren Sie den Bereich von Interesse als Punkte (1 -3 Pixel groß), Boxen, Linien, Ellipsen oder Kreise. Mehrere Kombinationen von Punkten oder Formen können auf das Bild befinden. </li></ol></li><li> Aufzeichnen einer Videosequenz <ol><li> Stellen Sie die Kamera auf 60 Hz und für die Aufnahme manuell gestoppt werden aufzuzeichnen. </li><li> Geben Sie die Stelle auf dem Computer oder einem externen Laufwerk, in dem die aufgenommene Videodatei platziert wird. </li><li> Beginnt die Aufnahme. Hinweis: Aufnahme auf eine externe Festplatte ist sehr zu empfehlen, da große Video-Dateien generiert. Aufgenommenen Videodateien können später bearbeitet werden, um nur den Teil mit dem ne enthalten seinwendigen Informationen. Dies wird erheblich reduziert die Dateigröße. </li><li> Senken Sie die Temperatur der Kammer schrittweise um 0,5 -1,0 ° C. Warten Sie, bis die Anlage Temperatur im Gleichgewicht mit Lufttemperatur und dann senken Sie die Temperatur wieder um 0,5 bis 1,0 ° C. Je nach Masse des Pflanzengewebes beobachtet und ihre Morphologie kann Äquilibrierung 10-15 min dauern. Somit geben eine Abkühlgeschwindigkeit von etwa 4 ° C / h beträgt. </li><li> Fahren Sie so fort, bis die Anlage friert und Beobachtungen abgeschlossen sind. Beenden Sie die Aufnahme, wenn der Gefrierprozess abgeschlossen ist. Hinweis: Das Pflanzengewebe wurde mit Luft Temperatur äquilibriert, wenn das Pflanzenmaterial und den Hintergrund dieselbe Farbe haben, da sie bei der gleichen Temperatur. Da der Hintergrundtemperatur und der Temperatur des Pflanzengewebes gleich sind, kann es schwierig sein, das Pflanzenmaterial zu visualisieren, bis wieder senken die Temperatur, und es ist die Temperaturdifferenz zwischen dem Pflanzengewebe undir Temperatur. </li></ol></li></ol>

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P., Margesin, R., Schinner, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Ice nucleation and deep supercooling in plants: new insights using infrared thermography. In: Cold adapted organisms. Ecology, physiology, enzymologyandmolecularbiology. , 105-118 (1999).</li><li>Franks, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Biophysics and biochemistry at low temperatures. , (1985).</li><li>Neuner, G., Kuprian, E., Hincha, D., Zuther, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Infrared+thermal+analysis+of+plant+freezing+processes.">Infrared thermal analysis of plant freezing processes.</a> Methods in Molecular Biology: Plant Cold Acclimation. , 91-98 (2014).</li></ol>

Die Autoren haben keine konkurrierenden finanziellen Interessen oder Interessenkonflikte.

Wasser hat die Fähigkeit, auf Temperaturen von weit unter 0 ° C und der Temperatur, bei der Wasser gefriert können ziemlich variabel sein, zu unterkühlen. 36 Die Temperaturgrenze für die Unterkühlung von reinem Wasser beträgt etwa -40 ° C und wird als die homogene Keimpunkt definiert. Wenn Wasser gefriert bei Temperaturen wärmer als -40 ° C wird durch die Anwesenheit von heterogenen Keimbildnern, dass gebracht ermöglichen kleinen Eis Embryonen zu bilden, die dann als Katalysator für die Bildung vo...

