Name: a whole cell bioreporter approach to assess transport and bioavailability of organic contaminants in water unsaturated systems
Uploaded: 2014-12-24T15:00:00
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Funding by the German Environmental Foundation (DBU) is acknowledged. The authors thank Ute Kuhlicke for technical help with CLSM analysis and Birgit W&#252;rz, Rita Remer, and Jana Reichenbach for skilled experimental help. The authors would particularly like to thank Prof. Jan Roelof van der Meer and Dr. Robin Tecon for fruitful discussion and providing the bioreporter strain. It contributes to the &#8216;Chemicals in the Environment&#8217; (CITE) research program of the Helmholtz Association.

1. Preparazione di piatti, Scivoli e incubazione Chambers <ol><li> Preparare il seguente materiale per ogni microcosmo: un grande plastico Petri fondo piatto (d = 10 cm), uno modificato (vedi punto 1.2) piccolo plastica Petri fondo piatto (d = 5 cm) con coperchi e un conteggio vetrino da camera con tre cavità. </li><li> Prendete il numero desiderato di Petri parti inferiori piatto (d = 5 cm). Rimuovere parte della tesa con una sega per adattarsi esattamente una diapositiva (26 mm di bordo). Per sterilizzare il sistem...

<ol>
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Plant Biol. 57, 233-266 (2006).</li><li>Furuno, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mycelia+promote+active+transport+and+spatial+dispersion+of+polycyclic+aromatic+hydrocarbons.">Mycelia promote active transport and spatial dispersion of polycyclic aromatic hydrocarbons.</a> Environ. Sci. Technol. 46, 5463-5470 (2012).</li><li>Gao, Y., Cheng, Z., Ling, W., Huang, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Arbuscular+mycorrhizal+fungal+hyphae+contribute+to+the+uptake+of+polycyclic+aromatic+hydrocarbons+by+plant+roots.">Arbuscular mycorrhizal fungal hyphae contribute to the uptake of polycyclic aromatic hydrocarbons by plant roots.</a> Bioresour. Technol. 101, 6895-6901 (2010).</li><li>Semple, K. T., Morriss, A. W. J., Paton, G. 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R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+green+fluorescent+protein-based+bacterial+biosensor+for+analysing+phenanthrene+fluxes.">A new green fluorescent protein-based bacterial biosensor for analysing phenanthrene fluxes.</a> Environ. Microbiol. 8, 697-708 (2006).</li><li>Schamfuss, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impact+of+mycelia+on+the+accessibility+of+fluorene+to+PAH-degrading+bacteria.">Impact of mycelia on the accessibility of fluorene to PAH-degrading bacteria.</a> Environ. Sci. Technol. 47, 6908-6915 (2013).</li><li>Smibert, R. M., Krieg, R. M., Gerhardt, P., Murray, R. G. E., Costilow, R. N., Nester, E. W., Wood, W. A., Krieg, N. R., Phillips, G. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Manual of methods for general bacteriology. 19, 409-443 (1981).</li><li>Wu, C. H., Warren, H. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Natural+autofluorescence+in+fungi+and+its+correlation+with+viability.">Natural autofluorescence in fungi and its correlation with viability.</a> Mycologia. 76, 1049-1058 (1984).</li><li>Schneider, C. A., Rasband, W. S., Eliceiri, K. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NIH+Image+to+ImageJ:+25+years+of+image+analysis.">NIH Image to ImageJ: 25 years of image analysis.</a> Nat. Methods. 9, 671-675 (2012).</li><li>Pedersen, C., Sylvia, D., Mukerji, K. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ch.+8+Concepts+in+Mycorrhizal+Research.">Ch. 8 Concepts in Mycorrhizal Research.</a> Handbook of Vegetation Science. Vol. 19, 195-222 (1996).</li><li>Furuno, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fungal+mycelia+allow+chemotactic+dispersal+of+polycyclic+aromatic+hydrocarbon-degrading+bacteria+in+water-unsaturated+systems.">Fungal mycelia allow chemotactic dispersal of polycyclic aromatic hydrocarbon-degrading bacteria in water-unsaturated systems.</a> Environ. Microbiol. 12, 1391-1398 (2010).</li></ol>

