Agradecemos a Keith Shigeteni de contribuição no insetário e Sostrom Natasha para ajudar com computação gráfica. Este trabalho foi financiado pelo USDA-ARS. Opiniões, resultados, conclusões ou recomendações expressas nesta publicação são as dos autores e não refletem necessariamente a opinião do USDA. O USDA é um provedor de igualdade de oportunidades e empregador.

1. Prepare ovos do hospedeiro para o parasitismo mosca da fruta <ol><li> Prepare um substrato para os ovos de mosca de fruta a ser parasitados por preparar agar (alternativamente, Gelcarin GP812, FMC Biopolymer, Ewing NJ; uma opção mais rentável) pratos cheios, 10 cm de comprimento de cada lado e 1,5 cm de profundidade. Prepare ágar na concentração de 9 g por litro de água. </li><li> Preencha pratos até a borda com ágar líquido (cerca de 70ml) e deixe esfriar e solidificar (pelo menos 45 minutos). </li><li> Uma vez que os blocos de agar são sólidos, aplique uma camada de papel de seda (uma única camada de tecido &#39;Preferências&#39; Georgia Pacific lote), cobrindo a parte superior de cada bloco. O tecido deve ficar facilmente para o agar por causa da umidade da superfície. </li><li> Aplicar um volume de 0,5 ml de ovos de mosca de fruta em suspensão na água para a superfície coberta de tecido dos blocos de ágar. No nosso insetário, isso é equivalente a cerca de 6000 ovos de Bactrocera dorsalis (12000 ovos por ml). </li><li> Use um pano limpo 2,5 centímetros de largura pincel umedecido em água para espalhar os ovos mosca da fruta uniformemente sobre a superfície do tecido. </li><li> Coloque os ovos de mosca de fruta sob a tela de fundo do parasitóide gaiolas para permitir a oviposição. Toque a parte inferior da tela de fundo para remover parasitas mortos que podem obstruir os ovos. É melhor se a tela de baixo não toca os ovos. </li><li> Permitir que os parasitas para oviposit durante a noite e no dia seguinte, cerca de 21 horas. Isso é tempo suficiente para que uma alta porcentagem dos ovos a serem parasitados. Exposição mais longa superparasitization riscos. </li></ol> 2. Incubação e de pupação fruto hospedeiro parasitado moscas <ol><li> Prepare criação recipiente, colocando 3-3,5 blocos de ágar preparado de ovos parasitados por 1,25 litros de dieta de frutas fly 19 com ovos voltada para cima. </li><li> Coloque o recipiente dieta com ovos em um riser sobre uma camada de 1,5 cm de profundidade grau de vermiculita fina ou areia lavada em um recipiente maior pupação. Cubra o recipiente pupação com uma tampa e fita adesiva nas bordas da fita adesiva com. Vermiculita pode ser reutilizado após peneiração (veja abaixo). </li><li> Verifique se há espaço suficiente entre a parte superior do recipiente dieta e a tampa do frasco pupação de larvas de mosca se arrastar para fora e &#39;pop&#39; 20 no vemiculite abaixo. </li><li> Mover recipientes pupação para uma sala a 27 ° C e 80% de umidade relativa (UR) por uma semana. Recipientes cubra com um pano escuro para os primeiros quatro dias depois remover o tecido para os dias restantes. </li><li> Remover dieta larval após uma semana, uma vez que as larvas estalaram e entrou na vermiculita para pupação. Se houver larvas nos vermiculite, permitir-lhes pupate durante a noite. </li><li> Peneira as pupas da vermiculita. Comece com uma peneira grossa (cerca de 3 mm), em seguida, use uma peneira para remover qualquer lado restantes aglomerados de vermiculita. </li></ol> 3. Classificação e seleção <ol><li> Pupa contendo F. arisanus pode ser parcialmente segregado de moscas de frutas unparasitized por tamanho. Coletar pupa entre 1,65 e 2,26 milímetros de diâmetro, como estes irão ser enriquecido para moscas parasitadas. 12 pupários que são menores contêm principalmente parasitóides subdimensionado, pupa maiores contêm principalmente moscas. Note também que para as pupas parasitadas a porcentagem de fêmeas se correlaciona positivamente com o tamanho 12. </li><li> Uma variedade de métodos podem ser empregados para classificar pupas por tamanho. Vamos descrever um método mecânico usando um tamanho personalizado classificador 21. </li><li> Pupas lugar no funil no topo de um alimentador de vibração que lentamente alimenta o classificador de tamanho. </li><li> Alimentar o pupas no classificador de tamanho, que consiste de um par de barras de rolamento ligeiramente divergentes em um ângulo de 30 °. A queda de pupas individualmente por tamanho em uma matriz de slots que vazio em dez recipientes separados através de um funil. </li><li> Recolher os copos de pupas na faixa de tamanho de 1,65-2,26 mm e colocá-los em uma gaiola de emergência por mais 7 dias até que a maioria moscas emergiram as pupas unparasitized. </li><li> Em seguida, use um ventilador para separar pupários vazios daqueles insetos contendo ainda. </li></ol> 4. F. arisanus surgimento e manutenção <ol><li> Para manter a colônia de