Les auteurs tiennent à remercier tous les pêcheurs de homard de la Nouvelle Angleterre commerciales dont les bateaux nous ont permis d&#39;échantillonner - leur coopération est très appréciée. En outre, nous remercions Little Bay Lobster Company (Newington, NH) et fruits de mer de Champlin (Point Judith, RI) qui nous a permis de procéder à un échantillonnage à leurs installations. Les étudiants UNH suivantes ont fourni une aide précieuse tout au long de cette étude: Haley Blanc, Françoise Morrison, Sarah Havener, Audra Chaput, et mai Grose. Ce projet a été financé par des subventions NH Seagrant à WHW (projet # R/CFR-11) et une subvention du programme UNH Marine à JSG.

Partie A: Domaine Technique d&#39;échantillonnage Une. Mesures femmes <ol><li> Pour chaque homard femelle, mesurer la longueur de la carapace (CL) et la largeur de la deuxième segment abdominal à la plus proche de 1,0 mm en utilisant une paire d&#39;étriers. NOTE: Le deuxième segment abdominal est mesurée parce que la largeur de l&#39;abdomen (où les femmes portent des œufs) est un indicateur de la maturité sexuelle. En outre, il pourrait être avantageux de déterminer le stade de la mue pour correspondre à plus tard ces données avec le stade de la reproduction (voir l&#39;étape 1.2). </li><li> Couper une petite partie distale de l&#39;un des pléopodes abdominaux (un protocole méthodologique standard homards) et stocker l&#39;eau de mer propre dans pleopod pour les visionner plus tard avec un microscope de dissection 9. REMARQUE: Cette étape peut également révéler la présence de glandes de ciment sur ​​les pléopodes, qui sont une indication que les femmes se préparent à extruder leurs œufs 10. </li></ol> 2. Aiguille Insertion <ol><li> Insérez l&#39;aiguille de la seringue (18 G, voir le tableau 1) ~ 1 cm dans le réceptacle séminal, inclinées vers la queue à 45 ° ~ (figure 1). NOTE: Dans les homards avec des bouchons de spermatozoïdes intacts, l&#39;aiguille peut rencontrer une certaine résistance lors de l&#39;insertion, comme il est poussé à travers le bouchon de sperme avant de parvenir à la plus forte concentration de sperme au fond de la réceptacle séminal (Figure 1). NOTE: La présence de la fiche, en soi, est un indicateur fiable que les homards se sont accouplés. Chez certains animaux, il peut y avoir peu de résistance, peut-être parce que personne ne spermatophore est présent, le spermatophore est petit, il n&#39;a pas encore durci dans un bouchon, ou il peut avoir déjà été utilisé pour féconder un ou plusieurs couvées. </li></ol><img alt="Figure 1" fo:content-width="5in" fo:src="/files/ftp_upload/50498/50498fig1highres.jpg" src="/files/ftp_upload/50498/50498fig1.jpg" wide = &quot;600px&quot; /&gt; Figure 1. insertion de l&#39;aiguille et vue de homard réceptacle séminal. place de l&#39;aiguille pointe arrondie dans le réceptacle séminal de la langouste pour extraire un échantillon de sperme. L&#39;angle et la profondeur de l&#39;aiguille sont à la fois critique d&#39;obtenir des échantillons homogènes. Droite: Vue d&#39;un réceptacle séminal qui a été disséqué et partagée verticalement. Le bouchon de sperme occupe la partie supérieure ou plus à l&#39;extérieur du récipient, tandis que les spermatozoïdes sont situés dans le bas (du point le plus profond) du récipient. Il est important de noter que l&#39;échantillonnage en dessous de cette profondeur peut percer le récipient et entraîner dans un échantillon de sang contaminé. <a href="https://www.jove.com/files/ftp_upload/50498/50498fig1highres.jpg" target="_blank">Cliquez ici pour agrandir l&#39;image</a> . 3. Pénétrant Plug sperme <ol><li> Après l&#39;insertion initiale, s&#39;accentuer l&#39;angle de l&#39;aiguille à presque verticale (perpendiculaire à l&#39;axe du corps), et très lentement commencer à travaillervers le bas dans le réceptacle. </li><li> Gardez l&#39;aiguille inclinée à un angle de sorte que la pointe est dirigée légèrement vers la queue. Dans ce cas, la pointe de l&#39;aiguille est la compression de la partie supérieure du bouchon, jusqu&#39;à ce qu&#39;elle pénètre finalement dans la matrice. </li><li> Continuer à pousser lentement l&#39;aiguille vers le bas jusqu&#39;à sentir une résistance, une fois de plus - ce qui est le fond du récipient et où l&#39;échantillon est prélevé. </li></ol> 4. Retrait sperme <ol><li> Prélever un échantillon (souvent solide de matériau de bouchon, le sperme et le liquide d&#39;un autre fluide) à partir du fond du récipient et au dépôt de l&#39;échantillon dans un tube en plastique de 2,0 ml étiqueté. </li><li> Rincer le récipient avec ~ 0.1-0.5 ml d&#39;eau de mer froide. </li><li> Enlevez environ 0,3 ml de liquide et le placer dans le même tube. </li><li> des échantillons de sperme de magasin sur la glace jusqu&#39;à ce qu&#39;ils puissent être examinés dans le laboratoire. Partie B: Examen des échantillons de sperme dans le laboratoire NOTE: Les échantillons retirés des réceptacles séminaux de l individu<html> obsters devraient être examinés afin de déterminer si les spermatozoïdes étaient présents ou absents. </li><li> Retirer ~ 50 pi de liquide de chaque tube en plastique, placer sur une lame de verre avec une lamelle, et vue à 100X avec un microscope composé. </li><li> Note échantillons: sperme rares, nombreux, ou absent (cette étude n&#39;a pas quantifié le nombre de spermatozoïdes réels, figure 2). </li></ol><img alt="Figure 2" fo:content-width="5in" fo:src="/files/ftp_upload/50498/50498fig2highres.jpg" src="/files/ftp_upload/50498/50498fig2.jpg" width="600px" /> Figure 2. Images d&#39;échantillons de homard de sperme. Images d&#39;échantillons de sperme prélevés homard de réceptacle séminal de la femelle, comme vu avec un microscope composé à grossissement de 40X. L&#39;image à gauche montre un échantillon qui contenait de nombreux sperme tandis que l&#39;autre (à droite) illustre un échantillon bien maigre (barre d&#39;échelle = 10 pm).Jpg &quot;target =&quot;. _blank &quot;&gt; Cliquez ici pour agrandir l&#39;image.

