Ringraziamo Manel Bolivar per la sua assistenza nel lavoro sul campo. Siamo grati al &quot;Parco Naturale del Montgrí, les Illes Medes i el Baix Ter&quot; per il loro supporto ai nostri permessi di ricerca e di campionamento. Il manipolatore subacqueo è stato progettato da Ayelet Dadon-Pilosof e fabbricato dal Sig Pilosof. Questo lavoro è stato supportato dal progetto governo spagnolo CSI-Corallo [codice di autorizzazione CGL2013-43106-R per RC e MR] e da una borsa di FPU da &quot;Ministerio de Educación, Cultura y Deporte (MECD)&quot; a TM. Si tratta di un contributo della biogeochimica marina e il gruppo di ricerca Cambiamento Globale finanziato dal Governo Catalano [concessione numero 2014SGR1029] e ISF concessione 1280-1213 e BSF concessione 2.012.089 a G. Yahel.

<ol>
	<li>Gili, J. M., Coma, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Benthic+suspension+feeders:+their+paramount+role+in+littoral+marine+food+webs.">Benthic suspension feeders: their paramount role in littoral marine food webs.</a> Trends. Ecol. Evol. 13 (8), 316-321 (1998).</li><li>Reiswig, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+situ+pumping+activities+of+tropical+Demospongiae.">In situ pumping activities of tropical Demospongiae.</a> Mar. 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U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficiency+of+particle+retention+in+13+species+of+suspension+feeding+bivalves.">Efficiency of particle retention in 13 species of suspension feeding bivalves.</a> Ophelia. 17 (2), 239-246 (1978).</li><li>Mueller, 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=Natural+diet+of+coral-excavating+sponges+consists+mainly+of+dissolved+organic+carbon+(DOC).">Natural diet of coral-excavating sponges consists mainly of dissolved organic carbon (DOC).</a> PLoS ONE. 9 (2), e90152 (2014).</li><li>Gasol, J. M., Moran, X. A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+filtration+on+bacterial+activity+and+picoplankton+community+structure+as+assessed+by+flow+cytometry.">Effects of filtration on bacterial activity and picoplankton community structure as assessed by flow cytometry.</a> Aquat. Microb. Ecol. 16 (3), 251-264 (1999).</li><li>Koroleff, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determination+of+reactive+silicate.">Determination of reactive silicate.</a> New Baltic Manual, Cooperative Research Report Series A. 29, 87-90 (1972).</li><li>Murphy, J., Riley, J. P. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modified+single+solution+method+for+the+determination+of+phosphate+in+in+natural+waters.">Modified single solution method for the determination of phosphate in in natural waters.</a> Anal. Chim. Acta. 27, 31-36 (1962).</li><li>Shin, M. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Colorimetric+method+for+determination+of+nitrite.">Colorimetric method for determination of nitrite.</a> Ind.Eng.Chem. 13 (1), 33-35 (1941).</li><li>Wood, E. D., Armstrong, F. A. J., Richards, F. 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H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Marine+dissolved+organic+carbon:+Are+the+older+values+correct.">Marine dissolved organic carbon: Are the older values correct.</a> Mar. Chem. 56 (3-4), 265-277 (1997).</li><li>Holmes, R. M., Aminot, A., Kerouel, R., Hooker, B. A., Peterson, B. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+simple+and+precise+method+for+measuring+ammonium+in+marine+and+freshwater+ecosystems.">A simple and precise method for measuring ammonium in marine and freshwater ecosystems.</a> Can. J. Fish. Aquat. 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C., Panagiotopoulos, C., Semp&#233;r&#233;, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dissolved+organic+carbon+contamination+induced+by+filters+and+storage+bottles.">Dissolved organic carbon contamination induced by filters and storage bottles.</a> Water Res. 33 (8), 1956-1959 (1999).</li><li>Zhang, J. Z., Fischer, C. J., Ortner, P. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laboratory+glassware+as+a+contaminant+in+silicate+analysis+of+natural+water+samples.">Laboratory glassware as a contaminant in silicate analysis of natural water samples.</a> Water Res. 33 (12), 2879-2883 (1999).</li><li>Yoshimura, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Appropriate+bottles+for+storing+seawater+samples+for+dissolved+organic+phosphorus+(DOP)+analysis:+a+step+toward+the+development+of+DOP+reference+materials.">Appropriate bottles for storing seawater samples for dissolved organic phosphorus (DOP) analysis: a step toward the development of DOP reference materials.</a> Limnol. Oceanogr-meth. 11 (4), 239-246 (2013).</li><li>Strickland, J. D. H., Parsons, T. 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The authors have nothing to disclose.

operazioni preliminari Fiale Collector per Dom e analisi dei nutrienti Dal momento che i vasi da collezione possono interagire con micro-costituenti disciolti e le pareti campionatore può essere un substrato per i batteri crescita 30-34, sono stati testati diversi fiale per DOM e la raccolta dei nutrienti. Borosilicato non è raccomandato per la quantificazione silice 33,35, dal momento che le bott...