F. arisanus usar uma gaiola parasitóide segurando com uma frente de vidro removível, cerca de 25 cm de comprimento de cada lado. A gaiola tem um milímetro de triagem dois na parte superior e dois dos lados. A parte de trás da gaiola é coberto por um lençol de borracha com um furo de 9 cm para o acesso. O fundo da gaiola tem um recorte cobertas por dentro com 1-2mm de tela 2 grandes o suficiente para permitir a colocação de dois blocos de agar com ovos de mosca de fruta para oviposição do parasitóide. </li><li> As gaiolas holding também deve ter uma porção coloridas ao longo do trimestre inferior da fachada de vidro para minimizar a luz que pode criar crowding parasitóide ao longo da frente da gaiola, resultando em mortalidade elevada. </li><li> Coloque cerca de 11 g desses seleçãoted pupas em recipientes plásticos de aproximadamente 9 cm de diâmetro. Isso deve produzir cerca de 600-700 parasitas por gaiola. Seleção de tamanho deve resultar em cerca de 60% do sexo feminino. </li><li> Anexar tampas selecionados para os recipientes. As tampas devem ser cobertos com 2 mm 2 de triagem, o que permitirá adultos F. arsianus para deixar os recipientes após a emergência, mas, conterá qualquer moscas de frutas restantes. </li><li> Manter os parasitas a 24 ° C e cerca de 45% RH com uma ventilação 12:12 fotoperíodo e bom. Se a mortalidade alta é observada, movendo-se as gaiolas ao ar livre por algumas horas por dia ou ventilação pode aumentar a reduzi-lo. Parasitas deve estar pronto para oviposit em uma semana adicional e devem ser produtivo por mais duas semanas. </li><li> Streak não diluído mel girado ao longo do topo das gaiolas, pelo menos, três vezes por semana ou sempre que o mel disponível é seco. Aplique o mel girou em estrias estreitas usando um dedo. Note que, se as estrias são muito pesados ​​o mel vai gotejar para dentro da gaiola, se muito leve o mel vai secar rapidamente e exigem reaplicação freqüente. </li><li> Colocar os blocos de ágar (10 cm x 10 cm x 4 cm) nos topos das gaiolas para fornecer umidade para os parasitas. Estes devem ser mudadas duas vezes por semana. </li></ol> 5. Resultados representativos: Relatamos aqui os resultados de procedimentos de controle de qualidade realizados no F. USDA-ARS arisanus operação de criação em Hilo, Havaí entre 05 de janeiro e 22 junho de 2010. Criação no local de Hilo na escala descrita neste artigo de vídeo e protocolo foi iniciado em agosto de 2009, e os problemas iniciais e ajustes com o novo local tinha sido na sua maioria resolvidos por janeiro. Portanto, estes dados são representativos dos resultados de outros pesquisadores podem obter a partir de sua colônia, se seguir o protocolo, conforme descrito e ter experiência de criação de insetos em geral. Durante este período, fomos mantendo uma pequena colônia de F. arisanus: 4 parasitóide gaiolas segurando produzidos por semana, equivalente a cerca de 3.600 F. arisanus. Uma verificação inicial das taxas de parasitismo enriquecido foi conduzido tomando uma amostra de 2 g de pupas na faixa de tamanho de 1,65-2,26 mm (enriquecido), antes de mais uma semana de desenvolvimento em uma gaiola de retenção (isto é, imediatamente após a dieta larval foi removido e os pupas peneirado e tamanho classificadas. Referimo-nos a estas como &#39;pupas cedo &quot;). Durante os primeiros três meses de 2010 os parasitas significa proporção na amostra enriquecida foi de 0,46 (DP = 0,18). Durante o trimestre seguinte esta média foi de 0,58 (DP = 0,08), refletindo a estabilização dos procedimentos e criação de colônias no novo local. Note-se que as pupas enriquecido, que não estavam parasitados com sucesso incluem aqueles que não produzem emergentes e aqueles que produziram as moscas. Após mais uma semana na remoção gaiola realização de pupários vazios outra amostra de grama dois anos foi tomada (&#39;pupas tarde&#39;). Moscas da fruta representava menos de 1% desta amostra. O baixo percentual de moscas da fruta é parcialmente devido a com o anterior aparecimento de moscas em comparação com parasitas, assim a maioria voa permaneceu no recipiente de exploração. Finalmente, a Figura 1 apresenta o rendimento em massa de pupa de janeiro a junho de 2010. Os rendimentos são medidos a partir de pupas enriquecido na faixa de tamanho de interesse, 1,65-2,26 mm. A grande massa de &quot;início de pupas &#39;no início do ano indica uma alta proporção de moscas de frutas e pupas unemerged naquela época, enquanto que os ajustes foram sendo feitos para as novas instalações. <img src="/files/ftp_upload/2901/2901fig1.jpg" alt="Figura 1" /> Figura 1. Rendimento de enriquecido (pupal diâmetro 1,65 2,26 milímetros) precoce (imediatamente após a seleção do tamanho) e &#39;atrasado&#39; pupas (após a realização de uma semana e remover carcaças vazias) na ARS-USDA colônia ariasnus F. em Hilo, Hawaii , Janeiro - Junho 2010