Eisigen Temperaturen, die beim Pflanzen aktiv wachsenden auftreten kann tödlich sein, vor allem, wenn die Anlage hat wenig oder keine Frosttoleranz. Solche Frost Ereignisse haben oft verheerende Auswirkungen auf die landwirtschaftliche Produktion und können auch eine wichtige Rolle bei der Gestaltung der Gemeinschaftsstruktur in natürlichen Populationen von Pflanzen, vor allem im alpinen, subarktischen und arktischen Ökosysteme 1-6. Episoden starker Frühlingsfrösten haben große Auswirkungen auf Obstproduktion in den USA und Südamerika in den letzten Jahren 7-9 hatten und wurden von der frühen Beginn der warmen Wetter, gefolgt von mehr typische mittlere niedrigen Temperaturen verschärft. Die frühe warme Wetter induziert Knospen zu brechen, aktivieren das Wachstum der neuen Triebe, Blätter und Blumen, die alle sehr wenig bis gar keine Frosttoleranz 1,3,10-12 haben. Solche unberechenbar Wettermuster sind berichtet worden, um eine direkte Reflexion der fortschreitenden Klimawandel und erwartet, dass sie ein gemeinsames Wettermuster für die fores seineeable Zukunft 13. Die Bemühungen um die wirtschaftliche, effektive und umweltfreundliche Management-Techniken oder Agrochemikalien, die eine erhöhte Frosttoleranz bereitstellen kann einen begrenzten Erfolg für eine Vielzahl von Gründen, hatte aber das kann teilweise auf die Komplexität der Frosttoleranz und Gefriervermeidungsmechanismen in Pflanzen zurückgeführt werden. 14 Die adaptiven Mechanismen mit Frost Überleben in Pflanzen assoziiert sind traditionell in zwei Kategorien eingeteilt, das Einfrieren und Gefriertoleranz Vermeidung. Die erste Kategorie ist mit biochemischen Mechanismen, durch eine bestimmte Gruppe von Genen, die Pflanzen, die Spannungen mit der Gegenwart und der dehydratisierende Wirkung von Eis in seinen Geweben tolerieren ermöglichen geregelt verbunden. Während die letztere Kategorie ist typischerweise, aber nicht ausschließlich, mit strukturellen Aspekte einer Pflanze, die, wenn zu bestimmen, wann und wo sich Eis in einer Pflanze 14 verbunden. Trotz der Prävalenz von freeze Vermeidung als Ad-aptive Mechanismus hat wenig Forschung in der letzten Zeit für das Verständnis der zugrunde liegenden Mechanismen und Regulation der Gefriert Vermeidung gewidmet. Der Leser wird auf eine aktuelle Übersicht 15 für mehr Details zu diesem Thema verwiesen. Während die Bildung von Eis bei niedrigen Temperaturen kann wie ein einfacher Prozess erscheinen, viele Faktoren tragen zur Bestimmung der Temperatur, bei der Eis Keime in Pflanzengewebe und wie es breitet sich in der Anlage. Parameter, wie die Anwesenheit von extrinsischen und intrinsischen Eiskernbildnern heterogene vs. homogene Keimbildung, thermische Hysterese (Frostschutz) Proteine, das Vorhandensein von spezifischen Zucker und anderen Osmolyten, und eine Vielzahl von strukturellen Aspekte der Anlage spielen eine bedeutende Rolle des Gefrierprozesses in Pflanzen. Gemeinsam beeinflussen diese Parameter die Temperatur, bei der eine Pflanze friert, wo Eis initiiert und wie es wächst. Sie kann auch Auswirkungen auf die Morphologie der resultierenden Eiskristallen.Verschiedene Verfahren wurden verwendet, um den Gefrierprozeß in Pflanzen, die unter Laborbedingungen, einschließlich kernmagnetischer Resonanzspektroskopie (NMR) 16, Kernspintomographie (MRI) 17, Kryo-Mikroskopie 18-19 und Tieftemperatur-Rasterelektronenmikroskopie Studie (LTSEM ). 20 Freezing ganzer Pflanzen im Labor- und Feldeinstellungen hat sich jedoch hauptsächlich mit Thermoelementen überwacht. Die Verwendung von Thermoelementen, um ein Einfrieren zu studieren ist an der Freisetzung von Wärme (Schmelzwärme), wenn das Wasser einen Phasenübergang von einer Flüssigkeit zu einem Feststoff basiert. Einfrieren wird dann als exotherme Ereignis aufgezeichnet. 