Alimentatori sospensione bentonici giocano un ruolo essenziale nel funzionamento degli ecosistemi marini 1. Filtrando grandi volumi di acqua 2,3, rimuovono e espellono particolato (plancton e detriti) e composti disciolti 1 (e riferimenti) e sono un importante agente di bentonica-pelagica accoppiamento 4,5 e il ciclo dei nutrienti 6,7. Misurare accuratamente la presenza di particelle e composti disciolti rimossi ed escreto dal alimentatori sospensione bentonici (come spugne, ascidie, policheti, e bivalvi) è fondamentale per capire la loro fisiologia, metabolismo, e l&#39;ecologia alimentazione. Insieme con pompaggio misurazione della frequenza, ma consente anche una quantificazione dei flussi di nutrienti mediate da questi organismi e il loro impatto ecologico sulla qualità delle acque, nonché sui processi scala ecosistema. La scelta del metodo appropriato di misurazione di rimozione e di produzione tassi di particolato e com discioltochili di sospensione filtratori è fondamentale per ottenere dati affidabili relativi alla loro attività di alimentazione 8. Come sottolineato da Riisgård e altri, non appropriate metodologie di polarizzazione risultati, falsare le condizioni sperimentali, la produzione di stime errate di ingestione ed escrezione di determinate sostanze, e può portare a quantificazione errata dei flussi di nutrienti elaborati da questi organismi. I due metodi più frequentemente utilizzati per misurare le particelle e flussi di nutrienti disciolti in filtratori coinvolgere sia incubazione (tecniche indirette) o la raccolta simultanea di ambiente e acqua espirata (tecniche dirette). Tecniche di incubazione si basano sulla misura della velocità di variazione della concentrazione di particelle e nutrienti disciolti nell&#39;acqua incubate, e stima dei tassi di produzione o rimozione rispetto ai controlli adeguati 8. Tuttavia, allegando un organismo in una camera di incubazione può modificare la sua Feeding e pompaggio comportamento a causa di cambiamenti nel regime di flusso naturale, a causa di un calo di ossigeno e / o della concentrazione di cibo, o a causa di accumulo di composti di escrezione del 7,9 acqua incubazione (e riferimenti). Oltre agli effetti di isolamento e fornitura di acqua modificato, una grande polarizzazione di tecniche di incubazione deriva da effetti ri-filtrazione (vedi ad esempio 10). Sebbene alcuni di questi problemi metodologici sono stati superati utilizzando il giusto volume e la forma del recipiente di incubazione 11 o con l&#39;introduzione di un sistema di campana ricircolo in situ 12, questa tecnica spesso sottostima asportazione e di produzione. Quantificare il metabolismo dei composti disciolti quali azoto organico disciolto (DON) e carbonio (DOC) o nutritivi inorganici, ha dimostrato di essere particolarmente inclini a distorsioni causate da tecniche di incubazione 13. Alla fine degli anni &#39;60 e primi anni &#39;70, Henry Reiswig9,14,15 aperto la strada all&#39;applicazione di tecniche dirette per quantificare la rimozione di particelle da giganti spugne caraibiche, campionando separatamente l&#39;acqua inspirato ed espirato dagli organismi in situ. A causa della difficoltà di applicare la tecnica di Reiswig su sospensivori piccole e in condizioni subacquee più impegnative, la maggior parte della ricerca in questo campo è stata limitata al laboratorio (in vitro) impiegando lo più tecniche di incubazione indirette 16. Yahel e colleghi rimontati Reiswig di diretta in situ tecnica di lavorare in condizioni di minori dimensioni. Il loro metodo, chiamato Inex 16, si basa sul campionamento simultaneo subacquea dell&#39;acqua per via inalatoria (In) e esalato (Ex) da organismi indisturbati. La diversa concentrazione di una sostanza (ad esempio, batteri) tra una coppia di campioni (INEX) fornisce una misura della ritenzione (o produzione) della sostanza da parte dell&#39;animale. La tecnica Inex impiega tubi aperti esi basa sul jet excurrent prodotte dall&#39;attività di pompaggio dell&#39;organismo studiata per sostituire passivamente all&#39;acqua ambiente nel tubo di raccolta. Mentre Yahel e colleghi hanno applicato con successo questa tecnica per lo studio di oltre 15 sospensione di diversi alimentatori taxa (ad esempio, 17), il metodo è vincolato per l&#39;elevato livello di pratica e l&#39;esperienza necessarie, dalla dimensione minuscola di alcuni orifizi excurrent, e condizioni del mare. Per superare questi ostacoli, abbiamo sviluppato una tecnica alternativa basata su aspirazione controllata dell&#39;acqua campionato attraverso tubi minute (diametro esterno &lt;1,6 mm). Il nostro scopo era quello di creare un dispositivo semplice, affidabile ed economico che consenta pulita e controllata in campionamento dell&#39;acqua situ da un punto molto specifico, come l&#39;orifizio excurrent di alimentatori sospensione bentonici. Per essere efficace, il metodo deve essere non-intrusivo in modo da non influenzare il regime di flusso ambiente o modificare il behavior degli organismi studiati. Il dispositivo presentato qui è chiamato VacuSIP. È una semplificazione del sistema SIP sviluppato da Yahel et al. (2007) 18 per il campionamento punto ROV-based nel mare profondo. Il VacuSIP è notevolmente più conveniente che il SIP originale ed è stato adattato per il lavoro SCUBA-based. Il sistema è stato progettato secondo i principi presentati e testati da Wright e Stephens (1978) 19 e Møhlenberg e Riisgård (1978) 20 per le impostazioni di laboratorio. Sebbene il sistema VacuSIP stato progettato per studi situ del metabolismo di alimentatori