<ol>
	<li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> FAO and Agriculture Organization of the United Nations). Fisheries and Aquaculture Department. , (2012).</li><li>MacDiarmid, A., Butler, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sperm+economy+and+limitation+in+spiny+lobsters.">Sperm economy and limitation in spiny lobsters.</a> Behav. Ecol. Sociobiol. 46, 14-24 (1999).</li><li>Kendall, M. S., Wolcott, D. L., Wolcott, T. G., Hines, A. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+male+size+and+mating+history+on+sperm+content+of+ejaculates+of+the+blue+crab+Callinectes+sapidus.">Influence of male size and mating history on sperm content of ejaculates of the blue crab Callinectes sapidus.</a> 230, 235-240 (2002).</li><li>Sato, T., Ashidate, M., Goshima, S. <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+male+mating+frequency+and+male+size+on+ejaculate+size+and+reproductive+success+of+female+spiny+king+crab,+Parolithodes+brevipes.">Effects of male mating frequency and male size on ejaculate size and reproductive success of female spiny king crab, Parolithodes brevipes.</a> Mar. Ecol. Prog. Ser. 296, 251-262 (2005).</li><li>Sato, T., Goshima, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impacts+of+male-only+fishing+and+sperm+limitation+in+manipulated+populations+of+an+unfished+crab,+Hapalogaster.">Impacts of male-only fishing and sperm limitation in manipulated populations of an unfished crab, Hapalogaster.</a> 313, 193-204 (2006).</li><li>Rondeau, A., Sainte-Marie, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Variable+mate-guarding+time+and+sperm+allocation+by+male+snow+crabs+(Chionoecetes+opilio)+in+response+to+sexual+competition,+and+their+impact+on+the+mating+success+of+females.">Variable mate-guarding time and sperm allocation by male snow crabs (Chionoecetes opilio) in response to sexual competition, and their impact on the mating success of females.</a> Biol. Bull. 201, 204-217 (2001).</li><li>Waddy, S. L., Aiken, D. E., Bauer, R. T., Martin, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Mating and insemination in the Ameri can lobster,Homarus americanus. Crustacean sexual behavior. , 126-144 (1990).</li><li>Johnson, K. J., Goldstein, J. S., Watson, W. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Two+methods+for+determining+the+fertility+status+in+early-stage+American+lobster,+Homarus+americanus,+eggs.">Two methods for determining the fertility status in early-stage American lobster, Homarus americanus, eggs.</a> J. Crust. Biol. 31, 693-700 (2011).</li><li>Aiken, D. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Proecdysis,+setal+development,+and+molt+prediction+in+the+American+lobster+(Homarus).">Proecdysis, setal development, and molt prediction in the American lobster (Homarus).</a> 30, 1337-1344 (1973).</li><li>Aiken, D. E., Waddy, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cement+gland+development,+ovary+maturation,+and+reproductive+cycles+in+the+American+lobster+Homarus+americanus.">Cement gland development, ovary maturation, and reproductive cycles in the American lobster Homarus americanus.</a> J. Crust. Biol. 2, 315-327 (1982).</li><li>Pugh, T. L., Goldstein, J. S., Lavalli, K. L., Clancy, M., Watson, W. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=At-sea+determination+of+female+American+lobsters+(Homarus+americanus)+mating+activity:+Patterns+vs.+expectations.">At-sea determination of female American lobsters (Homarus americanus) mating activity: Patterns vs. expectations.</a> Fish. Res. 147, 327-337 (2013).</li><li>Bauer, R. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phylogenetic+trends+in+sperm+transfer+and+storage+complexity+in+decapod+crustaceans.">Phylogenetic trends in sperm transfer and storage complexity in decapod crustaceans.</a> J. Crust. Biol. 6, 313-325 (1986).</li><li>Talbot, P., Helluy, S., Factor, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reproduction+and+embryonic+development.">Reproduction and embryonic development.</a> Biology of the Lobster: Homarus americanus. , 177-216 (1995).</li><li>Cooper, R. A., Uzmann, J. R., Cobb, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Ecology of juvenile and adult Homarus. The biology and management of lobsters. 2, 97-142 (1980).</li><li>Estrella, B. T., McKiernan, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Catch-per-unit-effort+and+biological+parameters+from+the+Massachusetts+coastal+lobster+(Homarus+americanus)+resource:+description+and+trends.">Catch-per-unit-effort and biological parameters from the Massachusetts coastal lobster (Homarus americanus) resource: description and trends.</a> NOAA Tech. Rep. NMFS. 81, (1989).</li><li>Waddy, S. L., Aiken, D. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multiple+fertilization+and+consecutive+spawning+in+large+American+lobster,+Homarus+americanus.">Multiple fertilization and consecutive spawning in large American lobster, Homarus americanus.</a> Can. J. Fish. Aquat. Sci. 43, 2291-2294 (1986).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Atlantic+States+Marine+Fisheries+Commission+(ASMFC).+American+lobster+stock+assessment+report+for+peer+review.">Atlantic States Marine Fisheries Commission (ASMFC). American lobster stock assessment report for peer review.</a> ASMFC Stock Assessment. , (2009).</li></ol>

