Os autores agradecem o financiamento do Monitoramento do Serviço Florestal de Avaliação de Proteção à Saúde EUA Florestal e programas de desenvolvimento tecnológico especiais que apoiaram o desenvolvimento, modificação e aplicação do protocolo descrito aqui para identificar árvores que são resistentes e suscetíveis ao inseto escala de faia. Também somos gratos ao Holden Arboretum, Kirtland, OH, por sua contínua parceria e apoio do nosso programa de triagem resistência faia americano.

Um material vegetal:. Mature Campo árvores, vasos de mudas, ou vasos de Enxertos <ol><li> Para os testes de campo, selecione árvores americanos saudáveis ​​maduros faia que não apresentem sinais de infestação de escala ou de doença para os testes de resistência possível. Visivelmente também terão de ser identificado para ser utilizado como um controlo (Figura 1) árvores susceptíveis. </li><li> Para testar mudas ou enxertos de vasos, recolher e germinar frutos da faia, conforme descrito no Koch &amp; Carey, 2004 ou descendente enxerto, conforme descrito no Carey et al, 2013. </li><li> Crescer mudas ou enxertos em envasamento mix solo com 47 g de micronutrientes, 477 g fertilizante de liberação lenta 15N-3.9P-9.9K, 700 g grosseiro perlita e 75 g de sulfato de alumínio por 2,8 cu. saco de pé. Se necessário mais tarde na estação de crescimento, fertilizar plantas semanais com solúvel 17N-1.3P-14.1K a 200 ppm de nitrogênio. </li><li> Durante a estação de crescimento, manter as plantas em uma casa de sombra. Permitem que as plantas vão dormentes fora no outono antes de movê-lospara uma instalação de armazenamento com temperatura controlada (~ 4 ° C) a partir de novembro a abril. </li></ol> 2. Recolha de Beech Escala Ovos <ol><li> Em um estande BBD-infestado, inspecionar árvores altamente infestados (facilmente identificados pela sua &quot;caiada&quot; aparência, Figura 1A), com uma lupa de mão para confirmar a presença de ovos, normalmente abundantes a partir de meados de julho a meados de agosto. </li><li> Use um pincel para escovar suavemente os pedaços de cera branca de cochonilhas de adultos, ovos e outros detritos, em um saco lacrado coleção um galão de plástico (Figura 1B). Colete de um mínimo de três árvores diferentes de pelo menos 12 m de distância. </li><li> Se necessário, armazenar os ovos no saco de recolha selado durante até duas semanas a 4 ° C. Um pequeno pedaço de fita (2,5 cm quadrados) de espuma de polietileno com amortecimento para dentro do saco, para impedir que os ovos de secar. </li><li> Para separar os ovos escala (0,15 x 0,25 mm) do que os adultos (0,60 mm) e debris, a construção de um peneiro utilizando uma peça curta de 2 &#39;de tubo de PVC e de um acoplamento para suportar uma peça quadrada de 250 micra de malha de nylon (Figura 2). </li><li> Esvaziar a mistura de adultos, cera, ovos e detritos a partir do saco de recolha para o crivo e utilizar um pequeno pincel de pintura para estimular suavemente os ovos para passar através da malha para uma placa de Petri de vidro abaixo. Pratos de plástico de Petri devem ser evitados porque eles detêm eletricidade mais estática, tornando-se difícil de mover os ovos. Ovos antes e após crivagem são mostrados na Figura 3. </li><li> Ovos purificada pode ser armazenada na caixa de Petri, a 4 ° C durante pelo menos uma semana. Para evitar que os ovos de secar, cole um pedaço úmido de espuma para a tampa e lacre com Parafilm. </li></ol> 3. Ovo Viabilidade Ensaios <ol><li> Para avaliar a viabilidade do ovo, com uma seringa de 10 cc para aplicar um anel fino de geleia de petróleo em torno da circunferência da parte inferior de uma placa de Petri de 60 milímetros de vidro (Figura 4A </strong&gt;). </li><li> Transferir cerca de 100 ovos para o centro do anel, coloque a tampa sobre a placa de Petri e selar com Parafilm. Permitir que a placa de Petri selado a permanecer sem perturbação à temperatura ambiente durante 2 semanas, ou 3 semanas, se os ovos foram armazenados a 4 ° C antes de se iniciar o ensaio. </li><li> Eclosão das ninfas vai ficar preso no vaselina e pode ser facilmente contado, e ovos vazias são facilmente distinguidos dos