sospensione bentonici, può essere utilizzato anche per studi di laboratorio e ovunque sia richiesto un campione di acqua da sorgente puntiforme controllata e pulita. Il sistema è particolarmente utile quando è richiesto l&#39;integrazione per periodi prolungati (min-ore) o in filtrazioni situ. Il VacuSIP è stato utilizzato con successo presso il laboratorio Yahel a partire dal 2011, e ha anchestato impiegato in due recenti studi dei flussi di nutrienti mediati da specie di spugne caraibiche e mediterranee 21 (Morganti et al. submitted). L&#39;utilizzo di campionatori specifici, la durata di campionamento prolungato, e le condizioni del campo, in cui si applica VacuSIP, comporta alcune differenze rispetto a protocolli standard di oceanografici per la raccolta, il filtraggio e la conservazione dei campioni per analiti sensibili. Per ridurre il rischio di contaminazione da parte del sistema VacuSIP o il rischio di modificazione dell&#39;acqua campionata dall&#39;attività batterica dopo la raccolta, abbiamo testato varie procedure di filtrazione e stoccaggio situ. Diversi dispositivi di filtraggio, recipienti di raccolta e procedure memorizzazione sono stati esaminati per conseguire la tecnica più adatta per l&#39;analisi dei disciolto inorganico (PO 4 3-, NO x -, NH 4 +, SiO 4) ed organici (DOC + DON) composti, e ultra-plancton (&lt;1081; m) e il particolato organico (POC + campionamento PON). Per ridurre ulteriormente il rischio di contaminazione, in particolare in condizioni di campo, il numero di fasi di manipolazione è stato ridotto al minimo. Il formato visivo in cui è presentato il metodo è orientato per facilitare riproducibilità e per ridurre il tempo necessario per applicare efficacemente la tecnica. Panoramica del sistema Per campionare in situ pompata acqua sospensivori con orifizi exhalant piccole 2 mm, l&#39;attività di pompaggio di ciascun campione viene prima visualizzata rilasciando filtrata fluoresceina tinti acqua di mare accanto all&#39;orifizio inalante (s) e osservando il suo flusso dall&#39;apertura excurrent 16 (vedi anche figura 2B in 18). L&#39;acqua inalato ed espirato dal campione di studio (incurrent e excurrent) vengono quindi simultaneamente campionati con l&#39;impiego di una coppia di tubi minute installati sul manipolatore fuoriserie o due dei &quot;arms &quot;di un treppiede portatile flessibile capovolta (Figura 1 e complementare Video 1). L&#39;acqua inalata dall&#39;organismo studio viene raccolto posizionando accuratamente l&#39;estremità prossimale di un tubo all&#39;interno o vicino all&#39;apertura inalazione dell&#39;organismo studio. Un identico tubo viene quindi posizionato all&#39;interno dell&#39;orifizio excurrent. Questa operazione richiede cura per evitare il contatto o disturbo dell&#39;animale, per esempio, mediante risospensione dei sedimenti. per iniziare il campionamento, un subacqueo perfora un setto nel recipiente di raccolta con una siringa attaccata alla estremità distale di ciascun tubo, consentendo la pressione dell&#39;acqua esterna di forzare l&#39;acqua campionata nel vaso attraverso il tubo di campionamento. l&#39;aspirazione viene iniziata dal vuoto creato in precedenza nei flaconi e dalla differenza di pressione tra l&#39;acqua esterna e il contenitore del campione evacuato . Per garantire una raccolta di acqua pulita espirata e per evitare l&#39;aspirazione accidentale di ambiacqua ent 16, la frequenza di campionamento dell&#39;acqua deve essere mantenuto a un tasso significativamente più basso (&lt;10%) rispetto alla portata excurrent. Il tasso di aspirazione è controllata dalla lunghezza del tubo e il diametro interno (ID). Il piccolo diametro interno garantisce inoltre un volume morto trascurabile (&lt;200 ml per metro di tubo). Campionamento per periodi prolungati (minuti a ore) permette di integrare il patchiness intrinseca della maggior parte delle sostanze di interesse. Per assicurare che i campioni siano adeguatamente conservati in sessioni di campionamento subacqueo prolungati nonché per il trasporto al laboratorio, una in situ filtrazione è raccomandato per analiti sensibili. La selezione di vasi di campionamento, il montaggio di filtraggio, e tubi sono dettate dagli organismi di studio e la domanda di ricerca specifica. Il protocollo descritto di seguito presuppone che un profilo metabolico completo è di interesse (per una panoramica si veda la Figura 2). Tuttavia, la natura modulare del protocollo consente fo facile modifica per accogliere i metodi di campionamento più semplici o anche molto diverse. Per un profilo metabolico completo, il protocollo di campionamento dovrebbe comprendere le seguenti fasi: (1) la visualizzazione di flusso; (2) l&#39;alimentazione di campionamento ultra-plancton (plancton &lt;10 micron); (3) Il campionamento nutrienti inorganici assorbimento e l&#39;escrezione (usando filtri in linea); (4) Campionamento disciolto assorbimento organico e l&#39;escrezione (usando filtri in linea); (5) di alimentazione particolato e l&#39;escrezione (usando filtri in linea); (6) Ripetere il punto 2 (ultra-plancton alimentazione come controllo della qualità); (7) Portata visualizzazione. Quando logisticamente fattibile, si raccomanda che le misurazioni profilo metabolico sono combinati con pompaggio rate (ad esempio, il metodo a velocità fronte del colorante, in 16), nonché con le misurazioni respirazione. Queste misure devono essere prese all&#39;inizio e alla fine della sessione di campionamento. Per la misurazione della respirazione, optodes subacquee o micro-elettrodi sono preferibili.