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Nós não temos conflitos de interesse para relatar.

Neste artigo de vídeo protocolo e acompanhamento que temos descrito e demonstrado um protocolo otimizado para a criação de F. ariasnus, um parasitóide de moscas da fruta Tephritid, em um ambiente de laboratório. Este protocolo tem sido aperfeiçoado ao longo dos anos, para minimizar a quantidade de equipamentos de trabalho e especializado necessário para manter uma colônia. Notamos que uma bem estabelecida colônia, produtiva e estável de moscas da fruta host é necessária para qualquer tentativa de criação de parasitóides. No nosso insetário usamos Bactrocera dorsalis como uma mosca de fruta de acolhimento, mas outras espécies têm sido mostrados para ser os anfitriões competentes, bem 14-17. Vários aspectos da F. arisanus manutenção da colônia ainda precisam ser exploradas. Estes incluem a taxa de adaptação desta espécie de laboratório em 10 de mecanismos de aprendizagem 22 e as mudanças genéticas que podem ocorrer com adaptação e mecanismos que podem estar envolvidos na plasticidade anfitrião mosca da fruta. Nos últimos dez anos houve vários exemplos da propagação de moscas Dacine todo o mundo, em especial os da Bactrocera género: B. dorsalis na Polinésia Francesa, B. carambolae em partes da América do Sul, B. invadens na África e B. zonata na África e Mediterrâneo do Norte. Testing 3,23,18 a eficácia dos agentes de controle biológico, como F. arisanus contra outras espécies Bactrocera deve ser uma alta prioridade, e é nossa esperança que a aplicação dos métodos descritos neste protocolo e artigo de vídeo que o acompanha, vai acelerar a pesquisa sobre F. arisanus em uma ampla variedade de locais 24. Por fim, novas pesquisas utilizando este método como ponto de partida também pode fornecer informações importantes para a colonização de parasitóides de ovos romance 25,1

an optimized protocol for rearing fopius arisanus, a parasitoid of tephritid fruit flies