ovos não eclodidos por sua cor e brilho (Figura 4B). Calcular a viabilidade através da divisão do número de ninfas por a soma dos ovos vazios mais os restantes ovos completos. Boa viabilidade deve ser em 75% a 90% de ovos eclodidos gama. </li></ol> 4. Screening Resistência Escala de Grande Mature Faias no Campo <ol><li> Para um teste quantitativo, conte a 500 ovos usando um microscópio de dissecação e usando uma pequena espátula polvilhe-os suavemente em todo o centro de um pré-umedecido, de células abertas de 10 x 15 x 1.3 cm rectângulo de espuma de polietileno. Paraamortecer espuma, molhá-lo, em seguida, espremer o máximo de água possível. Para um teste qualitativo, 500 ovos pode ser contado para fora e colocados em um pequeno frasco de vidro e uma linha de &quot;enchimento&quot; desenhado que pode ser usado para medir lotes adicionais de cerca de 500 ovos. </li><li> Coloque a almofada de espuma para a árvore de teste com o lado do ovo virado contra a casca. Segure o disco no lugar com uma corda, corda, barbante ou fio de hardware plástico revestido. De plástico ou de material à base de metal que deve ser usado ao invés de materiais de fibras naturais, que são mais facilmente eliminado por animais selvagens. </li><li> Linha de cima e dos lados de um cm pedaço de vapor permeável envoltório casa à prova de água com adesivo de silicone de acetato com base pré-cortado 23 x 30 e coloque-a sobre a almofada de teste de espuma. Pressionar as bordas do envoltório casa da árvore para criar um selo à prova de água. Coloque fio de nylon ou arame revestido de plástico hardware ao redor da árvore e da casa envoltório, para segurá-la no lugar, enquanto os conjuntos adesivas (Figura 5). </li><li> Coloque um mínimo de doiscompressas de teste em cada árvore de ensaio, de preferência, em lados opostos do fuste. Em cada local, coloque almofadas de teste em pelo menos duas árvores suscetíveis (com infestação de escala natural, óbvio) como controle. Antes de colocar as almofadas de teste em árvores de controle, remova quaisquer insetos ou ovos escala que ocorrem naturalmente usando uma escova de cerdas firmes. Nota: as almofadas de espuma com ovos também pode ser colocado em árvores com o objectivo de ovos escala criação, o que pode ser particularmente útil em zonas onde o nível de infestação é baixo. </li></ol> 5. Screening Resistência Escala de vasos de mudas ou enxertos <ol><li> Para o teste, selecione árvores em vasos com diâmetro mínimo de 1 cm (pinça 5 cm acima linha do solo), que são altos o suficiente para ter pelo menos 2 almofadas de teste separados colocadas sobre eles. Podar pequenos ramos laterais quando necessário para dar espaço para as almofadas de teste. </li><li> Use um microscópio de dissecação para contar a 150 ovos e espalhá-los ao longo de um pré-cortados e umedecido 2,5 x 7,6 x 1,3 centímetros polyethy de células abertasalmofada de espuma lene. </li><li> Cole a almofada de espuma de mudas com o lado do ovo contra a casca, usando o fio revestido de plástico. Enrole um pequeno quadrado de casa envoltório ao redor da muda logo acima da espuma e selá-lo com silicone à base de acetato (Figura 6A). </li><li> Incluir conhecido famílias suscetíveis de mudas ou enxertos como controles. </li></ol> 6. Coleta de Dados Nota: Cerca de 52-57 semanas após a colocação dos blocos de espuma e ovos nas árvores de teste, os dados podem ser coletados. É importante que este não é feito até adultos após ter começado a pôr ovos de modo a que a sua capacidade para reproduzir pode ser determinada. Em algumas árvores resistentes, não é incomum ver um pequeno número de adultos, mas sem estabelecer reprodução ocorrendo. <ol><li> Remova cuidadosamente a almofada de espuma e contar o número de cochonilhas adulto estabelecidos na casca usando uma lente mão ou lupa (10X). </li><li> Quando a almofada de espuma é removido, ele não é uncommon para alguns adultos ea maioria dos aglomerados de ovos para ser puxado para fora da árvore com a espuma (ver Figura 7). Usando um microscópio de dissecação, a contagem dos aglomerados de ovos e adultos que tenham permanecido ligado à almofada de espuma. </li></ol>