21-23 Obwohl Thermoelemente sind das typische Verfahren der Wahl bei der Untersuchung Gefrier in Pflanzen hat ihre Verwendung vielen Einschränkungen, die die Menge an Informationen während eines Gefrier Ereignis erhalten begrenzen. Zum Beispiel mit Thermoelementen ist es schwierig, fast unmöglich, zu bestimmen, wo das Eis in Pflanzen angezogen, wie sie ausbreitet,wenn es breitet sich mit gleichmäßiger Rate, und wenn einige Gewebe eisfrei bleiben. Fortschritte in der hochauflösenden Infrarot-Thermografie (HRIT) 24-27 haben jedoch deutlich erhöht die Fähigkeit, Informationen über den Gefrierprozess in ganzen Pflanzen zu erhalten, vor allem, wenn in einem Differential Imaging-Modus verwendet. 28-33 In diesem Bericht haben wir beschreiben die Verwendung dieser Technologie, um verschiedene Aspekte des Gefrierprozesses und verschiedene Parameter, die beeinflussen, wo und bei welcher Temperatur und Eis in Pflanzen begonnen zu studieren. Ein Protokoll vorgestellt werden, die die Fähigkeit des Eis-Nukleation aktive (INA) Bakterium Pseudomonas syringae (Cit-7) als extrinsische Nukleator Einleitung Einfrieren in eine krautige Pflanze auf einem hohen, unter Null Temperatur handeln demonstrieren wird. Hochauflösende Infrarot-Kamera Das Protokoll und die Beispiele in diesem Bericht dokumentiert nutzen eine hochauflösende Infrarot-Video-Radiometer. Das Radiometer (1) liefert eine Kombination von Infrarot- und sichtbaren Spektrums Bilder und Temperaturdaten. Die spektrale Empfindlichkeit der Kamera ist im Bereich von 7,5 bis 13,5 um und liefert 640 x 480 Pixel Auflösung. Sichtbaren Spektrums erzeugt Bilder durch die integrierte Kamera mit IR-Bilder in Echtzeit, was die Auslegung komplexer, Wärmebilder erleichtert fusioniert werden. Eine Reihe von Linsen für die Kamera verwendet werden, um Nahaufnahmen und mikroskopische Beobachtungen zu machen. Die Kamera kann in einem Standalone-Modus verwendet werden, oder eine Schnittstelle mit einem Laptop mit propietary Software gesteuert. Die Software kann verwendet werden, um eine Vielzahl von in den aufgezeichneten Videos eingebettet thermische Daten zu erhalten. Es ist wichtig anzumerken, dass eine Vielzahl von Infrarot-Radiometer sind kommerziell verfügbar. Daher ist es wichtig, dass die Forscher über ihre vorgesehene Anwendung mit einem erfahrenen Produktingenieur und dass der Forscher testen die Fähigkeit eines spezific Radiometer, um die erforderlichen Informationen zur Verfügung stellen. Das bildgebende Radiometer in der beschriebenen Protokoll verwendet wird, in einem Acryl-Box mit Styropor i n, um die Exposition während der Erwärmung und Abkühlung Protokolle, um Kondensation abzuschrecken isoliert (Abbildung 2) platziert. Dieser Schutz wird nicht für alle Kameras oder Anwendungen benötigt.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Infrared Camera</td>
			<td>FLIR</td>
			<td>SC-660</td>
			<td>Many models available depending on application</td>
		</tr>
		<tr>
			<td>Infrared Analytical Software</td>
			<td>FLIR</td>
			<td>ResearchIR 4.10.2.5</td>
			<td align="right">$3,500</td>
		</tr>
		<tr>
			<td rowspan="2">Pseudomonas syringae (strain Cit-7)</td>
			<td rowspan="2"></td>
			<td rowspan="2"></td>
			<td rowspan="2"><a href="mailto:icelab@berkeley.edu">Kindly provided by Dr. Steven Lindow, University of California&#160; Berkeley icelab@berkeley.edu</a></td>
		</tr>
		<tr>
		</tr>
		<tr>
			<td>Pseudomonas Agar F</td>
			<td>Fisher Scientific</td>
			<td>DF0448-17-1</td>
			<td></td>
		</tr>
	</tbody>
</table>