La configurazione microcosmo presentato dimostrato adatto per studiare la biodisponibilità di sostanze chimiche spazialmente separate per degradare organismi dopo l&#39;assorbimento e il trasporto da miceli. Potenziale di trasporto del gas in fase di composti volatili parzialmente è impedito e le cellule di marcatura già batteriche può essere visualizzato senza preparazione del campione elaborato e quindi con il minimo disturbo del sistema sensibile. Allo stesso tempo, l&#39;analisi chimica del campione può essere ...

Suolo è densamente popolato da una vasta gamma di microrganismi come batteri 1,2. Tuttavia, le condizioni in questo habitat sono impegnativi, soprattutto in termini di disponibilità di acqua 3. I batteri in modo permanente devono cercare le condizioni ottimali in ambienti eterogenei 4, ma l&#39;assenza di film d&#39;acqua continuo si traduce in mobilità limitata 5 ostacolando loro di diffondersi liberamente. Inoltre, i tassi di diffusione di soluti (ad esempio, nutrienti) sono abbassati in condizioni insaturi 6. Così, batteri e sostanze nutrienti sono spesso separati fisicamente e l&#39;accessibilità dei nutrienti è limitato a 3. Di conseguenza, un vettore di trasporto per i composti chimici che non richiede un acqua-fase continua potrebbe contribuire a superare queste limitazioni. In realtà, molti microrganismi, come funghi e oomiceti hanno sviluppato una forma di crescita filamentosa che permette loro di crescere attraverso pori pieni d&#39;aria e raggiungere in tal modo mobilizing anche fisica nutrienti 7 e 8 sostanze carboniose separato su lunghe distanze. Essi possono anche fungere da vettori di trasporto biologici che forniscono zuccheri e altre fonti di energia ai batteri 9. Assorbimento e trasporto in organismi miceliari è stato anche dimostrato di inquinanti organici idrofobici come gli idrocarburi policiclici aromatici (IPA) in Pythium ultimum 10 o nei funghi micorrizici arbuscolari 11. Dal PAH sono contaminanti solubili in acqua onnipresenti e scarsamente 12 nel suolo, trasporti miceli mediata potrebbe contribuire ad aumentare la biodisponibilità dei contaminanti per i potenziali degraders batteriche. Considerando che l&#39;importo totale del trasporto inquinante può essere quantificato direttamente chimica significa 10, la biodisponibilità dei contaminanti trasportati da miceli a batteri degradanti e altri organismi non può essere valutata facilmente. Il protocollo che segue presenta un metodo per valutare l&#39;impatto di MyCelia sulla biodisponibilità contaminante degraders batteriche in modo diretto; permette la raccolta di informazioni sull&#39;impatto spazio-temporale dei contaminanti sugli ecosistemi microbici. Descriviamo come impostare un elaborato sistema di microcosmo insaturo imitando interfacce aria-acqua nel suolo collegando una sorgente puntiforme PAH separati fisicamente con PAH-degradanti marcatura già batteri tramite vettori di trasporto miceliari. Poiché è escluso il trasporto aereo, l&#39;effetto del trasporto miceliale basata sulla PAH biodisponibilità dei batteri può essere studiata in modo isolato. Più in dettaglio, tre anelli PAH fluorene, l&#39;organismo miceliale Pythium ultimum e la marcatura già batterio Burkholderia sartisoli RP037-Mche 13 sono stati applicati nelle configurazioni microcosmo descritti. Il batterio B. sartisoli RP037-Mche è stato originariamente costruito per studiare flussi fenantrene alla cella 14 ed esprime migliorati proteina fluorescente verde (eGFP) per effetto del flusso di PAHla cella, mentre il rosso fluorescente mCherry è espressa costitutivamente. Informazioni dettagliate sulla costruzione giornalista è data da Tecon et al. 13 Nei test preliminari, il batterio non ha rivelato alcuna nuoto e solo molto lenta capacità sciamatura. Era in grado di migrare lentamente ife di Pythium ultimum quando applicato come una sospensione densa sopra il ife. Poiché i batteri sono stati incorporati in agarosio nella seguente protocollo, migrazione ife non si è verificato. Utilizzando microscopio confocale a scansione laser (CLSM), i batteri marcatura già possono essere visualizzati direttamente in microcosmi ed espressione di eGFP possono essere quantificati in relazione alla quantità di cellule (proporzionale al segnale mCherry) con l&#39;aiuto del ImageJ software. Questo permette di confrontare biodisponibilità qualitativamente in diversi scenari (cioè, superiore o inferiore). FLU è risultato essere biodisponibile dopo trasporto micelio di P. ultimum (cioè,era superiore a quello di un controllo negativo). Inoltre, il protocollo descrive come quantificare l&#39;ammontare totale del trasporto miceli mediato tramite mezzi chimici e verificare la biodisponibilità dei contaminanti con fibre di vetro siliconato (fibre SPME) in microcosmi identici. I risultati con questa configurazione microcosmo sono stati pubblicati e discussi per la combinazione di P. ultimum, fluorene e B. sartisoli RP037-Mche 15. Qui, l&#39;attenzione si concentra sulla descrizione metodo dettagliato e l&#39;individuazione di potenziali insidie ​​del protocollo di fornire questa conoscenza per i potenziali ulteriori applicazioni. Ulteriori applicazioni possono coinvolgere vari fungine, specie batteriche (ad esempio, da siti contaminati), e altri contaminanti (ad esempio, i pesticidi) o contaminante di alimentazione (ad esempio, i terreni di età).