<ol>
	<li>Houston, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+technique+to+artificially+infest+beech+bark+with+the+scale,+Cryptococcus+fagisuga+(Lindinger).">A technique to artificially infest beech bark with the scale, Cryptococcus fagisuga (Lindinger).</a> U.S. Forest Service. , (1982).</li><li>Houston, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=American+beech+resistance+to+Cryptococcus+fagisuga.">American beech resistance to Cryptococcus fagisuga.</a> Proceeding, IUFRO beech bark disease working party conference. , 38-42 (1983).</li><li>Ehrlich, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+beech+bark+disease:+a+nectria+disease+of+Fagus,+following+Cryptococcus+fagi.">The beech bark disease: a nectria disease of Fagus, following Cryptococcus fagi.</a> 10, 593-692 (1934).</li><li>Papaik, M. J., Canham, C. D., Latty, E. F., Woods, K. D. <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+an+introduced+pathogen+on+resistance+to+natural+disturbance:+beech+bark+disease+and+withdrow.">Effects of an introduced pathogen on resistance to natural disturbance: beech bark disease and withdrow.</a> 35, 1832-1843 (2005).</li><li>Miller-Weeks, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Current+status+of+beech+bark+disease+in+New+England+and+New+York.">Current status of beech bark disease in New England and New York.</a> Proceeding, IUFRO beech bark disease working party conference. , 21-23 (1983).</li><li>Houston, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Beech+bark+disease:+The+aftermath+forests+are+structured+for+a+new+outbreak.">Beech bark disease: The aftermath forests are structured for a new outbreak.</a> J. For. 73, 660-663 (1975).</li><li>Twery, M. J., Patterson, W. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Variations+in+beech+bark+disease+and+its+effects+on+species+composition+and+structure+of+northern+hardwood+stands+in+central+New.">Variations in beech bark disease and its effects on species composition and structure of northern hardwood stands in central New.</a> 14, 565-574 (1984).</li><li>Jakubus, W. J., McLaughlin, C. R., Jensen, P. G., McNulty, S. A., Evans, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alternate+year+beechnut+production+and+its+influence+on+bear+and+marten+populations.">Alternate year beechnut production and its influence on bear and marten populations.</a> Beech Bark Disease: Proceedings of the Beech Bark Disease Symposium. , 79-87 (2005).</li><li>Koch, J. L., Carey, D. W., Mason, M. E., Nelson, C. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+beech+scale+resistance+in+full-+and+half-sib+families.">Assessment of beech scale resistance in full- and half-sib families.</a> Can. J. For. Res. 40, 265-272 (2010).</li><li>Koch, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Beech+bark+disease:+The+oldest+&#8220;new&#8221;+threat+to+American+beech+in+the+United+States.">Beech bark disease: The oldest &#8220;new&#8221; threat to American beech in the United States.</a> Outlooks Pest Manag. 21, 64-68 (2010).</li><li>Koch, J. L., Mason, M. E., Carey, D. W., Palmieri, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Screening+for+resistance+to+beech+bark+disease:+improvements+and+results+from+seedlings+and+grafted+field+selections.">Screening for resistance to beech bark disease: improvements and results from seedlings and grafted field selections.</a> Proceedings of the 4th International Workshop on Genetics of Host-Parasite Interactions in Forestry. , (2012).</li><li>Leak, W. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fifty+year+impacts+of+the+beech+bark+disease+in+the+Bartlett+Experimental+Forest,+New+Hampshire.">Fifty year impacts of the beech bark disease in the Bartlett Experimental Forest, New Hampshire.</a> Northern J. Appl. For. 23, 141-143 (2006).</li><li>Koch, J. L., Carey, D. W., Yaussy, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Controlled+cross-pollinations+with+American+beech+trees+that+are+resistant+to+beech+bark+disease.">Controlled cross-pollinations with American beech trees that are resistant to beech bark disease.</a> Proceedings of the 14th Central Hardwood Forest Conference. , 358-364 (2004).</li><li>Carey, D. W., Mason, M. E., Bloese, P., Koch, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hot+callusing+for+propagation+of+American+beech+by+grafting.">Hot callusing for propagation of American beech by grafting.</a> HortSci. 48, 620-624 (2013).</li><li>Koch, J. L., Carey, W. W., Evans, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+genetics+of+resistance+of+American+beech+to+beech+bark+disease:+knowledge+through+2004.">The genetics of resistance of American beech to beech bark disease: knowledge through 2004.</a> Beech Bark Disease: Proceedings of the Beech Bark Disease Symposium. , 98-105 (2005).</li></ol>