Fopius arisanus (Sonan) é um parasitóide de moscas da fruta importante Tephritid por pelo menos duas razões. Primeiro, é o um dos únicos três parasitóides opiine conhecido para infectar o hospedeiro durante a fase de ovo 1. Em segundo lugar, tem uma ampla gama de hospedeiros em potencial de frutas voar. Talvez devido à sua história de vida, F. arisanus tem sido um sucesso usado para controle biológico de moscas da fruta em várias regiões tropicais 2-4. Um impedimento para o amplo uso do F. arisanus para controle de mosca da fruta é que é difícil estabelecer uma colônia de laboratório estável 5-9. Apesar desta dificuldade, na década de 1990 pesquisadores do USDA desenvolveu um método confiável para manter populações de laboratório de F. arisanus 10-12. Há um interesse significativo em F. arisanus 13,14 biologia, especialmente quanto à sua capacidade de colonizar uma grande variedade de hosts Tephritid 14-17; interesse é especialmente dirigido pelo alarmante disseminação de pragas mosca Bactrocera frutas para novos continentes na última década 18. Pesquisa mais aprofundada sobre F. arisanus e implementações adicionais desta espécie como um agente de controle biológico irá beneficiar de otimizações e melhorias de métodos de criação. Neste protocolo e artigo vídeo associado descrevemos um método otimizado para a criação de F. arisanus com base em uma abordagem descrita anteriormente 12. O método descrito aqui permite criação de F. arisanus em pequena escala, sem a utilização de frutas, utilizando materiais disponíveis em regiões tropicais em todo o mundo e com as exigências de trabalho relativamente baixa manual.

Watch this Scientific Journal Video about An Optimized Protocol for Rearing Fopius arisanus, a Parasitoid of Tephritid Fruit Flies at JoVE.com

An Optimized Protocol for Rearing Fopius arisanus, a Parasitoid of Tephritid Fruit Flies

Fopius arisanus é um parasitóide de ovos-larvas de moscas da fruta Tephritid que é utilizado com sucesso no controle biológico dessas pragas importantes tropical. Descrevemos aqui um protocolo otimizado para a criação de F. arisanus em pequena escala, utilizando materiais facilmente disponíveis.

Um protocolo Optimizado para Criação Fopius arisanus, Um parasitóide de Frutas Tephritid Flies

Fopius arisanus (Sonan) is an important parasitoid of Tephritid fruit flies for at least two reasons. First, it is the one of only three opiine ...

an-optimized-protocol-for-rearing-fopius-arisanus-parasitoid

Research

JoVE Journal

Biology

An Optimized Protocol for Rearing Fopius arisanus, a Parasitoid of Tephritid Fruit Flies

14.7K visualizações.  US Pacific Basin Agricultural Research Center. Fopius arisanus é um parasitóide de ovos-larvas de moscas da fruta Tephritid que é utilizado com sucesso no controle biológico dessas pragas importantes tropical. Descrevemos aqui um protocolo otimizado para a criação de F. arisanus em pequena escala, utilizando materiais facilmente disponíveis.

Vídeo: An Optimized Protocol for Rearing Fopius arisanus, a Parasitoid of Tephritid Fruit Flies

Studying Aggression in Drosophila (fruit flies)

Aggression is an innate behavior that evolved in the framework of defending or obtaining resources. This complex social behavior is influenced by genetic, hormonal and environmental factors. In many organisms, aggression is critical to survival but controlling and suppressing aggression in distinct contexts also has become increasingly important. In recent years, invertebrates have become increasingly useful as model systems for investigating the genetic and systems biological basis of complex social behavior. This is in part due to the diverse repertoire of behaviors exhibited by these organisms. In the accompanying video, we outline a method for analyzing aggression in Drosophila whose design encompasses important eco-ethological constraints. Details include steps for: making a fighting chamber; isolating and painting flies; adding flies to the fight chamber; and video taping fights. This approach is currently being used to identify candidate genes important in aggression and in elaborating the neuronal circuitry that underlies the output of aggression and other social behaviors.

Studying Aggression in Drosophila fruit flies

Agressão a estudar em Drosophila (mosca da fruta)

Aggression is an innate behavior that evolved in the framework of defending or obtaining resources.  This complex social behavior is influenced by ...

Biologia

Protocol for Mosquito Rearing (A. gambiae)

This protocol describes mosquito rearing in the insectary. The insectary rooms are maintained at 28&deg;C and ~80% humidity, with a 12 hr. day/night cycle. For this procedure, you'll need mosquito cages, 10% sterile sucrose solution, paper towels, beaker, whatman filter paper, glass feeders, human blood and serum, water bath, parafilm, distilled water, clean plastic trays, mosquito food (described below), mosquito net to cover the trays, vacuum, and a collection chamber to collect adults.