the use of high-resolution infrared thermography (hrit) for the study of ice nucleation and ice propagation in plants

Freezing events that occur when plants are actively growing can be a lethal event, particularly if the plant has no freezing tolerance. Such frost events often have devastating effects on agricultural production and can also play an important role in shaping community structure in natural populations of plants, especially in alpine, sub-arctic, and arctic ecosystems. Therefore, a better understanding of the freezing process in plants can play an important role in the development of methods of frost protection and understanding mechanisms of freeze avoidance. Here, we describe a protocol to visualize the freezing process in plants using high-resolution infrared thermography (HRIT). The use of this technology allows one to determine the primary sites of ice formation in plants, how ice propagates, and the presence of ice barriers. Furthermore, it allows one to examine the role of extrinsic and intrinsic nucleators in determining the temperature at which plants freeze and evaluate the ability of various compounds to either affect the freezing process or increase freezing tolerance. The use of HRIT allows one to visualize the many adaptations that have evolved in plants, which directly or indirectly impact the freezing process and ultimately enables plants to survive frost events.

Ice-Keimbildungsaktivität der Ice + Bakterium Pseudomonas syringae (Stamm Cit-7) Ein 10 ul Tropfen Wasser und 10 ul Wasser, P. syringae (Cit-7) wurden auf der Oberfläche eines abaxial Hosta Blatt gelegt (Hosta spp.) (Figur 4). Wie dargestellt, ist der Wassertropfen, die die INA Bakterien erstarrte erste und war für das Blatt zu induzieren zu frieren, während der Tropfen Wasser auf der Blattoberfläche blieb nicht gefrorenen verantwortlich. ...

Watch this Scientific Journal Video about The Use of High-resolution Infrared Thermography HRIT for the Study of Ice Nucleation and Ice Propagation in Plants at JoVE.com

The Use of High-resolution Infrared Thermography HRIT for the Study of Ice Nucleation and Ice Propagation in Plants

Here we present a protocol that allows one to visualize sites of ice formation and avenues of ice propagation in plants utilizing high resolution infrared thermography (HRIT).

Die Verwendung von hochauflösenden Infrarot-Thermografie (HRIT) für das Studium der Ice Ice Keimbildung und Ausbreitung in Pflanzen

Freezing events that occur when plants are actively growing can be a lethal event, particularly if the plant has no freezing tolerance. Such frost events ...

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The Use of High-resolution Infrared Thermography (HRIT) for the Study of Ice Nucleation and Ice Propagation in Plants

9.5K Aufrufe.  Agricultural Research Service (USDA-ARS), Kearneysville, WV. Here we present a protocol that allows one to visualize sites of ice formation and avenues of ice propagation in plants utilizing high resolution infrared thermography (HRIT).

Serie: The Use of High-resolution Infrared Thermography HRIT for the Study of Ice Nucleation and Ice Propagation in Plants

Expression of Recombinant Cellulase Cel5A from Trichoderma reesei in Tobacco Plants

Cellulose degrading enzymes, cellulases, are targets of both research and industrial interests. The preponderance of these enzymes in difficult-to-culture organisms, such as hyphae-building fungi and anaerobic bacteria, has hastened the use of recombinant technologies in this field. Plant expression methods are a desirable system for large-scale production of enzymes and other industrially useful proteins. Herein, methods for the transient expression of a fungal endoglucanase, Trichoderma reesei Cel5A, in Nicotiana tabacum are demonstrated. Successful protein expression is shown, monitored by fluorescence using an mCherry-enzyme fusion protein. Additionally, a set of basic tests are used to examine the activity of transiently expressed T. reesei Cel5A, including SDS-PAGE, Western blotting, zymography, as well as&#160;fluorescence and dye-based substrate degradation assays. The system described here can be used to produce an active cellulase in a short time period, so as to assess the potential for further production in plants through constitutive or inducible expression systems.

Expression of Recombinant Cellulase Cel5A from Trichoderma reesei in Tobacco Plants

Tobacco plants were used to produce a fungal cellulase, TrCel5A, via a transient expression system. The expression could be monitored using a fluorescent fusion protein, and the protein activity was characterized post-expression.

Expression von rekombinanten Cellulase aus Cel5A<em&gt; Trichoderma reesei</em&gt; In Tabakpflanzen

Cellulose degrading enzymes, cellulases, are targets of both research and industrial interests. The preponderance of these enzymes in difficult-to-culture ...