Os autores declaram que não têm interesses financeiros concorrentes.

Os passos críticos necessários para o sucesso deste ensaio incluem a execução de um teste de viabilidade sobre os ovos e a utilização de controlos sensíveis em cada local de ensaio, e com materiais de vasos. Também é importante o uso de mais de um bloco de teste por árvore ou mudas. Descobrimos que, tanto no campo e em testes mais controlados sobre vasos de plantas, uma fonte freqüente de erro pode ser falha pad. Por exemplo, no campo, mesmo com a modificação de que cobre a almofada de ensaio com o envoltó...

Doença casca Beech (BBD) teve um impacto negativo sobre a faia americana na América do Norte desde a introdução do inseto escala faia invasivo, Cryptococcus fagisuga, na província canadense de Nova Escócia, no final de 1890 3. Este complexo de insetos-doença é iniciada quando o inseto escala faia insere seu estilete alimentação na casca criando pequenas fissuras que fornecem entrada para a infecção por uma das espécies de fungos Neonectria (Neonectria ditissima ou Neonectria faginata). Tal como os micélios fúngicos crescer, grandes áreas de tecido podem morrer, eventualmente anelamento completamente a árvore. Os danos da doença enfraquece a árvore, tornando-se propenso à rotura no 4 ventos fortes. Os níveis de mortalidade na primeira onda da doença foram relatados para ser tão alta quanto 50% 5. Árvores sobreviventes são muitas vezes deformado severamente como cancros formar reduzindo o valor da árvore como um produto de madeira.0; Essas árvores têm uma propensão para o root-brotação que leva à formação de &quot;moitas faia&quot; que impedem outras espécies mais desejáveis ​​de estabelecer, reduzindo o valor econômico e ecológico do stand 6. Embora a doença de casca de faia não é susceptível de conduzir à extinção da faia americana, altera ficar composição e saúde levando a uma diminuição nos alimentos e habitat para a vida selvagem 7,8. Em povoamentos afectados por BBD há muitos anos, as árvores que permanecem livres de quaisquer sintomas da doença têm sido relatados. Ensaios inoculação artificial têm demonstrado que estas árvores são resistentes ao inseto escala 2. Sem infestação escala, há poucas oportunidades para infecção Neonectria, minimizando o impacto do fungo. Mortalidade em grande escala em faia americano devido à infecção Neonectria na ausência de infestação de escala antes nunca temforam relatados, por isso os resultados de resistência a insectos escala faia em resistência à BBD. Pesquisas recentes sobre a gestão de BBD centrou-se na identificação, propagação, reprodução e retenção de faias americanas com resistência ao inseto escala de faia. Estudos genéticos mostraram que a resistência ao inseto escala é hereditária e cuidadosa seleção e melhoramento de árvores resistentes podem resultar em melhorias significativas em uma única geração 9. Este achado tem alimentado os esforços de gestores estaduais e floresta nacional nos Estados Unidos para estabelecer pomares regionais de sementes de faia americano resistente para fornecer uma fonte de diversidade genética de sementes resistentes BBD para plantios de restauração 10,11. A pesquisa também indicou que a manipulação genética silvicultural de suporte através da remoção de árvores susceptíveis e retenção de árvores resistentes podem resultar em stand melhoria 9,12. &lt;/ P&gt; Gestão de BBD através de atividades de melhoria de árvores ou através da realização de prescrições silviculturais requer a capacidade de selecionar para e distinguir entre árvores escala resistentes e suscetíveis de faia. Os métodos aqui apresentados foram adaptados a partir de um método introduzido pela primeira vez por Dave Houston para inocular artificialmente mudas com ovos escala faia 1. O método pode ser utilizado como uma ferramenta de rastreio para identificar loci característica quantitativa (QTL) associado com a resistência ou para distinguir entre mudas vasos resistentes e sensíveis ou rametes enxertados em estudos genéticos. Alternativamente, ele pode ser usado para o rastreio de árvores adultas no campo para identificar plantas resistentes para o desenvolvimento de um pomar de sementes, ou a retenção no campo. Árvores sensíveis, podem ser identificados e removidos para minimizar os impactos das doenças.

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a technique to screen american beech for resistance to the beech scale insect (cryptococcus fagisuga lind.)

Beech doença casca (BBD) resulta em altos níveis de mortalidade inicial, deixando para trás árvores sobreviventes que estão muito enfraquecidos e deformados. A doença é iniciada pela alimentação de actividades da cochonilha faia invasiva, Cryptococcus fagisuga, o que gera os pontos de entrada para a infecção por uma das espécies Neonectria de fungo. Sem infestação escala, há pouca oportunidade para a infecção fúngica. Usando ovos escala para infestar artificialmente árvores saudáveis ​​em povoamentos fortemente impactadas BBD demonstrado que essas árvores eram resistentes à parcela inseto escala do complexo de doenças 1. Aqui apresenta-se um protocolo que temos desenvolvido, com base na técnica de infestação artificial de Houston 2, o qual pode ser utilizado para rastrear para árvores de escala-resistente no campo e em pequenas plântulas envasadas e enxertos. A identificação de árvores de escala-resistente é uma componente importante da gestão da BBD através da árvoreprogramas de melhoria e manipulação silvicultural.