Protocol for Mosquito Rearing A. gambiae

This video illustrates the general techniques used to rear Anopheles gambiae in the laboratory. The methods for caring for laboratory mosquitoes are demonstrated through all stages of the organism's life cycle from larvae to pupae to blood-feeding adults.

Protocolo para a Criação Mosquito (A. gambiae)

This protocol describes mosquito rearing in the insectary. The insectary rooms are maintained at 28&deg;C and ~80% humidity, with a 12 hr. day/night ...

Isolation and Biophysical Study of Fruit Cuticles

The cuticle, a hydrophobic protective layer on the aerial parts of terrestrial plants, functions as a versatile defensive barrier to various biotic and abiotic stresses and also regulates water flow from the external environment.1 A biopolyester (cutin) and long-chain fatty acids (waxes) form the principal structural framework of the cuticle; the functional integrity of the cuticular layer depends on the outer 'epicuticular' layer as well as the blend consisting of the cutin biopolymer and 'intracuticular' waxes.2 Herein, we describe a comprehensive protocol to extract waxes exhaustively from commercial tomato (Solanum lycopersicum) fruit cuticles or to remove epicuticular and intracuticular waxes sequentially and selectively from the cuticle composite. The method of Jetter and Sch&auml;ffer (2001) was adapted for the stepwise extraction of epicuticular and intracuticular waxes from the fruit cuticle.3,4 To monitor the process of sequential wax removal, solid-state cross-polarization magic-angle-spinning (CPMAS) 13C NMR spectroscopy was used in parallel with atomic force microscopy (AFM), providing molecular-level structural profiles of the bulk materials complemented by information on the microscale topography and roughness of the cuticular surfaces. To evaluate the cross-linking capabilities of dewaxed cuticles from cultivated wild-type and single-gene mutant tomato fruits, MAS 13C NMR was used to compare the relative proportions of oxygenated aliphatic (CHO and CH2O) chemical moieties.
Exhaustive dewaxing by stepwise Soxhlet extraction with a panel of solvents of varying polarity provides an effective means to isolate wax moieties based on the hydrophobic characteristics of their aliphatic and aromatic constituents, while preserving the chemical structure of the cutin biopolyester. The mechanical extraction of epicuticular waxes and selective removal of intracuticular waxes, when monitored by complementary physical methodologies, provides an unprecedented means to investigate the cuticle assembly: this approach reveals the supramolecular organization and structural integration of various types of waxes, the architecture of the cutin-wax matrix, and the chemical composition of each constituent. In addition, solid-state 13C NMR reveals differences in the relative numbers of CHO and CH2O chemical moieties for wild-type and mutant red ripe tomato fruits. The NMR techniques offer exceptional tools to fingerprint the molecular structure of cuticular materials that are insoluble, amorphous, and chemically heterogeneous. As a noninvasive surface-selective imaging technique, AFM furnishes an effective and direct means to probe the structural organization of the cuticular assembly on the nm-&mu;m length scale.

Aerial plant organs are protected by the cuticle, a supramolecular biopolyester-wax assembly. We present protocols to monitor selective removal of epi- and intracuticular waxes from tomato fruit cuticles on molecular and micro scales by solid-state NMR and atomic force microscopy, respectively, and to assess the cross-linking capacity of engineered cuticular biopolyesters.

Isolamento e Estudo de Biofísica de Cutículas Frutas

The cuticle, a hydrophobic protective layer on the aerial parts of terrestrial plants, functions as a versatile defensive barrier to various biotic and ...