<table><tbody><tr><td>GorillaPod, Original</td><td>Joby</td><td>GP000001</td><td>flexible portable tripod&amp;nbsp;</td></tr><tr><td>Flangeless Ferrule</td><td>IDEX Health &amp;amp; Science&amp;nbsp;</td><td>P-200X</td><td>1/16&quot; in Blue/pk</td></tr><tr><td>Male Nut</td><td>IDEX Health &amp;amp; Science</td><td>P-205X&amp;nbsp;</td><td>1/16&quot; in Green/10&amp;nbsp;pk</td></tr><tr><td>Female to Female Luer</td><td>IDEX Health &amp;amp; Science&amp;nbsp;</td><td>P-658</td><td></td></tr><tr><td>Female-Male Luer</td><td>IDEX Health &amp;amp; Science&amp;nbsp;</td><td>P-655</td><td></td></tr><tr><td>Peek Tubing (254&amp;nbsp;&amp;micro;m ID)</td><td>IDEX Health &amp;amp; Science&amp;nbsp;</td><td>1531</td><td>1/16&quot; OD x 0.01&amp;nbsp;in ID x 5&amp;nbsp;ft length. Alternative ID can be used.</td></tr><tr><td>Two component resin epoxy</td><td>IVEGOR</td><td>9257</td><td>Mix well the two component resin before use</td></tr><tr><td>(TOC) EPA Vials</td><td>Cole -Parmer</td><td>03756-20</td><td>40 ml glass vials. Manufactured also by Thomas Scientific (ref. number 9711F09).</td></tr><tr><td>HDPE Vials</td><td>Wheaton</td><td>986701 (E78620)</td><td>20 ml high-density polyethylene vials</td></tr><tr><td>Vacuette Z no additive</td><td>Greiner bio-one</td><td>455001</td><td>pre-vacuum by the manufacturer&amp;nbsp;</td></tr><tr><td>Septum Sample Bottles</td><td>Thomas Scientific</td><td>1755C01</td><td>250 ml glass bottles&amp;nbsp;</td></tr><tr><td>Septum Cap 1</td><td>Wheaton</td><td>W240844SP (E7865R)</td><td>22-400 for HDPE vials&amp;nbsp;</td></tr><tr><td>Septum Cap 2</td><td>Wheaton</td><td>W240846 (1078-5553)</td><td>24-400 for glass vials and bottles. Also manufactured by Thermo Scientific National (ref. 03-377-42).</td></tr><tr><td>In-line stainless steel Swinney Filter holders</td><td>Pall</td><td>516-9067</td><td>13 mm of diameter</td></tr><tr><td>PTFE Seal Washer</td><td>Pall&amp;nbsp;</td><td>516-8064</td><td>ring for stainless steel filter holders</td></tr><tr><td>TCLP Glass Filters</td><td>Pall&amp;nbsp;</td><td>516-9126</td><td>binder-free glass fiber filters, 13&amp;nbsp;mm of diameter, pore size 0.7&amp;nbsp;&amp;micro;m</td></tr><tr><td>Polycarbonate Filter Holders</td><td>Cole -Parmer</td><td>17295</td><td>13 mm of diameter</td></tr><tr><td>Isopore Membrane Filters</td><td>Millipore</td><td>GTTP01300</td><td>13&amp;nbsp;mm of diameter, pore size 0.2&amp;nbsp;&amp;micro;m</td></tr><tr><td>Contrad 70 Solution&amp;nbsp;</td><td>Decon Labs</td><td>1002</td><td>highly soluble basic mix of anionic and non-ionic surfactant solution&amp;nbsp;</td></tr><tr><td>Sterile Syringe Filters</td><td>VWR International Eurolab S.L.</td><td>514-0061P</td><td>25&amp;nbsp;mm of diameter , pore size 0.2&amp;nbsp;&amp;micro;m</td></tr><tr><td>Fluorescein</td><td>Sigma-Aldrich</td><td>(old ref.28802) 46955-100G</td><td>100&amp;nbsp;g</td></tr><tr><td>Holdex, disposable,sterile</td><td>Greiner bio-one</td><td>450263</td><td>sterile, single-use tube holder with off-center luer for Vacuette</td></tr><tr><td>Sterile Needles</td><td>IcoGammaPlus</td><td>5160</td><td>0.7&amp;nbsp;mm x 30&amp;nbsp;mm</td></tr><tr><td>Cryovials Nalgene</td><td>Nalgene</td><td>V5007(Cat. No.5000-0020)</td><td>2&amp;nbsp;ml</td></tr><tr><td>Cryobox carton&amp;nbsp;</td><td>Rubilabor</td><td>M-600</td><td>145x145x55&amp;nbsp;mm p/microtube 1.5&amp;nbsp;ml</td></tr><tr><td>Orthophosphoric Acid</td><td>Sigma</td><td>79617 or Alternatively use Ultra-Pure Hydrochloric acid, final concertation 0.1%</td><td></td></tr><tr><td>Paraformaldehyde</td><td>Sigma</td><td>P6148</td><td>500&amp;nbsp;g</td></tr><tr><td>Glutaraldehyde</td><td>Merck</td><td>8,206,031,000</td><td>25%, 1 L</td></tr><tr><td>Hand Vacuum Pump&amp;nbsp;</td><td>B&amp;uuml;rkle&amp;nbsp;</td><td>5620-2181 or Alternatively use standard laboratory vacuum pump</td><td></td></tr></tbody></table>