Forschung

Umwelt

Enhancement of the Initial Growth Rate of Agricultural Plants by Using Static Magnetic Fields

Electronic devices and high-voltage wires induce magnetic fields. A magnetic field of 1,300-2,500 Gauss (0.2 Tesla) was applied to Petri dishes containing seeds of Garden Balsam (Impatiens balsamina), Mizuna (Brassica rapa var. japonica), Komatsuna (Brassica rapa var. perviridis), and Mescluns (Lepidium sativum). We applied magnets under the culture dish. During the 4 days of application, we observed that the stem and root length increased. The group subjected to magnetic field treatment (n = 10) showed a 1.4 times faster rate of growth compared with the control group (n = 11) in a total of 8 days (p &#60;0.0005). This rate is 20% higher than that reported in previous studies. The tubulin complex lines did not have connecting points, but connecting points occur upon the application of magnets. This shows complete difference from the control, which means abnormal arrangements. However, the exact cause remains unclear. These results of growth enhancement of applying magnets suggest that it is possible to enhance the growth rate, increase productivity, or control the speed of germination of plants by applying static magnetic fields. Also, magnetic fields can cause physiological changes in plant cells and can induce growth. Therefore, stimulation with a magnetic field can have possible effects that are similar to those of chemical fertilizers, which means that the use of fertilizers can be avoided.

The goal of this protocol is to demonstrate the acceleration of the initial growth rate of plants by applying static magnetic fields with no external energy.

Verbesserung des Anfangswachstumsrate von landwirtschaftlichen Pflanzen durch statische Magnetfelder

Electronic devices and high-voltage wires induce magnetic fields. A magnetic field of 1,300-2,500 Gauss (0.2 Tesla) was applied to Petri dishes containing ...

Use of a Battery of Chemical and Ecotoxicological Methods for the Assessment of the Efficacy of Wastewater Treatment Processes to Remove Estrogenic Potency

Endocrine Disrupting Compounds pose a substantial risk to the aquatic environment. Ethinylestradiol (EE2) and estrone (E1) have recently been included in a watch list of environmental pollutants under the European Water Framework Directive. Municipal wastewater treatment plants are major contributors to the estrogenic potency of surface waters. Much of the estrogenic potency of wastewater treatment plant (WWTP) effluents can be attributed to the discharge of steroid estrogens including estradiol (E2), EE2 and E1 due to incomplete removal of these substances at the treatment plant. An evaluation of the efficacy of wastewater treatment processes requires the quantitative determination of individual substances most often undertaken using chemical analysis methods. Most frequently used methods include Gas Chromatography-Mass Spectrometry (GCMS/MS) or Liquid Chromatography-Mass Spectrometry (LCMS/MS) using multiple reaction monitoring (MRM). Although very useful for regulatory purposes, targeted chemical analysis can only provide data on the compounds (and specific metabolites) monitored. Ecotoxicology methods additionally ensure that any by-products produced or unknown estrogenic compounds present are also assessed via measurement of their biological activity. A number of in vitro bioassays including the Yeast Estrogen Screen (YES) are available to measure the estrogenic activity of wastewater samples. Chemical analysis in conjunction with in vivo and in vitro bioassays provides a useful toolbox for assessment of the efficacy and suitability of wastewater treatment processes with respect to estrogenic endocrine disrupting compounds. This paper utilizes a battery of chemical and ecotoxicology tests to assess conventional, advanced and emerging wastewater treatment processes in laboratory and field studies.

Endocrine Disrupting Compounds (EDC) pose a substantial risk to the aquatic environment. Municipal wastewater treatment plants are major contributors to the estrogenic potency of surface waters. The methodology provided in this paper allows for an assessment of the efficacy and suitability of wastewater treatment processes with respect to EDC removal.

Die Verwendung einer Batterie von chemischen und ökotoxikologischen Methoden zur Beurteilung der Wirksamkeit von Abwasserbehandlungsprozesse zu entfernen Estrogenic Potency

Endocrine Disrupting Compounds pose a substantial risk to the aquatic environment. Ethinylestradiol (EE2) and estrone (E1) have recently been included in ...