Fruit Volatile Analysis Using an Electronic Nose

Numerous and diverse physiological changes occur during fruit ripening, including the development of a specific volatile blend that characterizes fruit aroma. Maturity at harvest is one of the key factors influencing the flavor quality of fruits and vegetables1. The validation of robust methods that rapidly assess fruit maturity and aroma quality would allow improved management of advanced breeding programs, production practices and postharvest handling. 
Over the last three decades, much research has been conducted to develop so-called electronic noses, which are devices able to rapidly detect odors and flavors2-4. Currently there are several commercially available electronic noses able to perform volatile analysis, based on different technologies. The electronic nose used in our work (zNose, EST, Newbury Park, CA, USA), consists of ultra-fast gas chromatography coupled with a surface acoustic wave sensor (UFGC-SAW). This technology has already been tested for its ability to monitor quality of various commodities, including detection of deterioration in apple5; ripeness and rot evaluation in mango6; aroma profiling of thymus species7; C6 volatile compounds in grape berries8; characterization of vegetable oil9 and detection of adulterants in virgin coconut oil10. 
This system can perform the three major steps of aroma analysis: headspace sampling, separation of volatile compounds, and detection. In about one minute, the output, a chromatogram, is produced and, after a purging cycle, the instrument is ready for further analysis. The results obtained with the zNose can be compared to those of other gas-chromatographic systems by calculation of Kovats Indices (KI). Once the instrument has been tuned with an alkane standard solution, the retention times are automatically converted into KIs. However, slight changes in temperature and flow rate are expected to occur over time, causing retention times to drift. Also, depending on the polarity of the column stationary phase, the reproducibility of KI calculations can vary by several index units11. A series of programs and graphical interfaces were therefore developed to compare calculated KIs among samples in a semi-automated fashion. These programs reduce the time required for chromatogram analysis of large data sets and minimize the potential for misinterpretation of the data when chromatograms are not perfectly aligned.
We present a method for rapid volatile compound analysis in fruit. Sample preparation, data acquisition and handling procedures are also discussed.

A rapid method for volatile compound analysis in fruit is described. The volatile compounds present in the headspace of a homogenate of the sample are rapidly separated and detected with ultra-fast gas chromatography (GC) coupled with a surface acoustic wave (SAW) sensor. A procedure for data handling and analysis is also discussed.

Análise de Frutas volátil Usando um Nariz Eletrônico

Numerous and diverse physiological changes occur during fruit ripening, including the development of a specific volatile blend that characterizes fruit ...

Optimized Protocol for Retinal Wholemount Preparation for Imaging and Immunohistochemistry

Working with delicate tissue can be a complicating factor when performing immunohistochemical assessment. Here, we present a method that utilizes a ring-supported hydrophilized PTFE membrane to provide structural support to both living and fixed tissue during immunohistochemical processing, which allows for the use of a variety of protocols that would otherwise cause damage to the tissue. First, this is demonstrated with bolus loading of fluorescent markers into living retinal tissue. This method allows for quick visualization of targeted structures, while the membrane support maintains tissue integrity during the injection and allows for easy transfer of the preparation for further imaging or processing.
Second, a procedure for antibody staining in tissue fixed with carbodiimide is described. Though paraformaldehyde fixation is more common, carbodiimide fixation provides a superior substrate for the visualization of synaptic proteins. A limitation of carbodiimide is that the resulting fixed tissue is relatively fragile; however, this is overcome with the use of the supporting membrane. Retinal tissue is used to demonstrate these techniques, but they may be applied to any fragile tissue.

The protocol described here is for structural assessment of a wholemount retinal preparation. This includes descriptions of tissue dissection, mounting onto a hydrophilized polytetrafluoroethylene (PTFE) membrane insert, bolus loading with fluorescent markers, and a comparison of fixation with carbodiimide and paraformaldehyde for immunohistochemical analysis of cellular and synaptic components.

Working with delicate tissue can be a complicating factor when performing immunohistochemical assessment. Here, we present a method that utilizes a ...

Laboratory Rearing of Stable Flies and Other Muscoid Diptera

Stable flies, Stomoxys calcitrans, are serious pests of livestock, humans, companion animals and wildlife worldwide. During the last 20+ years, changes in agronomic practices resulted in serious outbreaks of stable flies in several countries. These outbreaks disrupted livestock production and human recreation resulting in public demands for increasing research and management efforts for this pest. A simple and inexpensive procedure for rearing stable flies for laboratory studies is presented. The procedure uses locally available diet components, equipment and supplies. The procedure can be adapted for rearing other muscoid flies including face fly (Musca autumnalis), horn fly (Haematobia irritans), and house fly (Musca domestica). The procedure produces stable fly puparia averaging 12.5 mg and ~35% egg to adult survival. Approximately 3000 flies are produced in each pan.