Les auteurs déclarent aucun conflit d&#39;intérêt.

Cette vidéo et papier contour et démontrer une méthode pour déterminer si les homards femelles se sont accouplés avec succès. Cette approche est suffisamment simple pour qu&#39;il puisse être utilisé pour l&#39;échantillonnage à grande échelle de homards en mer par une variété d&#39;utilisateurs. La méthode est basée sur la détection de la présence d&#39;un spermatophore dans le réceptacle séminal qui est passée du sexe masculin au homards femelles pendant l&#39;accouplement. Ces spermatophores durci...

<html> Homard d&#39;Amérique (Homarus americanus) est l&#39;une des pêches les plus productives de l&#39;Atlantique Nord (~ 56 000 tonnes en 2011, d&#39;une valeur plus de 390 millions $) 1. Cependant, il ya un manque général de compréhension en ce qui concerne la dynamique de reproduction de cette espèce dans les populations sauvages. Produire des estimations plus précises de la reproduction, y compris le nombre et la taille des individus participant activement à la reproduction, peut améliorer le processus d&#39;évaluation des stocks. Ainsi, les femelles qui sont empêchés de réaliser leurs potentialités reproductives raison de limitation de spermatozoïdes ont été identifiés comme une inquiétude pour plusieurs de crustacés marines exploitées commercialement, notamment: langoustes de 2, crabes bleus 3, crabes royaux 4, crabes 5 et crabes des neiges 6. L&#39;objectif global est de déterminer si la limitation de sperme pourrait être un facteur dans certaines régions de la pêche au homard américain ainsi. nt &quot;&gt; Les premiers signes d&#39;un problème potentiel de limitation du sperme chez les homards ont été observés pendant la conduite d&#39;une étude de marquage sans rapport avec les homards femelles ovigères (avec des oeufs visiblement intactes, récemment extrudés). Récupération des rapports de pêcheurs a indiqué que ~ 15% de ces animaux avait chuté leurs couvées après seulement 1-2 mois. L&#39;hypothèse de travail est que certains homards portaient des oeufs qui n&#39;ont pas été fécondés et, par conséquent, «tombé» 7. données préliminaires d&#39;une étude qui a suivi ont confirmé le fait que les homards extruder des oeufs, même si elles ne sont pas accouplés avec succès et ces œufs non fécondés ne sont effectués pour ~ 1 mois 8. Par conséquent, compte tenu de l&#39;observation des femelles portant des oeufs non fécondés dans les populations naturelles, nous avons cherché à déterminer la mesure dans laquelle la limitation de sperme pourrait contribuer à un niveau sous-maximal de reproduction par homards sexuellement matures. Pour atteindre cet objectif, une technique a été développée pour détecter la spermatophore déposé par les hommes pendant l&#39;accouplement. Ce papier et vidéo décrivent une méthode simple non invasif développé pour déterminer le succès d&#39;accouplement des femelles homards. La technique peut être utilisée rapidement et de façon fiable à la mer, soit à bord de la recherche ou des navires de pêche commerciale. Les détails de ce procédé de prélèvement, ainsi que quelques résultats représentatifs sont présentés pour illustrer l&#39;application de la technique.