A Figura 6 mostra uma plântulas resistentes (C) e duas mudas susceptíveis (D, E) exibem diferentes graus de susceptibilidade. Um exemplo do que uma árvore madura susceptível parece 57 semanas após o teste de infestação artificial foi configurado é mostrado na Figura 5B. Quando a espuma é retirado da árvore, não é incomum para cochonilhas e seus cachos de ovos para permanecer preso para a espuma, como mostrado na Figura 7, qu...

Watch this Scientific Journal Video about A Technique to Screen American Beech for Resistance to the Beech Scale Insect Cryptococcus fagisuga Lind. at JoVE.com

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

Doença casca Beech é iniciada através da alimentação atividades do inseto escala de faia que criar pontos de entrada de fungos na casca. Árvores que são resistentes ao inseto escala também são resistentes às doenças. Aqui apresentamos o protocolo que temos desenvolvido para a tela faias individuais de resistência à escala de faia.

Uma técnica para Screen americana Beech para Resistência ao Beech Scale Insect (<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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Research

JoVE Journal

Environment

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

9.5K visualizações.  US Forest Service.  Doença casca Beech é iniciada através da alimentação atividades do inseto escala de faia que criar pontos de entrada de fungos na casca. Árvores que são resistentes ao inseto escala também são resistentes às doenças. Aqui apresentamos o protocolo que temos desenvolvido para a tela faias individuais de resistência à escala de faia. 

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

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

Despite being one of the most productive fisheries in the Northwest Atlantic, much remains unknown about the natural reproductive dynamics of American lobsters. Recent work in exploited crustacean populations (crabs and lobsters) suggests that there are circumstances where mature females are unable to achieve their full reproductive potential due to sperm limitation. To examine this possibility in different regions of the American lobster fishery, a reliable and noninvasive method was developed for sampling large numbers of female lobsters at sea. This method involves inserting a blunt-tipped needle into the female&#39;s seminal receptacle to determine the presence or absence of a sperm plug and to withdraw a sample that can be examined for the presence of sperm. A series of control studies were conducted at the dock and in the laboratory to test the reliability of this technique. These efforts entailed sampling 294 female lobsters to confirm that the presence of a sperm plug was a reliable indicator of sperm within the receptacle and thus, mating. This paper details the methodology and the results obtained from a subset of the total females sampled. Of the 230 female lobsters sampled from George&#39;s Bank and Cape Ann, MA (size range = 71-145 mm in carapace length), 90.3% were positive for sperm. Potential explanations for the absence of sperm in some females include: immaturity (lack of physiological maturity), breakdown of the sperm plug after being used to fertilize a clutch of eggs, and lack of mating activity. The surveys indicate that this technique for examining the mating success of female lobsters is a reliable proxy that can be used in the field to document reproductive activity in natural populations.

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

Fishery-induced changes to exploited crustacean fisheries, such as the American lobster fishery, could potentially influence their reproductive dynamics, leading to a reduction in mating success. This study&#39;s goal was to develop a noninvasive method for ascertaining the mating success of female lobsters that may be physiologically or functionally mature.

Um método não invasivo para<em&gt; Em situ</em&gt; Determinação do sucesso de acasalamento em fêmeas lagostas americanas (<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 ...

Ambiente

Video Tracking Protocol to Screen Deterrent Chemistries for Honey Bees

The European honey bee, Apis mellifera L., is an economically and agriculturally important pollinator that generates billions of dollars annually. Honey bee colony numbers have been declining in the United States and many European countries since 1947. A number of factors play a role in this decline, including the unintentional exposure of honey bees to pesticides. The development of new methods and regulations are warranted to reduce pesticide exposures to these pollinators. One approach is the use of repellent chemistries that deter honey bees from a recently pesticide-treated crop. Here, we describe a protocol to discern the deterrence of honey bees exposed to select repellent chemistries. Honey bee foragers are collected and starved overnight in an incubator 15 h prior to testing. Individual honey bees are placed into Petri dishes that have either a sugar-agarose cube (control treatment) or sugar-agarose-compound cube (repellent treatment) placed into the middle of the dish. The Petri dish serves as the arena that is placed under a camera in a light box to record the honey bee locomotor activities using video tracking software. A total of 8 control and 8 repellent treatments were analyzed for a 10 min period with each treatment was duplicated with new honey bees. Here, we demonstrate that honey bees are deterred from the sugar-agarose cubes with a compound treatment whereas honey bees are attracted to the sugar-agarose cubes without an added compound.

The loss of honey bee colonies presents a challenge to crop pollination services. Current pollinator protection practices warrant an alternative approach to minimize the contact of honey bees to harmful pesticides using repellent chemistries. Here, we provide detailed methods for a visual tracking protocol to screen deterrents for bees.