vacusip, an improved inex method for in situ measurement of particulate and dissolved compounds processed by active suspension feeders

Benthic suspension feeders play essential roles in the functioning of marine ecosystems. By filtering large volumes of water, removing plankton and detritus, and excreting particulate and dissolved compounds, they serve as important agents for benthic-pelagic coupling. Accurately measuring the compounds removed and excreted by suspension feeders (such as sponges, ascidians, polychaetes, bivalves) is crucial for the study of their physiology, metabolism, and feeding ecology, and is fundamental to determine the ecological relevance of the nutrient fluxes mediated by these organisms. However, the assessment of the rate by which suspension feeders process particulate and dissolved compounds in nature is restricted by the limitations of the currently available methodologies. Our goal was to develop a simple, reliable, and non-intrusive method that would allow clean and controlled water sampling from a specific point, such as the excurrent aperture of benthic suspension feeders, in situ. Our method allows simultaneous sampling of inhaled and exhaled water of the studied organism by using minute tubes installed on a custom-built manipulator device and carefully positioned inside the exhalant orifice of the sampled organism. Piercing a septum on the collecting vessel with a syringe needle attached to the distal end of each tube allows the external pressure to slowly force the sampled water into the vessel through the sampling tube. The slow and controlled sampling rate allows integrating the inherent patchiness in the water while ensuring contamination free sampling. We provide recommendations for the most suitable filtering devices, collection vessel, and storing procedures for the analyses of different particulate and dissolved compounds. The VacuSIP system offers a reliable method for the quantification of undisturbed suspension feeder metabolism in natural conditions that is cheap and easy to learn and apply to assess the physiology and functional role of filter feeders in different ecosystems.

L&#39;ottimizzazione dei metodi di raccolta di acqua di mare Selezione di fiale da collezione e la procedura di pulizia recipienti di raccolta compatibili VacuSIP dovrebbero avere un setto che permette di campionamento venga avviata forando con una siringa. Essi dovrebbero resistere alla pressione sottomarina elevata (2-3 b...

Watch this Scientific Journal Video about VacuSIP, an Improved InEx Method for In Situ Measurement of Particulate and Dissolved Compounds Processed by Active Suspension Feeders at JoVE.com

VacuSIP, an Improved InEx Method for In Situ Measurement of Particulate and Dissolved Compounds Processed by Active Suspension Feeders

We introduce the VacuSIP, a simple, non-intrusive, and reliable method for clean and accurate point sampling of water. The system was developed and evaluated for the simultaneous collection of the water inhaled and exhaled by benthic suspension feeders in situ, to cleanly measure removal and excretion of particulate and dissolved compounds.

VacuSIP, un metodo migliorato per Inex<em&gt; In Situ</em&gt; Misura di particolato e sciolto composti elaborati da Alimentatori sospensioni attive

Benthic suspension feeders play essential roles in the functioning of marine ecosystems. By filtering large volumes of water, removing plankton and ...

vacusip-an-improved-inex-method-for-situ-measurement-particulate

Research

JoVE Journal

Environment

VacuSIP, an Improved InEx Method for In Situ Measurement of Particulate and Dissolved Compounds Processed by Active Suspension Feeders

10.8K Views.  Centre d’Estudis Avançats de Blanes (CEAB-CSIC). We introduce the VacuSIP, a simple, non-intrusive, and reliable method for clean and accurate point sampling of water. The system was developed and evaluated for the simultaneous collection of the water inhaled and exhaled by benthic suspension feeders in situ, to cleanly measure removal and excretion of particulate and dissolved compounds.

Video: VacuSIP, an Improved InEx Method for In Situ Measurement of Particulate and Dissolved Compounds Processed by Active Suspension Feeders

Quantitatively Measuring In situ Flows using a Self-Contained Underwater Velocimetry Apparatus (SCUVA)

The ability to directly measure velocity fields in a fluid environment is necessary to provide empirical data for studies in fields as diverse as oceanography, ecology, biology, and fluid mechanics. Field measurements introduce practical challenges such as environmental conditions, animal availability, and the need for field-compatible measurement techniques. To avoid these challenges, scientists typically use controlled laboratory environments to study animal-fluid interactions. However, it is reasonable to question whether one can extrapolate natural behavior (i.e., that which occurs in the field) from laboratory measurements. Therefore, in situ quantitative flow measurements are needed to accurately describe animal swimming in their natural environment.
We designed a self-contained, portable device that operates independent of any connection to the surface, and can provide quantitative measurements of the flow field surrounding an animal. This apparatus, a self-contained underwater velocimetry apparatus (SCUVA), can be operated by a single scuba diver in depths up to 40 m. Due to the added complexity inherent of field conditions, additional considerations and preparation are required when compared to laboratory measurements. These considerations include, but are not limited to, operator motion, predicting position of swimming targets, available natural suspended particulate, and orientation of SCUVA relative to the flow of interest. The following protocol is intended to address these common field challenges and to maximize measurement success.