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			<td height="40" style="height:40px;width:163px;">Calipers: Mitutoyo Digimatic</td>
			<td style="width:129px;">Mitutoyo</td>
			<td style="width:135px;">500-196-20</td>
			<td style="width:231px;">http://www.globalindustrial.com</td>
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			<td height="10" style="height:10px;width:163px;"></td>
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			<td height="40" style="height:40px;width:163px;">3 mL Luer-Lok&#8482; syringe</td>
			<td style="width:129px;">Beckton-Dickinson</td>
			<td style="width:135px;">309585</td>
			<td style="width:231px;">www.bd.com</td>
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		<tr height="10">
			<td height="10" style="height:10px;width:163px;"></td>
			<td style="width:129px;"></td>
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			<td height="60" style="height:60px;width:163px;">Monoject aluminum hub blunt needles, 18-G x 1</td>
			<td style="width:129px;">Webster Veterinary</td>
			<td style="width:135px;">8881202348</td>
			<td style="width:231px;">www.mywebstervet.com</td>
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			<td height="10" style="height:10px;width:163px;"></td>
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			<td height="40" style="height:40px;width:163px;">2.0 mL plastic storage tubes</td>
			<td style="width:129px;">Eppendorf&#160;</td>
			<td style="width:135px;">wu-06333-72</td>
			<td style="width:231px;">http://www.coleparmer.com</td>
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			<td height="40" style="height:40px;width:163px;">Styrofoam cooler with ice</td>
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			<td height="40" style="height:40px;width:163px;">Compound light microscope</td>
			<td style="width:129px;">Olympus&#160;</td>
			<td style="width:135px;">BH System</td>
			<td style="width:231px;">http://www.olympusamerica.com</td>
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			<td height="10" style="height:10px;width:163px;"></td>
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			<td height="40" style="height:40px;width:163px;">Glass sides with cover slip</td>
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			<td style="width:231px;">75 mm x 25 mm, ~ 1 mm thickness</td>
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			<td height="41" style="height:41px;width:163px;">Pipettor with disposable tips</td>
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a noninvasive method for in situ determination of mating success in female american lobsters (homarus americanus)

En dépit d&#39;être l&#39;une des pêches les plus productives de l&#39;Atlantique Nord-Ouest, il reste encore beaucoup sur la dynamique de reproduction naturelle de homards américains. Des travaux récents dans les populations de crustacés exploités (crabes et homards) suggère qu&#39;il existe des circonstances où les femelles matures sont incapables d&#39;atteindre leur potentiel de reproduction complet en raison de la limitation de sperme. Pour examiner cette possibilité dans les différentes régions de la pêche au homard, une méthode fiable et non invasive a été développé pour échantillonner un grand nombre de femelles homards en mer. Ce procédé consiste à insérer une aiguille à pointe arrondie dans le réceptacle séminal de la femelle pour déterminer la présence ou l&#39;absence d&#39;un bouchon de spermatozoïdes et de retirer un échantillon qui peut être examinée pour la présence de spermatozoïdes. Une série d&#39;études de contrôle ont été menées sur le quai et dans le laboratoire pour tester la fiabilité de cette technique. Ces efforts ont entraîné échantillonnage 294 homards femelles pour confirmer que la présence ofa fiche de sperme était un indicateur fiable de spermatozoïdes dans le réceptacle et donc, l&#39;accouplement. Ce document détaille la méthodologie et les résultats obtenus à partir d&#39;un sous-ensemble de l&#39;ensemble des femmes de l&#39;échantillon. Parmi les 230 homards femelles prélevées dans le banc Georges et Cape Ann, MA (gamme de taille = 71 à 145 mm de longueur de carapace), 90,3% étaient positifs pour le sperme. Explications possibles de l&#39;absence de spermatozoïdes dans certaines femelles comprennent: l&#39;immaturité (manque de maturité physiologique), la répartition de la prise de sperme après avoir été utilisé pour fertiliser une couvée d&#39;oeufs, et le manque d&#39;activité de l&#39;accouplement. Les enquêtes indiquent que cette technique pour examiner le succès d&#39;accouplement des femelles homards est un indicateur fiable qui peut être utilisé dans le domaine de documenter l&#39;activité de reproduction dans les populations naturelles.

<html> Les données présentées ici sont pour un sous-ensemble de homards femelles qui proviennent à la fois du littoral et les centres offshore. Le groupe de nearshore composée de 44 homards qui ont été capturés dans les eaux côtières près de Cape Ann, Massachusetts et transportés dans des glacières au laboratoire pour examen. Un total de 186 homards en mer ont été capturés sur le banc Georges et échantillonné à un centre de détention commercial dans le New Hampshire. Des ?...