Protocolo de rastreamento de vídeo para a tela de quimioterapia para abelhas

The European honey bee, Apis mellifera L., is an economically and agriculturally important pollinator that generates billions of dollars annually. Honey ...

Lab-Scale Model to Evaluate Odor and Gas Concentrations Emitted by Deep Bedded Pack Manure

A lab-scaled simulated bedded pack model was developed to study air quality and nutrient composition of deep-bedded packs used in cattle mono-slope facilities. This protocol has been used to effectively evaluate many different bedding materials, environmental variables (temperature, humidity), and potential mitigation treatments that can improve air quality in commercial deep-bedded mono-slope facilities. The model is dynamic and allows researchers to easily collect many chemical and physical measurements from the bedded pack. Weekly measurements, collected over the course of six to seven weeks, allows sufficient time to see changes in air quality measurements over time as the bedded pack matures. The data collected from the simulated bedded packs is within the range of concentrations previously measured in commercial deep-bedded mono-slope facilities. Past studies have demonstrated that 8 - 10 experimental units per treatment are sufficient to detect statistical differences among the simulated bedded packs. The bedded packs are easy to maintain, requiring less than 10 minutes of labor per bedded packs per week to add urine, feces, and bedding. Sample collection using the gas sampling system requires 20 - 30 minutes per bedded pack, depending on the measurements that are being collected. The use of lab-scaled bedded packs allows the researcher to control variables such as temperature, humidity, and bedding source that are difficult or impossible to control in a research or commercial facility. While not a perfect simulation of &#34;real-world&#34; conditions, the simulated bedded packs serve as a good model for researchers to use to examine treatment differences among bedded packs. Several lab-scale studies can be conducted to eliminate possible treatments before trying them in a research or commercial-sized facility.

A protocol has been developed to measure gases, odors, and nutrient composition in lab-scaled bedded manure packs, which can be used to study ways to improve air quality in commercial cattle facilities using deep-bedded manure packs.

Laboratório-escala modelo para avaliar o Odor e as concentrações de gases emitidas pelo profundo com camas Pack estrume

A lab-scaled simulated bedded pack model was developed to study air quality and nutrient composition of deep-bedded packs used in cattle mono-slope ...

Double-stranded RNA Oral Delivery Methods to Induce RNA Interference in Phloem and Plant-sap-feeding Hemipteran Insects

Phloem and plant sap feeding insects invade the integrity of crops and fruits to retrieve nutrients, in the process damaging food crops. Hemipteran insects account for a number of economically substantial pests of plants that cause damage to crops by feeding on phloem sap. The brown marmorated stink bug (BMSB), Halyomorpha halys (Heteroptera: Pentatomidae) and the Asian citrus psyllid (ACP), Diaphorina citri Kuwayama (Hemiptera: Liviidae) are hemipteran insect pests introduced in North America, where they are an invasive agricultural pest of high-value specialty, row, and staple crops and citrus fruits, as well as a nuisance pest when they aggregate indoors. Insecticide resistance in many species has led to the development of alternate methods of pest management strategies. Double-stranded RNA (dsRNA)-mediated RNA interference (RNAi) is a gene silencing mechanism for functional genomic studies that has potential applications as a tool for the management of insect pests. Exogenously synthesized dsRNA or small interfering RNA (siRNA) can trigger highly efficient gene silencing through the degradation of endogenous RNA, which is homologous to that presented. Effective and environmental use of RNAi as molecular biopesticides for biocontrol of hemipteran insects requires the in vivo delivery of dsRNAs through feeding. Here we demonstrate methods for delivery of dsRNA to insects: loading of dsRNA into green beans by immersion, and absorbing of gene-specific dsRNA with oral delivery through ingestion. We have also outlined non-transgenic plant delivery approaches using foliar sprays, root drench, trunk injections as well as clay granules, all of which may be essential for sustained release of dsRNA. Efficient delivery by orally ingested dsRNA was confirmed as an effective dosage to induce a significant decrease in expression of targeted genes, such as juvenile hormone acid O-methyltransferase (JHAMT) and vitellogenin (Vg). These innovative methods represent strategies for delivery of dsRNA to use in crop protection and overcome environmental challenges for pest management.

This article demonstrates novel techniques developed for oral delivery of double-stranded RNA (dsRNA) through the vascular tissues of plants for RNA interference (RNAi) in phloem sap feeding insects.

Métodos de entrega Double-stranded RNA Oral para induzir a interferência do RNA no floema e insetos Hemipteran planta-sap-alimentação

Phloem and plant sap feeding insects invade the integrity of crops and fruits to retrieve nutrients, in the process damaging food crops. Hemipteran ...