Quantitatively Measuring In situ Flows using a Self-Contained Underwater Velocimetry Apparatus SCUVA

This protocol provides instructions on how to use a self-contained underwater velocimetry apparatus (SCUVA), which is designed for quantification of in situ animal-generated flows. In addition, this protocol addresses challenges posed by field conditions, and includes operator motion, predicting position of animals, and orientation of SCUVA.

Misura quantitativamente In situ Flussi utilizzando un autorespiratore subacqueo Velocimetry (SCUVA)

The ability to directly measure velocity fields in a fluid environment is necessary to provide empirical data for studies in fields as diverse as ...

Ricerca

Bioingegneria

Clean Sampling and Analysis of River and Estuarine Waters for Trace Metal Studies

Most of the trace metal concentrations in ambient waters obtained a few decades ago have been considered unreliable owing to the lack of contamination control. Developments of some techniques aiming to reduce trace metal contamination in the last couple of decades have resulted in concentrations reported now being orders of magnitude lower than those in the past. These low concentrations often necessitate preconcentration of water samples prior to instrumental analysis of samples. Since contamination can appear in all phases of trace metal analyses, including sample collection (and during preparation of sampling containers), storage and handling, pretreatments, and instrumental analysis, specific care needs to be taken in order to reduce contamination levels at all steps. The effort to develop and utilize "clean techniques" in trace metal studies allows scientists to investigate trace metal distributions and chemical and biological behavior in greater details. This advancement also provides the required accuracy and precision of trace metal data allowing for environmental conditions to be related to trace metal concentrations in aquatic environments.
This protocol that is presented here details needed materials for sample preparation, sample collection, sample pretreatment including preconcentration, and instrumental analysis. By reducing contamination throughout all phases mentioned above for trace metal analysis, much lower detection limits and thus accuracy can be achieved. The effectiveness of "clean techniques" is further demonstrated using low field blanks and good recoveries for standard reference material. The data quality that can be obtained thus enables the assessment of trace metal distributions and their relationships to environmental parameters.

Special care using "clean techniques" is required to properly collect and process water samples for trace metal studies in aquatic environments. A protocol for sampling, processing, and analytical procedures with the aim of obtaining reliable environmental monitoring data and results with high sensitivity for detailed trace metal studies is presented.

Pulire campionamento e analisi del fiume e di estuario Acque per Trace Studi di metallo

Most of the trace metal concentrations in ambient waters obtained a few decades ago have been considered unreliable owing to the lack of contamination ...

Ambiente

Continuous Instream Monitoring of Nutrients and Sediment in Agricultural Watersheds

Pollutant concentrations and loads in watersheds vary considerably with time and space. Accurate and timely information on the magnitude of pollutants in water resources is a prerequisite for understanding the drivers of the pollutant loads and for making informed water resource management decisions. The commonly used &#34;grab sampling&#34; method provides the concentrations of pollutants at the time of sampling (i.e., a snapshot concentration) and may under- or overpredict the pollutant concentrations and loads. Continuous monitoring of nutrients and sediment has recently received more attention due to advances in computing, sensing technology, and storage devices. This protocol demonstrates the use of sensors, sondes, and instrumentation to continuously monitor in situ nitrate, ammonium, turbidity, pH, conductivity, temperature, and dissolved oxygen (DO) and to calculate the loads from two streams (ditches) in two agricultural watersheds. With the proper calibration, maintenance, and operation of sensors and sondes, good water quality data can be obtained by overcoming challenging conditions such as fouling and debris buildup. The method can also be used in watersheds of various sizes and characterized by agricultural, forested, and/or urban land.

With the advancement of technology and the rise in end-user expectations, the need and use of higher temporal resolution data for pollutant load estimation has increased. This protocol describes a method for continuous in situ water quality monitoring to obtain higher temporal resolution data for informed water resource management decisions.

Instream continuo monitoraggio delle sostanze nutrienti e sedimenti nei bacini agricoli

Pollutant concentrations and loads in watersheds vary considerably with time and space. Accurate and timely information on the magnitude of pollutants in ...

Capturing Flow-weighted Water and Suspended Particulates from Agricultural Canals During Drainage Events

The purpose of this study is to describe the methods used to capture flow-weighted water and suspended particulates from farm canals during drainage discharge events. Farm canals can be enriched by nutrients such as phosphorus (P) that are susceptible to transport. Phosphorus in the form of suspended particulates can significantly contribute to the overall P loads in drainage water. A settling tank experiment was conducted to capture suspended particulates during discrete drainage events. Farm canal discharge water was collected in a series of two 200 L settling tanks over the entire duration of the drainage event, so as to represent a composite subsample of the water being discharged. Imhoff settling cones are ultimately used to settle out the suspended particulates. This is achieved by siphoning water from the settling tanks via the cones. The particulates are then collected for physico-chemical analyses.

Nutrients present in particulate form can contribute significantly to the overall loads in agricultural drainage waters. This study describes a novel method to capture flow-weighted water and suspended particulates from farm canal drainage over the entire duration of the drainage event.

Acquisizione ponderati per il flusso di acqua e particelle in sospensione dai canali agricoli durante eventi di drenaggio

The purpose of this study is to describe the methods used to capture flow-weighted water and suspended particulates from farm canals during drainage ...