Watch this Scientific Journal Video about A Noninvasive Method For In situ Determination of Mating Success in Female American Lobsters Homarus americanus at JoVE.com

A Noninvasive Method For In situ Determination of Mating Success in Female American Lobsters Homarus americanus

Les changements induits par la pêche-à pêche de crustacés exploités, comme la pêche au homard américain, pourraient influer sur leur dynamique de reproduction, conduisant à une réduction dans le succès de l&#39;accouplement. L&#39;objectif de cette étude était de développer une méthode non invasive pour déterminer le succès d&#39;accouplement des homards femelles qui peuvent être physiologiquement ou fonctionnellement mature.

Une méthode non invasive pour<em&gt; In situ</em&gt; Détermination du succès de l&#39;accouplement chez les femelles homards américains (<em&gt; Homarus americanus</em&gt;)

Despite being one of the most productive fisheries in the Northwest Atlantic, much remains unknown about the natural reproductive dynamics of American ...

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A Noninvasive Method For In situ Determination of Mating Success in Female American Lobsters (Homarus americanus)

10.9K vues.  University of New Hampshire.  Les changements induits par la pêche-à pêche de crustacés exploités, comme la pêche au homard américain, pourraient influer sur leur dynamique de reproduction, conduisant à une réduction dans le succès de l'accouplement. L'objectif de cette étude était de développer une méthode non invasive pour déterminer le succès d'accouplement des homards femelles qui peuvent être physiologiquement ou fonctionnellement mature. 

Vidéo: A Noninvasive Method For In situ Determination of Mating Success in Female American Lobsters Homarus americanus

A Technique to Screen American Beech for Resistance to the Beech Scale Insect (Cryptococcus fagisuga Lind.)

Beech bark disease (BBD) results in high levels of initial mortality, leaving behind survivor trees that are greatly weakened and deformed.&#160;The disease is initiated by feeding activities of the invasive beech scale insect, Cryptococcus fagisuga, which creates entry points for infection by one of the Neonectria species of fungus.&#160;Without scale infestation, there is little opportunity for fungal infection. Using scale eggs to artificially infest healthy trees in heavily BBD impacted stands demonstrated that these trees were resistant to the scale insect portion of the disease complex1.&#160;Here we present a protocol that we have developed, based on the artificial infestation technique by Houston2, which can be used to screen for scale-resistant trees in the field and in smaller potted seedlings and grafts.&#160;The identification of scale-resistant trees is an important component of management of BBD through tree improvement programs and silvicultural manipulation.

A Technique to Screen American Beech for Resistance to the Beech Scale Insect Cryptococcus fagisuga Lind.

Beech bark disease is initiated by feeding activities of the beech scale insect that create fungal entry points in the bark.&#160;Trees that are resistant to the scale insect are also disease resistant.&#160;Here we present the protocol we have developed to screen individual beech trees for beech scale resistance.

Une technique à l&#39;écran hêtre américain pour la résistance à la cochenille Hêtre (<em&gt; Cryptococcus fagisuga</em&gt; Lind.)

Beech bark disease (BBD) results in high levels of initial mortality, leaving behind survivor trees that are greatly weakened and deformed.&#160;The ...

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Environnement

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.

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, une méthode Inex améliorée pour<em&gt; In Situ</em&gt; Mesure des particules et composés dissous transformés par suspensivores actifs

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

Optimized Procedure for Determining the Adsorption of Phosphonates onto Granular Ferric Hydroxide using a Miniaturized Phosphorus Determination Method

This paper introduces a procedure to investigate the adsorption of phosphonates onto iron-containing filter materials, particularly granular ferric hydroxide (GFH), with little effort and high reliability. The phosphonate, e.g., nitrilotrimethylphosphonic acid (NTMP), is brought into contact with the GFH in a rotator in a solution buffered by an organic acid (e.g., acetic acid) or Good buffer (e.g., 2-(N-morpholino)ethanesulfonic acid) [MES] and N-cyclohexyl-2-hydroxyl-3-aminopropanesulfonic acid [CAPSO]) in a concentration of 10 mM for a specific time in 50 mL centrifuge tubes. Subsequently, after membrane filtration (0.45 &#181;m pore size), the total phosphorus (total P) concentration is measured using a specifically developed determination method (ISOmini). This method is a modification and simplification of the ISO 6878 method: a 4 mL sample is mixed with H2SO4 and K2S2O8 in a screw cap vial, heated to 148-150 &#176;C for 1 h and then mixed with NaOH, ascorbic acid and acidified molybdate with antimony(III) (final volume of 10 mL) to produce a blue complex. The color intensity, which is linearly proportional to the phosphorus concentration, is measured spectrophotometrically (880 nm). It is demonstrated that the buffer concentration used has no significant effect on the adsorption of phosphonate between pH 4 and 12. The buffers, therefore, do not compete with the phosphonate for adsorption sites. Furthermore, the relatively high concentration of the buffer requires a higher dosage concentration of oxidizing agent (K2S2O8) for digestion than that specified in ISO 6878, which, together with the NaOH dosage, is matched to each buffer. Despite the simplification, the ISOmini method does not lose any of its accuracy compared to the standardized method.