<table border="0" cellpadding="0" cellspacing="0" width="1158">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Name of Reagent/ Equipment</td>
			<td style="width:195px;">Company</td>
			<td style="width:128px;">Catalog Number</td>
			<td style="width:523px;">Comments/Description</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Confocal Microscope</td>
			<td style="width:195px;">Leica</td>
			<td style="width:128px;"></td>
			<td>TCS SP5X, LAS AF - Version 2.6.1; or equivalent CLSM</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:313px;">GC HP 7890 Series GC and Agilent 5975C MSD</td>
			<td>Agilent</td>
			<td style="width:128px;"></td>
			<td>an equivalent GC/MS may be used</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">GC capillary column J&#38;W 121-5522&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;</td>
			<td>Agilent</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Cork borer</td>
			<td style="width:195px;">Fisher Scientific</td>
			<td align="right">12863952</td>
			<td>or any other</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Cover slips</td>
			<td style="width:195px;">Marienfeld</td>
			<td style="width:128px;">107222</td>
			<td>High performance, No.1.5H</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">GC/MS insterts</td>
			<td style="width:195px;">WICOM</td>
			<td style="width:128px;">WIC 47080</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">GC/MS vials 2 ml</td>
			<td style="width:195px;">WICOM</td>
			<td style="width:128px;">WIC 41150</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Lids / septa for screw cap vials</td>
			<td style="width:195px;">DIONEX</td>
			<td>49463 / 049464&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Lids for GC/MS vials</td>
			<td style="width:195px;">WICOM</td>
			<td style="width:128px;">WIC 43948/B</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Objective Slides</td>
			<td style="width:195px;">Menzel</td>
			<td style="width:128px;"></td>
			<td>ordinary</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:313px;">PDMS coated glass fibers</td>
			<td style="width:195px;">Polymicro Technologies, Inc.</td>
			<td style="width:128px;"></td>
			<td>V (PDMS) = 13.55 &#177; 0.02 &#181;L m-1</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:313px;">Petri Dishes small / big</td>
			<td style="width:195px;">Greiner</td>
			<td style="width:128px;">633-102 / 628-102</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Screw cap vials 40 ml</td>
			<td style="width:195px;">DIONEX</td>
			<td>48783</td>
			<td>other glass vials may be used</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Screw cap vials 60 ml</td>
			<td style="width:195px;">DIONEX</td>
			<td>48784</td>
			<td>other glass vials may be used</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:313px;">Acenaphthylene d08</td>
			<td style="width:195px;">Dr. Ehrenstorfer</td>
			<td style="width:128px;">C 20510100</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Acetone</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">9372.2</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Activated carbon</td>
			<td style="width:195px;">Sigma-Aldrich</td>
			<td style="width:128px;">242276-1kg</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Agarose</td>
			<td style="width:195px;">Carl Roth</td>
			<td align="right">2267.4</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Fluorene</td>
			<td style="width:195px;">Fluka</td>
			<td style="width:128px;">46880</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:313px;">Kanamycin sulfate</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">T832.2</td>
			<td>50 mg L-1</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Methanol</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">P7171</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Minimal Medium:</td>
			<td style="width:195px;"></td>
			<td style="width:128px;"></td>
			<td>100 mL solution 1 + 25 mL solution 2 + 5 mL solution 3 ad. 1000 mL aqua dest</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">&#160; Solution 1</td>
			<td style="width:195px;"></td>
			<td style="width:128px;"></td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:313px;">&#160;&#160;&#160; Ammonium sulfate&#160;</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">3746.1</td>
			<td>5 g L-1&#160;</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;width:313px;">&#160;&#160;&#160; Magnesium chloride x 6 H2O</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">2189.1</td>
			<td>1 g L-1</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;width:313px;">&#160;&#160;&#160; Calcium nitrate x 4 H2O</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">P740.1</td>
			<td>0.5 g L-1</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">&#160; Solution 2</td>
			<td style="width:195px;"></td>
			<td style="width:128px;"></td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:313px;">&#160;&#160;&#160; Disodium phosphate</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">P030.1</td>
			<td>55.83 g L-1</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:313px;">&#160;&#160;&#160; Monopotassium phosphate</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">3904.1</td>
			<td>20 g L-1</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">&#160; Solution 3</td>
			<td style="width:195px;"></td>
			<td style="width:128px;"></td>