Continuous Hydrologic and Water Quality Monitoring of Vernal Ponds

Vernal ponds, also referred to as vernal pools, provide critical ecosystem services and habitat for a variety of threatened and endangered species. However, they are vulnerable parts of the landscapes that are often poorly understood and understudied. Land use and management practices, as well as climate change are thought to be a contribution to the global amphibian decline. However, more research is needed to understand the extent of these impacts. Here, we present methodology for characterizing a vernal pond's morphology and detail a monitoring station that can be used to collect water quantity and quality data over the duration of a vernal pond's hydroperiod. We provide methodology for how to conduct field surveys to characterize the morphology and develop stage-storage curves for a vernal pond. Additionally, we provide methodology for monitoring the water level, temperature, pH, oxidation-reduction potential, dissolved oxygen, and electrical conductivity of water in a vernal pond, as well as monitoring rainfall data. This information can be used to better quantify the ecosystem services that vernal ponds provide and the impacts of anthropogenic activities on their ability to provide these services.

Understanding the ecosystem services and processes provided by vernal ponds and the impacts of anthropogenic activities on their ability to provide these services requires intensive hydrologic monitoring. This sampling protocol using in-situ monitoring equipment was developed to understand the impact of anthropogenic activities on water levels and quality.

Continuo idrologici e monitoraggio della qualità di Vernal stagni di acqua

Vernal ponds, also referred to as vernal pools, provide critical ecosystem services and habitat for a variety of threatened and endangered species. ...

An Ultra-clean Multilayer Apparatus for Collecting Size Fractionated Marine Plankton and Suspended Particles

The distributions of many trace elements in the ocean are strongly associated with the growth, death, and re-mineralization of marine plankton and those of suspended/sinking particles. Here, we present an all plastic (Polypropylene and Polycarbonate), multi-layer filtration system for collection of suspended particulate matter (SPM) at sea. This ultra-clean sampling device has been designed and developed specifically for trace element studies. Meticulous selection of all non-metallic materials and utilization of an in-line flow-through procedure minimizes any possible metal contamination during sampling. This system has been successfully tested and tweaked for determining trace metals (e.g., Fe, Al, Mn, Cd, Cu, Ni) on particles of varying size in coastal and open ocean waters. Results from the South China Sea at the South East Asia Time-Series (SEATS) station indicate that diurnal variations and spatial distribution of plankton in the euphotic zone can be easily resolved and recognized. Chemical analysis of size-fractionated particles in surface waters of the Taiwan Strait suggests that the larger particles (&#62;153 &#181;m) were mostly biologically derived, while the smaller particles (10 - 63 &#181;m) were mostly composed of inorganic matter. Apart from Cd, the concentrations of metals (Fe, Al, Mn, Cu, Ni) decreased with increasing size.

Plankton and suspended particles play a major role in the biogeochemical cycles in the ocean. Here, we provide an ultra-clean, low stress method for the collection of various sizes of particles and plankton at sea with the capability of handling large volumes of seawater.

Un apparato multistrato ultra-pulito per la raccolta di plancton marino dimensioni frazionato e particelle in sospensione

The distributions of many trace elements in the ocean are strongly associated with the growth, death, and re-mineralization of marine plankton and those ...

Design and Use of an Apparatus for Quantifying Bivalve Suspension Feeding at Sea

As shellfish aquaculture moves from coastal embayments and estuaries to offshore locations, the need to quantify ecosystem interactions of farmed bivalves (i.e., mussels, oysters, and clams) presents new challenges. Quantitative data on the feeding behavior of suspension-feeding mollusks is necessary to determine important ecosystem interactions of offshore shellfish farms, including their carrying capacity, the competition with the zooplankton community, the availability of trophic resources at different depths, and the deposition to the benthos. The biodeposition method is used to quantify feeding variables in suspension-feeding bivalves in a natural setting and represents a more realistic proxy than laboratory experiments. This method, however, relies upon a stable platform to satisfy the requirements that water flow rates supplied to the shellfish remain constant and the bivalves are undisturbed. A flow-through device and process for using the biodeposition method to quantify the feeding of bivalve mollusks were modified from a land-based format for shipboard use by building a two-dimensional gimbal table around the device. Planimeter data reveal a minimal pitch and yaw of the chambers containing the test shellfish despite boat motion, the flow rates within the chambers remain constant, and operators are able to collect the biodeposits (feces and pseudofeces) with sufficient consistency to obtain accurate measurements of the bivalve clearance, filtration, selection, ingestion, rejection, and absorption at offshore shellfish aquaculture sites.

A flow-through device for using the biodeposition method to quantify filtration and feeding behavior of bivalve mollusks was modified for shipboard use. A two-dimensional gimbal table built around the device isolates the apparatus from boat motion, thereby allowing the accurate quantification of bivalve filtration variables at offshore shellfish aquaculture sites.

Progettazione e l'uso di un apparato per quantificare bivalvi sospensione alimentazione in mare

As shellfish aquaculture moves from coastal embayments and estuaries to offshore locations, the need to quantify ecosystem interactions of farmed bivalves ...