This paper introduces a procedure to investigate the adsorption of phosphonates onto iron-containing filter materials, particularly granular ferric hydroxide, with little effort and high reliability. In a buffered solution, the phosphonate is brought into contact with the adsorbent using a rotator and then analyzed via a miniaturized phosphorus determination method.

Procédure optimisée pour la détermination de l’Adsorption de Phosphonates sur hydroxyde ferrique granulaire, en utilisant une méthode de dosage de phosphore miniaturisé

This paper introduces a procedure to investigate the adsorption of phosphonates onto iron-containing filter materials, particularly granular ferric ...

In Situ Hybridization Techniques for Paraffin-Embedded Adult Coral Samples

Corals are important ocean invertebrates that are critical for overall ocean health as well as human health. However, due to human impacts such as rising ocean temperatures and ocean acidification, corals are increasingly under threat. To tackle these challenges, advances in cell and molecular biology have proven to be crucial for diagnosing the health of corals. Modifying some of the techniques commonly used in human medicine could greatly improve researchers&#39; ability to treat and save corals. To address this, a protocol for in situ hybridization used primarily in human medicine and evolutionary developmental biology has been adapted for use in adult corals under stress.
The purpose of this method is to visualize the spatial expression of an RNA probe in adult coral tissue that has been embedded in paraffin and sectioned onto glass slides. This method focuses on removal of the paraffin and rehydration of the sample, pretreatment of the sample to ensure permeability of the sample, pre-hybridization incubation, hybridization of the RNA probe, and visualization of the RNA probe. This is a powerful method when using non-model organisms to discover where specific genes are expressed, and the protocol can be easily adapted for other non-model organisms. However, the method is limited in that it is primarily qualitative, because expression intensity can vary depending on the amount of time used during the visualization step and the concentration of the probe. Furthermore, patience is required, as this protocol can take up to 5 days (and in many cases, longer) depending on the probe being used. Finally, non-specific background staining is common, but this limitation can be overcome.

In Situ Hybridization Techniques for Paraffin-Embedded Adult Coral Samples

The goal of this protocol is to perform in situ hybridization on adult coral samples that have been embedded in paraffin and sectioned onto glass slides. This is a qualitative method used to visualize the spatial expression of an RNA anti-sense probe in paraffin-embedded tissues.

In Situ Techniques d’hybridation pour échantillons de corail adultes paraffine

Corals are important ocean invertebrates that are critical for overall ocean health as well as human health. However, due to human impacts such as rising ...

A Loop-mediated Isothermal Amplification (LAMP) Assay for Rapid Identification of Bemisia tabaci

The whitefly Bemisia tabaci (Gennadius) is an invasive pest of considerable importance, affecting the production of vegetable and ornamental crops in many countries around the world. Severe yield losses are caused by direct feeding, and even more importantly, also by the transmission of more than 100 harmful plant pathogenic viruses. As for other invasive pests, increased international trade facilitates the dispersal of B. tabaci to areas beyond its native range. Inspections of plant import products at points of entry such as seaports and airports are, therefore, seen as an important prevention measure. However, this last line of defense against pest invasions is only effective if rapid identification methods for suspicious insect specimens are readily available. Because the morphological differentiation between the regulated B. tabaci and close relatives without quarantine status is difficult for non-taxonomists, a rapid molecular identification assay based on the loop-mediated isothermal amplification (LAMP) technology has been developed. This publication reports the detailed protocol of the novel assay describing rapid DNA extraction, set-up of the LAMP reaction, as well as interpretation of its read-out, which allows identifying B. tabaci specimens within one hour. Compared to existing protocols for the detection of specific B. tabaci biotypes, the developed method targets the whole B. tabaci species complex in one assay. Moreover the assay is designed to be applied on-site by plant health inspectors with minimal laboratory training directly at points of entry. Thorough validation performed under laboratory and on-site conditions demonstrates that the reported LAMP assay is a rapid and reliable identification tool, improving the management of B. tabaci.

A Loop-mediated Isothermal Amplification LAMP Assay for Rapid Identification of Bemisia tabaci

This paper reports the protocol for a rapid identification assay for Bemisia tabaci based on loop-mediated isothermal amplification (LAMP) technology. The protocol requires minimal laboratory training and can, therefore, be implemented on-site at points of entry for plant imports such as seaports and airports.

Un test d’Amplification isotherme (lampe) médiée par la boucle pour une Identification rapide de Bemisia tabaci

The whitefly Bemisia tabaci (Gennadius) is an invasive pest of considerable importance, affecting the production of vegetable and ornamental crops in many ...