			<td>pH 6.0</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;">&#160;&#160;&#160; Disodium EDTA</td>
			<td style="width:195px;">MERCK</td>
			<td style="width:128px;">1084180250</td>
			<td>0.8 g L-1</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;">&#160;&#160;&#160; Iron(II) chloride x 4 H2O</td>
			<td style="width:195px;">MERCK</td>
			<td style="width:128px;">1038610250</td>
			<td>0.3 g L-1</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;">&#160;&#160;&#160; Cobalt(II) chloride x 6 H2O</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">T889.3</td>
			<td>4 mg L-1</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;">&#160;&#160;&#160; Manganese(II) chloride x 1 H2O&#160;&#160;&#160;&#160;&#160;&#160;&#160;</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">&#160; 4320.2</td>
			<td>10 mg L-1</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;">&#160;&#160;&#160; Copper(II) sulfate</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">&#160;P023.1</td>
			<td>1 mg L-1</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;">&#160;&#160;&#160; Sodium molybdate x 2 H2O</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">&#160;0274.1</td>
			<td>3 mg L-1</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;">&#160;&#160;&#160; Zinc chloride</td>
			<td style="width:195px;">MERCK</td>
			<td style="width:128px;">1088160250</td>
			<td>2 mg L-1</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;">&#160;&#160;&#160; Lithium chloride</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">P007.1</td>
			<td>0.5 mg L-1</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;">&#160;&#160;&#160; Tin(II) chloride x 2 H2O</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">4473.1</td>
			<td>0.5 mg L-1</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;">&#160;&#160;&#160; Boric acid</td>
			<td style="width:195px;">Riedel-de-Haen&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;</td>
			<td style="width:128px;">11606</td>
			<td>1 mg L-1</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;">&#160;&#160;&#160; Potassium bromide</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">A137.1</td>
			<td>2 mg L-1</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;">&#160;&#160;&#160; Potassium iodide</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">6750.1</td>
			<td>2 mg L-1</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;">&#160;&#160;&#160; Barium chloride</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">4453.1</td>
			<td>0.5 mg L-1</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">MMA</td>
			<td style="width:195px;"></td>
			<td style="width:128px;"></td>
			<td>Minimal medium + agarose 0.2 %</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:313px;">Phenanthrene d10</td>
			<td style="width:195px;">Dr. Ehrenstorfer</td>
			<td style="width:128px;">C 20920100</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:313px;">Potato Dextrose Agar:</td>
			<td style="width:195px;"></td>
			<td style="width:128px;"></td>
			<td>24 g L-1 broth + bacto-agar 1.5 %; pH 6.8</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">&#160;&#160;&#160; Potato Dextrose broth</td>
			<td>Difco/ Beckton Dickinson</td>
			<td style="width:128px;">254920</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">&#160;&#160;&#160; Bacto-agar</td>
			<td>Difco/ Beckton Dickinson</td>
			<td align="right">214040</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Sodium acetate x 3 hydr.</td>
			<td style="width:195px;">Carl Roth</td>
			<td style="width:128px;">6779.1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Sodium sulfate</td>
			<td style="width:195px;">MERCK</td>
			<td style="width:128px;">&#160;1066495000</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Toluene</td>
			<td style="width:195px;">MERCK</td>
			<td style="width:128px;">1083252500</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:313px;">mTY medium:</td>
			<td style="width:195px;"></td>
			<td style="width:128px;"></td>
			<td>3 g L-1 yeast extract, 5 g L-1 bacto tryptone and 50 mM NaCl</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">&#160;&#160;&#160; Yeast extract</td>
			<td style="width:195px;">Merck</td>
			<td>1037530500</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">&#160;&#160;&#160; Tryptone</td>
			<td style="width:195px;">Serva</td>
			<td>4864702</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">&#160;&#160;&#160; Sodium chloride</td>
			<td style="width:195px;">Carl Roth</td>
			<td>3957.1</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:313px;">imageJ with logi tool plugin</td>
			<td style="width:195px;"></td>
			<td style="width:128px;"></td>
			<td style="width:523px;">http://rsb.info.nih.gov/ij/download.html and http://downloads.openmicroscopy.org/bio-formats/4.4.10</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:313px;">Pythium ultimum strain 67-1</td>
			<td style="width:195px;"></td>
			<td style="width:128px;"></td>
			<td style="width:523px;">Obtained from the lab of Dr. Christoph Keel; Department of Fundamental Microbiology, University of Lausanne, Switzerland</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:313px;">Burkholderia sartisoli RP037-mChe</td>
			<td style="width:195px;"></td>
			<td style="width:128px;"></td>
			<td style="width:523px;">Obtained from the lab of Prof. Jan Roelof van der Meer; Department of Fundamental Microbiology, University of Lausanne, Switzerland</td>
		</tr>
	</tbody>
</table>