In Situ Chemotaxis Assay to Examine Microbial Behavior in Aquatic Ecosystems

Microbial behaviors, such as motility and chemotaxis (the ability of a cell to alter its movement in response to a chemical gradient), are widespread across the bacterial and archaeal domains. Chemotaxis can result in substantial resource acquisition advantages in heterogeneous environments. It also plays a crucial role in symbiotic interactions, disease, and global processes, such as biogeochemical cycling. However, current techniques restrict chemotaxis research to the laboratory and are not easily applicable in the field. Presented here is a step-by-step protocol for the deployment of the in situ chemotaxis assay (ISCA), a device that enables robust interrogation of microbial chemotaxis directly in the natural environment. The ISCA is a microfluidic device consisting of a 20 well array, in which chemicals of interest can be loaded. Once deployed in aqueous environments, chemicals diffuse out of the wells, creating concentration gradients that microbes sense and respond to by swimming into the wells via chemotaxis. The well contents can then be sampled and used to (1) quantify strength of the chemotactic responses to specific compounds through flow cytometry, (2) isolate and culture responsive microorganisms, and (3) characterize the identity and genomic potential of the responding populations through molecular techniques. The ISCA is a flexible platform that can be deployed in any system with an aqueous phase, including marine, freshwater, and soil environments.

Presented here is the protocol for an in situ chemotaxis assay, a recently developed microfluidic device that enables studies of microbial behavior directly in the environment.

In Situ Chemotaxis Assay per esaminare il comportamento microbico negli ecosistemi acquatici

Microbial behaviors, such as motility and chemotaxis (the ability of a cell to alter its movement in response to a chemical gradient), are widespread ...

Microparticle Manipulation by Standing Surface Acoustic Waves with Dual-frequency Excitations

We demonstrate a method for increasing the tuning ability of a standing surface acoustic wave (SSAW) for microparticles manipulation in a lab-on-a-chip (LOC) system. The simultaneous excitation of the fundamental frequency and its third harmonic, which is termed as dual-frequency excitation, to a pair of interdigital transducers (IDTs) could generate a new type of standing acoustic waves in a microfluidic channel. Varying the power and the phase in the dual-frequency excitation signals results in a reconfigurable field of the acoustic radiation force applied to the microparticles across the microchannel (e.g., the number and location of the pressure nodes and the microparticle concentrations at the corresponding pressure nodes). This article demonstrates that the motion time of the microparticle to only one pressure node can be reduced ~2-fold at the power ratio of the fundamental frequency greater than ~90%. In contrast, there are three pressure nodes in the microchannel if less than this threshold. Furthermore, adjusting the initial phase between the fundamental frequency and the third harmonic results in different motion rates of the three SSAW pressure nodes, as well as in the percentage of microparticles at each pressure node in the microchannel. There is a good agreement between the experimental observation and the numerical predictions. This novel excitation method can easily and non-invasively integrate into the LOC system, with a wide tenability and only a few changes to the experimental set-up.

A protocol for manipulating the microparticles in a microfluidic channel with a dual-frequency excitation is presented.

Manipolazione di microparticelle levandosi in piedi le onde acustiche superficiali con doppia frequenza eccitazioni

We demonstrate a method for increasing the tuning ability of a standing surface acoustic wave (SSAW) for microparticles manipulation in a lab-on-a-chip ...

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Dissolved Solute Sampling Across an Oxic-Anoxic Soil-Water Interface Using Microdialysis Profilers

Biogeochemical processes shift rapidly in both spatial (millimeter scale) and temporal (hour scale to day scale) dimensions at the oxic-anoxic interface in response to disturbances. Deciphering the rapid biogeochemical changes requires in situ, minimally invasive tools with high spatial and temporal sampling resolution. However, the available passive sampling devices are not very useful in many cases either due to their disposable nature or the complexity and extensive workload for sample preparation.
To address this problem, a microdialysis profiler with 33 individual polyethersulfone nanomembrane tubes (semipermeable, &#60;20 nm pore size) integrated into the one-dimensional skeleton (60 mm) was established to iteratively sample the dissolved compounds in porewater across the soil-water interface at a high resolution of 1.8 mm (outer diameter plus one spacing, i.e., 0.1 mm between probes). The sampling mechanism is based on the principle of concentration gradient diffusion. The automatic loading of degassed water allows minimal disturbance to the chemical species across the oxic-anoxic interface.
This paper describes the procedures of device setup and continuous porewater sampling across the soil-water interface on a daily basis. Concentration-depth profiles were selectively measured before (on Day 6) and after (on Day 7) disturbances induced by irrigation. The results showed that concentration-depth profiles were undergoing rapid changes, especially for redox-sensitive elements (i.e., iron and arsenic). These protocols can help investigate the biogeochemical responses across the soil-water interface under various disturbances caused by physical, chemical, and biological factors. The paper thoroughly discusses the advantages and disadvantages of this method for potential use in the environmental sciences.

A microdialysis profiler is described to sample dissolved porewater solutes across an oxic-anoxic soil-water interface in situ with minimal disturbance. This device is designed to capture rapid changes in concentration-depth profiles induced by disturbances at the soil-water interface and beyond.

Campionamento di soluti disciolti attraverso un'interfaccia ossa-anossica suolo-acqua utilizzando profilatori di microdialisi

Biogeochemical processes shift rapidly in both spatial (millimeter scale) and temporal (hour scale to day scale) dimensions at the oxic-anoxic interface ...

VacuSIP, un metodo migliorato per Inex

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