The Barnacle Balanus improvisus as a Marine Model - Culturing and Gene Expression

Barnacles are marine crustaceans with a sessile adult and free-swimming, planktonic larvae. The barnacle Balanus (Amphibalanus) improvisus is particularly relevant as a model for the studies of osmoregulatory mechanisms because of its extreme tolerance to low salinity. It is also widely used as a model of settling biology, in particular in relation to antifouling research. However, natural seasonal spawning yields an unpredictable supply of cyprid larvae for studies. A protocol for the all-year-round culturing of B. improvisus has been developed and a detailed description of all steps in the production line is outlined (i.e., the establishment of adult cultures on panels, the collection and rearing of barnacle larvae, and the administration of feed for adults and larvae). The description also provides guidance on troubleshooting and discusses critical parameters (e.g., the removal of contamination, the production of high-quality feed, the manpower needed, and the importance of high-quality seawater). Each batch from the culturing system maximally yields roughly 12,000 nauplii and can deliver four batches in a week, so up to almost 50,000 larvae per week can be produced. The method used to culture B. improvisus is, probably, to a large extent also applicable to other marine invertebrates with free-swimminglarvae. Protocols are presented for the dissection of various tissues from adults as well as the production of high-quality RNA for studies on gene expression. It is also described how cultured adults and reared cyprids can be utilized in a wide array of experimental designs for examining gene expression in relation to external factors. The use of cultured barnacles in gene expression is illustrated with studies of possible osmoregulatory roles of Na+/K+ ATPase and aquaporins.

The Barnacle Balanus improvisus as a Marine Model - Culturing and Gene Expression

The barnacle Balanus (Amphibalanus) improvisus is a model for studying osmoregulation and antifouling. However, natural seasonal spawning yields an unpredictable supply of cyprid larvae. Here, a protocol for the all-year-round culturing of B. improvisus is described, including the production of larvae. The use of cultured barnacles in gene expression studies is illustrated.

A craca Balanus improvisus como modelo Marine - cultivo e expressão gênica

Barnacles are marine crustaceans with a sessile adult and free-swimming, planktonic larvae. The barnacle Balanus (Amphibalanus) improvisus is particularly ...

Experimental Protocol for Examining Behavioral Response Profiles in Larval Fish: Application to the Neuro-stimulant Caffeine

Fish models and behaviors are increasingly used in the biomedical sciences; however, fish have long been the subject of ecological, physiological and toxicological studies. Using automated digital tracking platforms, recent efforts in neuropharmacology are leveraging larval fish locomotor behaviors to identify potential therapeutic targets for novel small molecules. Similar to these efforts, research in the environmental sciences and comparative pharmacology and toxicology is examining various behaviors of fish models as diagnostic tools in tiered evaluation of contaminants and real-time monitoring of surface waters for contaminant threats. Whereas the zebrafish is a popular larval fish model in the biomedical sciences, the fathead minnow is a common larval fish model in ecotoxicology. Unfortunately, fathead minnow larvae have received considerably less attention in behavioral studies. Here, we develop and demonstrate a behavioral profile protocol using caffeine as a model neurostimulant. Though photomotor responses of fathead minnows were occasionally affected by caffeine, zebrafish&#160;were markedly more sensitive for photomotor and locomotor endpoints, which responded at environmentally relevant levels. Future studies are needed to understand comparative behavioral sensitivity differences among fish with age and time of day, and to determine whether similar behavioral effects would occur in nature and be indicative of adverse outcomes at the individual or population levels of biological organization.

Here, we present a protocol to examine larval zebrafish and fathead minnow locomotor activities and photomotor responses (PMR) using an automated tracking software. When incorporated in common toxicology bioassays, analyses of these behaviors provide a diagnostic tool to examine chemical bioactivity. This protocol is described using caffeine, a model neurostimulant.

Protocolo experimental pela análise de perfis de resposta comportamental em peixes Larval: aplicação para a Neuro-estimulantes cafeína

Fish models and behaviors are increasingly used in the biomedical sciences; however, fish have long been the subject of ecological, physiological and ...

A Precise and Autonomous System for the Detection of Insect Emergence Patterns

Existing systems to measure insect emergence patterns have limitations; they are only partially automated and are limited in the maximum number of emerging insects they can detect. In order to obtain precise measurement of insect emergence, it is necessary for systems to be semi-automated and able to measure large numbers of emerging insects. We addressed these issues by designing and building a system that is automated and can measure emergence of up to 1200 insects. We modified the existing &#34;falling-ball&#34; system using Arduino microcontrollers to automate data collection and expand the sample size through multiple data channels. Multiple data channels enable the user to not only increase their sample size, but also allows for multiple treatments to be run simultaneously in a single experiment. Furthermore, we created an R script to automatically visualize the data as a bubble plot, while also calculating the median day and time of emergence. The current system was designed using 3D printing so the user can modify the system to be adjusted for different species of insects. The goal of this protocol is to investigate important questions in chronobiology and stress physiology, using this precise and automated system to measure insect emergence patterns.