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.

I risultati presentati qui sono già stati pubblicati in precedenza 15. Consultare l&#39;articolo per la discussione dettagliata meccanicistica e ambientale. Dopo la registrazione dell&#39;immagine mediante CLSM, una sporgenza massima intensità può essere effettuato utilizzando il rispettivo software microscopio o ImageJ per ottenere una prima impressione visiva del campione ei controlli (Figura 2). Più tardi, i set di dati possono essere proiettati in modo div...

Watch this Scientific Journal Video about A Whole Cell Bioreporter Approach to Assess Transport and Bioavailability of Organic Contaminants in Water Unsaturated Systems at JoVE.com

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

A Whole marcatura già cellulare approccio per valutare i trasporti e la biodisponibilità di contaminanti organici in acqua insaturi Sistemi

Un intero test marcatura già cella con Burkholderia sartisoli RP037-Mche è stato sviluppato per rilevare frazioni di un contaminante organico (cioè, fluorene) disponibile per la degradazione batterica dopo trasporto attivo da miceli ponte pori pieni d&#39;aria in un sistema modello insaturo acqua.

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

Research

JoVE Journal

Environment

EPA Method 1615. Measurement of Enterovirus and Norovirus Occurrence in Water by Culture and RT-qPCR. I. Collection of Virus Samples

Metodo EPA 1615. Misura di Enterovirus e Norovirus Evento in acqua per la cultura e RT-qPCR. I. Raccolta di campioni di virus

EPA Method 1615 was developed with a goal of providing a standard method for measuring enteroviruses and noroviruses in environmental and drinking waters. ...