Implantation of a New Micro Acoustic Tag in Juvenile Pacific Lamprey and American Eel

Juvenile Pacific Lamprey and American eels were used for laboratory evaluations to determine potential effects from tag implantation. Telemetry technology has been identified as a way to obtain more detailed information on movement and behavior across a broader spatial scale than is possible with other known technology. The purpose of this method is to provide a detailed step by step instruction on tag implantation for both lampreys and eel. For laboratory studies using actively migrating juvenile Pacific Lamprey (120-160 mm), we determined that the presence of the tag did not alter the swimming ability between tagged and untagged Individuals or have any significant tag loss (&lt;3%). Similar results were determined during laboratory testing of Yellow phase American Eels (113-175 mm). No mortality occurred during a 38-day holding period and there was minimal tag loss (3.8%). The presence of the tag did not have any significant effect on the swimming ability or survival of tagged eels compared to untagged controls and there was minimal tag loss.

This article illustrates the procedure for implanting a micro acoustic tag in juvenile Pacific Lamprey and American Eel. Our application is based on laboratory trials, which indicate that acoustic tags can be successfully implanted with no apparent effects on swimming ability or survival and minimal tag loss.

Implantation d’une nouvelle balise acoustique Micro en juvénile lamproie du Pacifique et l’anguille d’Amérique

Juvenile Pacific Lamprey and American eels were used for laboratory evaluations to determine potential effects from tag implantation. Telemetry technology ...

A Telemetric, Gravimetric Platform for Real-Time Physiological Phenotyping of Plant&ndash;Environment Interactions

Food security for the growing global population is a major concern. The data provided by genomic tools far exceeds the supply of phenotypic data, creating a knowledge gap. To meet the challenge of improving crops to feed the growing global population, this gap must be bridged.
Physiological traits are considered key functional traits in the context of responsiveness or sensitivity to environmental conditions. Many recently introduced high-throughput (HTP) phenotyping techniques are based on remote sensing or imaging and are capable of directly measuring morphological traits, but measure physiological parameters mainly indirectly.
This paper describes a method for direct physiological phenotyping that has several advantages for the functional phenotyping of plant&#8211;environment interactions. It helps users overcome the many challenges encountered in the use of load-cell gravimetric systems and pot experiments. The suggested techniques will enable users to distinguish between soil weight, plant weight and soil water content, providing a method for the continuous and simultaneous measurement of dynamic soil, plant and atmosphere conditions, alongside the measurement of key physiological traits. This method allows researchers to closely mimic field stress scenarios while taking into consideration the environment&#8217;s effects on the plants&#8217; physiology. This method also minimizes pot effects, which are one of the major problems in pre-field phenotyping. It includes a feed-back fertigation system that enables a truly randomized experimental design at a field-like plant density. This system detects the soil-water-content limiting threshold (&#952;) and allows for the translation of data into knowledge through the use of a real-time analytic tool and an online statistical resource. This method for the rapid and direct measurement of the physiological responses of multiple plants to a dynamic environment has great potential for use in screening for beneficial traits associated with responses to abiotic stress, in the context of pre-field breeding and crop improvement.

A Telemetric, Gravimetric Platform for Real-Time Physiological Phenotyping of Plant&amp;#8211;Environment Interactions

This high-throughput, telemetric, whole-plant water relations gravimetric phenotyping method enables direct and simultaneous real-time measurements, as well as the analysis of multiple yield-related physiological traits involved in dynamic plant&#8211;environment interactions.

Une plate-forme télémétrique et gravimétrique pour le phénotypage physiologique en temps réel des interactions plante-environnement

Food security for the growing global population is a major concern. The data provided by genomic tools far exceeds the supply of phenotypic data, creating ...

Mistletoe Eradicator - A Novel Tool for Simultaneous Mechanical and Chemical Control of Mistletoe

Some species of mistletoes, an aerial plant hemiparasites, play a Janus-like role in ecosystems by being rich biodiversity hotspots due to their trophic and topic associations with various organisms and at the same time acting as biological pirates, exhausting the host trees by stealing water and nutrients via the haustorium. Many crop and timber trees are attacked by mistletoes, for example, dwarf mistletoes parasitize conifers, representatives of genus Tapinanthus target guava, and Phoradendron mistletoe target pecans. Recently, an increasing infestation of the European mistletoe (Viscum album subsp. album L.) in monoculture stands of common walnuts (Juglans regia L.)&#160;was recorded for the first time during 2018 in different regions of Kashmir Valley, India. The common chemical approach based on the spraying of mistletoe exophytes with the growth regulators, ethephon and glyphosate, was not efficient since V. album resprouted in the next vegetation season. Hand pruning of mistletoe was time-consuming, and a polyethylene covering resulted in the localized rotting of host tissues. The most efficient approach to eradicate mistletoe was the use of a novel tool called Mistletoe Eradicator, which allows to use mechanical and chemical means of mistletoe control simultaneously. It consists of a manipulating pole with the terminal pruning saw having a container filled with the growth regulator, from which the liquid slowly drops onto the saw blade through the nozzle. It allows working both from the ground and in the tree canopy. After the targeted application of 5%-10% ethephon or glyphosate to the base of the just pruned V. album stems (stubs), exophyte resprouting was significantly reduced 9 months after the pruning as compared to the only pruned or sprayed exophytes. An alternative variant of Mistletoe Eradicator was constructed in Ukraine filled just with a food color additive to track the liquid dropping and was successfully tested.