Measurement of insect emergence patterns requires precision. Existing systems are only semi-automated and sample size is limited. We addressed these issues by designing a system using microcontrollers to precisely measure the time of emergence of large numbers of emerging insects.

Um sistema autónomo e preciso para a detecção de padrões de emergência de insetos

Existing systems to measure insect emergence patterns have limitations; they are only partially automated and are limited in the maximum number of ...

Composition and Distribution Analysis of Bioaerosols Under Different Environmental Conditions

Variable microorganisms in particulate matter (PM) under different environmental conditions may have significant impacts on human health. In this study, we described a protocol for multiple analyses of the biological compositions in environmental PM. Five experiments are presented: (1) PM number monitoring by using a laser particle counter; (2) PM collection by using a cyclonic aerosol sampler; (3) PM collection by using a high-volume air sampler with filters; (4) culturable microbes collection by the Andersen six-stage sampler; and (5) detection of biological composition of environmental PM by bacterial 16SrDNA and fungal ITS region sequencing. We selected hazy days and a livestock farm as two typical examples of the application in this protocol. In this study, these two sampling methods, cyclonic aerosol sampler and filter sampler, showed different sampling efficiency. The cyclonic aerosol sampler performed much better in terms of collecting bacteria, while these two methods showed the same efficiency in collecting fungi. Filter samplers can work under low temperature conditions while cyclonic aerosol samplers have a sampling limitation for temperature. A solid impacting sampler, such as an Andersen six-stage sampler, can be used to sample bioaerosols directly into the culture medium, which increases the survival rate of culturable microorganisms. However, this method mainly relies on culture while more than 99% of microbes cannot be cultured. DNA extracted from the culturable bacteria collected by the Andersen six-stage sampler and samples collected by cyclonic aerosol sampler and filter sampler were detected by bacterial 16S rDNA and fungal ITS region sequencing.All the methods above may have wide application in many fields of study, such as environmental monitoring and airborne pathogen detection. From these results, we can conclude that these methods can be used under different conditions and may help other researchers further explore the health impacts of environmental bioaerosols.

Understanding the biological composition of environmental particulate matter is important for the study of its significant impacts on human health and disease spread. Here, we used three types of bioaerosol sampling methods and a biological analysis of airborne microbes to better explore airborne microbial communities under different environmental conditions.

Composição e análise da distribuição de Bioaerossóis sob diferentes condições ambientais

Variable microorganisms in particulate matter (PM) under different environmental conditions may have significant impacts on human health. In this study, ...

Detached Leaf Assays to Simplify Gene Expression Studies in Potato During Infestation by Chewing Insect Manduca sexta

The multitrophic nature of gene expression studies of insect herbivory demands large numbers of biological replicates, creating the need for simpler, more streamlined herbivory protocols. Perturbations of chewing insects are usually studied in whole plant systems. While this whole organism strategy is popular, it is not necessary if similar observations can be replicated in a single detached leaf. The assumption is that basic elements required for signal transduction are present within the leaf itself. In the case of early events in signal transduction, cells need only to receive the signal from the perturbation and transmit that signal to neighboring cells which are assayed for gene expression.
The proposed method simply changes the timing of the detachment. In whole plant experiments, larvae are confined to a single leaf which is eventually detached from the plant and assayed for gene expression. If the order of excision is reversed, from last in whole plant studies, to first in the detached study, the feeding experiment is simplified.
Solanum tuberosum var. Kennebec is propagated by nodal transfer in a simple tissue culture medium and transferred to soil for further growth if desired. Leaves are excised from the parent plant and relocated to Petri dishes where the feeding assay is conducted with the larval stages of M. sexta. Damaged leaf tissue is assayed for the expression of relatively early events in signal transduction. Gene expression analysis identified infestation-specific Cys2-His2 (C2H2) transcription factors, confirming the success of using detached leaves in early response studies. The method is easier to perform than whole plant infestations and uses less space.

The presented method creates natural herbivore damaged plant tissue through the application of Manduca sexta larvae to detached leaves of potato. The plant tissue is assayed for expression of six transcription factor homologs involved in early responses to insect herbivory.

Ensaios foliares destacados para simplificar estudos de expressão gênica em batata durante a infestação por mastigação de insetos Manduca sexta

The multitrophic nature of gene expression studies of insect herbivory demands large numbers of biological replicates, creating the need for simpler, more ...

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.

Implantação de uma nova marca acústica Micro no juvenil Pacífico lampreia e enguia americana

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