Empirical, Metagenomic, and Computational Techniques Illuminate the Mechanisms by which Fungicides Compromise Bee Health

Growers often use fungicide sprays during bloom to protect crops against disease, which exposes bees to fungicide residues. Although considered &#34;bee-safe,&#34; there is mounting evidence that fungicide residues in pollen are associated with bee declines (for both honey and bumble bee species). While the mechanisms remain relatively unknown, researchers have speculated that bee-microbe symbioses are involved. Microbes play a pivotal role in the preservation and/or processing of pollen, which serves as nutrition for larval bees. By altering the microbial community, it is likely that fungicides disrupt these microbe-mediated services, and thereby compromise bee health. This manuscript describes the protocols used to investigate the indirect mechanism(s) by which fungicides may be causing colony decline. Cage experiments exposing bees to fungicide-treated flowers have already provided the first evidence that fungicides cause profound colony losses in a native bumble bee (Bombus impatiens). Using field-relevant doses of fungicides, a series of experiments have been developed to provide a finer description of microbial community dynamics of fungicide-exposed pollen. Shifts in the structural composition of fungal and bacterial assemblages within the pollen microbiome are investigated by next-generation sequencing and metagenomic analysis. Experiments developed herein have been designed to provide a mechanistic understanding of how fungicides affect the microbiome of pollen-provisions. Ultimately, these findings should shed light on the indirect pathway through which fungicides may be causing colony declines.

Microbial consortia within bumble bee hives enrich and preserve pollen for bee larvae. Using next generation sequencing, along with laboratory and field-based experiments, this manuscript describes protocols used to test the hypothesis that fungicide residues alter the pollen microbiome, and colony demographics, ultimately leading to colony loss.

Empirische, metagenomischen und Computertechniken beleuchten die Mechanismen, durch welche Fungizide Kompromiss Bienengesundheit

Growers often use fungicide sprays during bloom to protect crops against disease, which exposes bees to fungicide residues. Although considered ...

Automated, High-resolution Mobile Collection System for the Nitrogen Isotopic Analysis of NOx

Nitrogen oxides (NOx = NO + NO2) are a family of atmospheric trace gases that have great impact on the environment. NOx concentrations directly influence the oxidizing capacity of the atmosphere through interactions with ozone and hydroxyl radicals. The main sink of NOx is the formation and deposition of nitric acid, a component of acid rain and a bioavailable nutrient. NOx is emitted from a mixture of natural and anthropogenic sources, which vary in space and time. The collocation of multiple sources and the short lifetime of NOx make it challenging to quantitatively constrain the influence of different emission sources and their impacts on the environment. Nitrogen isotopes of NOx have been suggested to vary amongst different sources, representing a potentially powerful tool to understand the sources and transport of NOx. However, previous methods of collecting atmospheric NOx integrate over long (week to month) time spans and are not validated for the efficient collection of NOx in relevant, diverse field conditions. We report on a new, highly efficient field-based system that collects atmospheric NOx for isotope analysis at a time resolution between 30 min and 2 hr. This method collects gaseous NOx in solution as nitrate with 100% efficiency under a variety of conditions. Protocols are presented for collecting air in urban settings under both stationary and mobile conditions. We detail the advantages and limitations of the method and demonstrate its application in the field. Data from several deployments are shown to 1) evaluate field-based collection efficiency by comparisons with in situ NOx concentration measurements, 2) test the stability of stored solutions before processing, 3) quantify in situ reproducibility in a variety of urban settings, and 4) demonstrate the range of N isotopes of NOx detected in ambient urban air and on heavily traveled roadways.

Automated, High-resolution Mobile Collection System for the Nitrogen Isotopic Analysis of NOx

Previous work suggests that the nitrogen isotopic composition of atmospheric nitrogen oxides might distinguish the influence of different sources in the environment. We report on an automated, mobile, field-based method for the high collection efficiency of atmospheric NOx for N isotopic analysis at an hourly time resolution.

Automatisierte, hochauflösende Mobilsammelsystem für die Stickstoffisotopenanalyse von NO<sub&gt; x</sub

Nitrogen oxides (NOx = NO + NO2) are a family of atmospheric trace gases that have great impact on the environment. NOx concentrations directly influence ...

The Calibration and Use of Capacitance Sensors to Monitor Stem Water Content in Trees