The protocol describes an easy-to-handle tool called Mistletoe Eradicator which combines mechanical and chemical approaches and is efficient in the control of mistletoe infestation on the host trees. The manuscript uses the European mistletoe (Viscum album subsp. album L.) as an example.

Éradicateur de gui - Un nouvel outil pour le contrôle mécanique et chimique simultané du gui

Some species of mistletoes, an aerial plant hemiparasites, play a Janus-like role in ecosystems by being rich biodiversity hotspots due to their trophic ...

A Complete Method for Evaluating the Performance of Photocatalysts for the Degradation of Antibiotics in Environmental Remediation

Various antibiotics such as tetracycline, aureomycin, amoxicillin, and levofloxacin are found in large quantities in groundwater and soil systems, potentially leading to the development of resistant and multi-drug resistant bacteria, posing a threat to humans, animals, and environmental systems. Photocatalytic technology has attracted keen interest due to its rapid and stable treatment and direct use of solar energy. However, most studies evaluating the performance of semiconductor catalysts for the photocatalytic degradation of organic pollutants in water are currently incomplete. In this paper, a complete experimental protocol is designed to comprehensively evaluate the photocatalytic performance of semiconductor catalysts. Herein, rhombic dodecahedral silver phosphate was prepared by a simple solvent phase synthesis method at room temperature and atmospheric pressure. BrSubphthalocyanine/Ag3PO4 heterojunction materials were prepared by the solvothermal method. The catalytic performance of as-prepared materials for the degradation of tetracycline was evaluated by studying different influencing factors such as catalyst dosage, temperature, pH, and anions at atmospheric pressure using a 300 W xenon lamp as a simulated solar light source and a light intensity of 350 mW/cm2. Compared with the first cycle, the constructed BrSubphthalocyanine/Ag3PO4 maintained 82.0% of the original photocatalytic activity after five photocatalytic cycles, while the pristine Ag3PO4 maintained only 28.6%. The stability of silver phosphate samples was further tested by a five-cycle experiment. This paper provides a complete process for evaluating the catalytic performance of semiconductor catalysts in the laboratory for the development of semiconductor catalysts with potential for practical applications.

Presented here is a protocol to explore a universal set of experimental procedures for comprehensive laboratory evaluation of photocatalysts in the field of environmental purification, using the example of photocatalytic removal of antibiotic organic pollutant molecules from water by phthalocyanine sensitized silver phosphate composites.

Méthode complète d’évaluation de la performance des photocatalyseurs pour la dégradation des antibiotiques dans l’assainissement de l’environnement

Various antibiotics such as tetracycline, aureomycin, amoxicillin, and levofloxacin are found in large quantities in groundwater and soil systems, ...

In Situ Soil Moisture Sensors in Undisturbed Soils

Soil moisture directly affects operational hydrology, food security, ecosystem services, and the climate system. However, the adoption of soil moisture data has been slow due to inconsistent data collection, poor standardization, and typically short record duration. Soil moisture, or quantitatively volumetric soil water content (SWC), is measured using buried, in situ sensors that infer SWC from an electromagnetic response. This signal can vary considerably with local site conditions such as clay content and mineralogy, soil salinity or bulk electrical conductivity, and soil temperature; each of these can have varying impacts depending on the sensor technology.
Furthermore, poor soil contact and sensor degradation can affect the quality of these readings over time. Unlike more traditional environmental sensors, there are no accepted standards, maintenance practices, or quality controls for SWC data. As such, SWC is a challenging measurement for many environmental monitoring networks to implement. Here, we attempt to establish a community-based standard of practice for in situ SWC sensors so that future research and applications have consistent guidance on site selection, sensor installation, data interpretation, and long-term maintenance of monitoring stations.
The videography focuses on a multi-agency consensus of best-practices and recommendations for the installation of in situ SWC sensors. This paper presents an overview of this protocol along with the various steps essential for high-quality and long-term SWC data collection. This protocol will be of use to scientists and engineers hoping to deploy a single station or an entire network.

In Situ Soil Moisture Sensors in Undisturbed Soils

The determination of soil water content is a critical mission requirement for many state and federal agencies. This protocol synthesizes multi-agency efforts to measure soil water content using buried in situ sensors.

In situ Capteurs d’humidité du sol dans les sols non perturbés

Soil moisture directly affects operational hydrology, food security, ecosystem services, and the climate system. However, the adoption of soil moisture ...