Ricerca

Ambiente

A Small Volume Bioassay to Assess Bacterial/Phytoplankton Co-culture Using WATER-Pulse-Amplitude-Modulated (WATER-PAM) Fluorometry

A Small Volume Bioassay to Assess Bacterial/Phytoplankton Co-culture Using WATER-Pulse-Amplitude-Modulated WATER-PAM Fluorometry

Un piccolo volume Bioassay per valutare batterica / fitoplancton Co-cultura Utilizzando WATER-Pulse-modulata in ampiezza (WATER-PAM) fluorometria

Conventional methods for experimental manipulation of microalgae have employed large volumes of culture (20 ml to 5 L), so that the culture can be ...

EPA Method 1615. Measurement of Enterovirus and Norovirus Occurrence in Water by Culture and RT-qPCR. Part III. Virus Detection by RT-qPCR

Metodo EPA 1615. Misura di Enterovirus e Norovirus Presenza in Water di Cultura e RT-qPCR. Parte III. Rilevamento virus mediante RT-qPCR

EPA Method 1615 measures enteroviruses and noroviruses present in environmental and drinking waters. This method was developed with the goal of having a ...

Soil Lysimeter Excavation for Coupled Hydrological, Geochemical, and Microbiological Investigations

Terreno lisimetro scavo per accoppiate idrologici, geochimiche e microbiologiche Investigations

Studying co-evolution of hydrological and biogeochemical processes in the subsurface of natural landscapes can enhance the understanding of coupled ...

Implementation of Portable Emissions Measurement Systems (PEMS) for the Real-driving Emissions (RDE) Regulation in Europe

Implementation of Portable Emissions Measurement Systems PEMS for the Real-driving Emissions RDE Regulation in Europe

Implementazione di sistemi di misura portatili (PEMS) per il regolamento delle emissioni Real-guida (RDE) in Europa

Vehicles are tested in controlled and relatively narrow laboratory conditions to determine their official emission values and reference fuel consumption. ...

Vegetated Treatment Systems for Removing Contaminants Associated with Surface Water Toxicity in Agriculture and Urban Runoff

Sistemi di trattamento vegetale per la rimozione dei contaminanti associati alla tossicità dell&#39;acqua superficiale nell&#39;agricoltura e nella rottura urbana

Urban stormwater and agriculture irrigation runoff contain a complex mixture of contaminants that are often toxic to adjacent receiving waters. Runoff may ...

Controlled-release of Chlorine Dioxide in a Perforated Packaging System to Extend the Storage Life and Improve the Safety of Grape Tomatoes

A rilascio di biossido di cloro in un sistema di imballaggio forato per prolungare la durata di conservazione e migliorare la sicurezza dei Pomodori

A controlled-release chlorine dioxide (ClO2) pouch was developed by sealing a slurry form of ClO2 into semipermeable polymer film; the release properties ...

Measuring and Mapping Patterns of Soil Erosion and Deposition Related to Soil Carbonate Concentrations Under Agricultural Management

Misura e mappatura modelli di erosione e deposizione relazionati al suolo carbonato concentrazioni sotto gestione agricola

Spatial patterns of soil erosion and deposition can be inferred from differences in ground elevation mapped at appropriate time increments. Such changes ...

A Simple Planting Technique for Re-establishing Trees Where Frequent Inundation Occurs

Una semplice tecnica di impianto per ristabilire gli alberi dove si verifica frequente Inundation

Many of the Everglades tree islands have lost elevation over the past century and most of their trees have died such that they are now covered with ...

Improving Infrared Spectroscopy Characterization of Soil Organic Matter with Spectral Subtractions

Migliorare la caratterizzazione spettroscopia infrarossa di materia organica del suolo con sottrazioni spettrale

Soil organic matter (SOM) underlies numerous soil processes and functions. Fourier transform infrared (FTIR) spectroscopy detects infrared-active organic ...