Water transport and storage through the soil-plant-atmosphere continuum is critical to the terrestrial water cycle, and has become a major research focus area. Biomass capacitance plays an integral role in the avoidance of hydraulic impairment to transpiration. However, high temporal resolution measurements of dynamic changes in the hydraulic capacitance of large trees are rare. Here, we present procedures for the calibration and use of capacitance sensors, typically used to monitor soil water content, to measure the volumetric water content in trees in the field. Frequency domain reflectometry-style observations are sensitive to the density of the media being studied. Therefore, it is necessary to perform species-specific calibrations to convert from the sensor-reported values of dielectric permittivity to volumetric water content. Calibration is performed on a harvested branch or stem cut into segments that are dried or re-hydrated to produce a full range of water contents used to generate a best-fit regression with sensor observations. Sensors are inserted into calibration segments or installed in trees after pre-drilling holes to a tolerance fit using a fabricated template to ensure proper drill alignment. Special care is taken to ensure that sensor tines make good contact with the surrounding media, while allowing them to be inserted without excessive force. Volumetric water content dynamics observed via the presented methodology align with sap flow measurements recorded using thermal dissipation techniques and environmental forcing data. Biomass water content data can be used to observe the onset of water stress, drought response and recovery, and has the potential to be applied to the calibration and evaluation of new plant-level hydrodynamics models, as well as to the partitioning of remotely sensed moisture products into above- and belowground components.

The hydraulic capacitance of biomass is a key component of the vegetation water budget, which serves as a buffer against short and long-term drought stresses. Here, we present a protocol for the calibration and use of soil moisture capacitance sensors to monitor water content in the stems of large trees.

Die Kalibrierung und die Nutzung der Kapazität Sensoren Stem Wassergehalt in den Bäumen zu überwachen

Water transport and storage through the soil-plant-atmosphere continuum is critical to the terrestrial water cycle, and has become a major research focus ...

Stress Distribution During Cold Compression of Rocks and Mineral Aggregates Using Synchrotron-based X-Ray Diffraction

We report detailed procedures for performing compression experiments on rocks and mineral aggregates within a multi-anvil deformation apparatus (D-DIA) coupled with synchrotron X-radiation. A cube-shaped sample assembly is prepared and compressed, at room temperature, by a set of four X-ray transparent sintered diamond anvils and two tungsten carbide anvils, in the lateral and the vertical planes, respectively. All six anvils are housed within a 250-ton hydraulic press and driven inward simultaneously by two wedged guide blocks. A horizontal energy dispersive X-ray beam is projected through and diffracted by the sample assembly. The beam is commonly in the mode of either white or monochromatic X-ray. In the case of white X-ray, the diffracted X-rays are detected by a solid-state detector array that collects the resulting energy dispersive diffraction pattern. In the case of monochromatic X-ray, the diffracted pattern is recorded using a two-dimensional (2-D) detector, such as an imaging plate or a charge-coupled device (CCD) detector. The 2-D diffraction patterns are analyzed to derive lattice spacings. The elastic strains of the sample are derived from the atomic lattice spacing within grains. The stress is then calculated using the predetermined elastic modulus and the elastic strain. Furthermore, the stress distribution in two-dimensions allow for understanding how stress is distributed in different orientations. In addition, a scintillator in the X-ray path yields a visible light image of the sample environment, which allows for the precise measurement of sample length changes during the experiment, yielding a direct measurement of volume strain on the sample. This type of experiment can quantify the stress distribution within geomaterials, which can ultimately shed light on the mechanism responsible for compaction. Such knowledge has the potential to significantly improve our understanding of key processes in rock mechanics, geotechnical engineering, mineral physics, and material science applications where compactive processes are important.

We report detailed procedures for compression experiments on rocks and mineral aggregates within a multi-anvil deformation apparatus coupled with synchrotron X-radiation. Such experiments allow quantification of the stress distribution within samples, that ultimately sheds light on compaction processes in geomaterials.

Spannungsverteilung bei kalten Verdichtung von Steinen und mineralischen Zuschlagsstoffen mit Synchrotron-basierte x-Ray Diffraction

We report detailed procedures for performing compression experiments on rocks and mineral aggregates within a multi-anvil deformation apparatus (D-DIA) ...

Protocol for Producing Three-Dimensional Infrared Video of Freezing in Plants

Freezing in plants can be monitored using infrared (IR) thermography, because when water freezes, it gives off heat. However, problems with color contrast make 2-dimensions (2D) infrared images somewhat difficult to interpret. Viewing an IR image or the video of plants freezing in 3 dimensions (3D) would allow a more accurate identification of sites for ice nucleation as well as the progression of freezing. In this paper, we demonstrate a relatively simple means to produce a 3D infrared video of a strawberry plant freezing. Strawberry is an economically important crop that is subjected to unexpected spring freeze events in many areas of the world. An accurate understanding of the freezing in strawberry will provide both breeders and growers with more economical ways to prevent any damage to plants during freezing conditions.
The technique involves a positioning of two IR cameras at slightly different angles to film the strawberry freezing. The two video streams will be precisely synchronized using a screen capture software that records both cameras simultaneously. The recordings will then be imported into the imaging software and processed using an anaglyph technique. Using red-blue glasses, the 3D video will make it easier to determine the precise site of ice nucleation on leaf surfaces.

Here, we present a protocol to image a strawberry plant freezing in 3 dimensions. Two infrared cameras positioned at slightly different angles are used to produce a red-blue anaglyph video to observe the freezing of the plant in 3 dimensions.

Protokoll zur Herstellung von dreidimensionalen Infrarot-Video des Einfrierens in Pflanzen

Freezing in plants can be monitored using infrared (IR) thermography, because when water freezes, it gives off heat. However, problems with color contrast ...

Long-term Video Tracking of Cohoused Aquatic Animals: A Case Study of the Daily Locomotor Activity of the Norway Lobster (Nephrops norvegicus)

We present a protocol related to a video-tracking technique based on the background subtraction and image thresholding that makes it possible to individually track cohoused animals. We tested the tracking routine with four cohoused Norway lobsters (Nephrops norvegicus) under light-darkness conditions for 5 days. The lobsters had been individually tagged. The experimental setup and the tracking techniques used are entirely based on the open source software. The comparison of the tracking output with a manual detection indicates that the lobsters were correctly detected 69% of the times. Among the correctly detected lobsters, their individual tags were correctly identified 89.5% of the times. Considering the frame rate used in the protocol and the movement rate of lobsters, the performance of the video tracking has a good quality, and the representative results support the validity of the protocol in producing valuable data for research needs (individual space occupancy or locomotor activity patterns). The protocol presented here can be easily customized and is, hence, transferable to other species where the individual tracking of specimens in a group can be valuable for answering research questions.

Long-term Video Tracking of Cohoused Aquatic Animals: A Case Study of the Daily Locomotor Activity of the Norway Lobster Nephrops norvegicus

Here we present a protocol to individually track animals over a long period of time. It uses computer vision methods to identify a set of manually constructed tags by using a group of lobsters as case study, simultaneously providing information on how to house, manipulate, and mark the lobsters.

Langfristige Video Tracking Cohoused Wassertiere: A Case Study of der Bewegungsorgane Tagesaktivität der Kaisergranat (Nephrops Norvegicus)

We present a protocol related to a video-tracking technique based on the background subtraction and image thresholding that makes it possible to ...

Single-throughput Complementary High-resolution Analytical Techniques for Characterizing Complex Natural Organic Matter Mixtures

Natural organic matter (NOM) is composed of a highly complex mixture of thousands of organic compounds which, historically, proved difficult to characterize. However, to understand the thermodynamic and kinetic controls on greenhouse gas (carbon dioxide [CO2] and methane [CH4]) production resulting from the decomposition of NOM, a molecular-level characterization coupled with microbial proteome analyses is necessary. Further, climate and environmental changes are expected to perturb natural ecosystems, potentially upsetting complex interactions that influence both the supply of organic matter substrates and the microorganisms performing the transformations. A detailed molecular characterization of the organic matter, microbial proteomics, and the pathways and transformations by which organic matter is decomposed will be necessary to predict the direction and magnitude of the effects of environmental changes. This article describes a methodological throughput for comprehensive metabolite characterization in a single sample by direct injection Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS), gas chromatography mass spectrometry (GC-MS), nuclear magnetic resonance (NMR) spectroscopy, liquid chromatography mass spectrometry (LC-MS), and proteomics analysis. This approach results in a fully-paired dataset which improves statistical confidence for inferring pathways of organic matter decomposition, the resulting CO2 and CH4 production rates, and their responses to environmental perturbation. Herein we present results of applying this method to NOM samples collected from peatlands; however, the protocol is applicable to any NOM sample (e.g., peat, forested soils, marine sediments, etc.).

This protocol describes a single throughput for complementary analytical and omics techniques culminating in a fully-paired characterization of natural organic matter and microbial proteomics in different ecosystems. This approach permits robust comparisons for identifying metabolic pathways and transformations important for describing greenhouse gas production and predicting responses to environmental change.