A procedure for rearing stable flies (Stomoxys calcitrans) is presented. The procedure uses locally available materials for diet components, equipment and supplies.

Laboratório de criação de moscas estáveis e outros Muscoid Diptera

Stable flies, Stomoxys calcitrans, are serious pests of livestock, humans, companion animals and wildlife worldwide. During the last 20+ years, changes in ...

Simple Homemade Tools to Handle Fruit Flies&#8212;Drosophila melanogaster

The fruit fly, Drosophila melanogaster, is widely used both in biological research and biology education. Handling adult flies is common but difficult in practice, as adult flies fly. Demonstrated here is how to make some simple and cost-effective tools to address difficult issues in the handling of Drosophila. Holes in foam stoppers are made and pipette tips or funnels are inserted into the holes. Flies then move only in one direction into the pipette tip/funnel assemblage, allowing efficient control of the transfer of adult Drosophila into or out of a vial. Existing protocols have been modified for cool-anesthetizing flies by chilling in crushed ice and transferring them onto a cold, hard icepack surface. The icepack is covered with a piece of medical gauze that keeps immobilized flies from the condensed water when examined under a stereomicroscope. The flies are finally euthanized for counting and sorting or discarded by microwaving. A bottle-shaped cage has also been developed for collecting eggs, as well as a labor-saving device and accompanying protocol for cleaning glass culture vials.

Simple Homemade Tools to Handle Fruit Flies&amp;#8212;&lt;em&gt;Drosophila melanogaster&lt;/em&gt;

Described here is the use of several homemade tools to transfer, chill, and kill adult Drosophila, as well as to clean glass culture vials and collect eggs. These tools are easy to make and are rather efficient in handling Drosophila.

Ferramentas caseiras simples para lidar com moscas de frutas —Drosophila melanogaster

The fruit fly, Drosophila melanogaster, is widely used both in biological research and biology education. Handling adult flies is common but difficult in ...

Topical Application Bioassay to Quantify Insecticide Toxicity for Mosquitoes and Fruit Flies

The continued use of insecticides for public health and agriculture has led to widespread insecticide resistance and hampering of control methods. Insecticide resistance surveillance of mosquito populations is typically done through Centers for Disease Control and Prevention (CDC) bottle bioassays or World Health Organization (WHO) tube tests. However, these methods can result in a high degree of variability in mortality data due to variable insecticide contact with&#160;the insect, the relatively small numbers of organisms tested, extensive variation in mass between populations, and constantly changing environmental conditions, leading to variable outcomes. This paper presents the topical application bioassay, adapted as a high-throughput phenotypic bioassay for both mosquitoes and fruit flies, to test large numbers of insects along&#160;a range of insecticide concentrations.
This assay 1) ensures consistent treatment and insecticide contact with every organism, 2) produces highly specific dose-response curves that account for differences in average mass between strains and sexes (which is particularly important for field-collected organisms), and 3) allows for the calculation of statistically rigorous median lethal doses (LD50), which are necessary for resistance ratio comparisons-an alternative surveillance approach from diagnostic dose mortality, which is also used for larvicide resistance surveillance. This assay will be a complementary tool for accurately phenotyping mosquito populations and, as illustrated using fruit flies, is easily adaptable for use with other insects. We argue that this assay will help fill the gap between genotypic and phenotypic insecticide resistance in multiple insect species.

We describe the methodology and importance of the topical application bioassay to measure insecticide susceptibility in mosquitoes and fruit flies. The presented assay is high-throughput, utilizes insect mass-thus allowing for calculating a mass-relativized lethal dose instead of concentration-and likely has lower variability than other similar methods.

Bioensaio de aplicação tópico para quantificar toxicidade de inseticida para mosquitos e moscas de frutas

The continued use of insecticides for public health and agriculture has led to widespread insecticide resistance and hampering of control methods. ...

Methods for Rearing the Parasitoid Ganaspis brasiliensis, a Promising Biological Control Agent for the Invasive Drosophila suzukii

Native to East Asia, the spotted-wing drosophila, Drosophila suzukii (Matsumura) (Diptera: Drosophilidae), has established widely in the Americas, Europe, and parts of Africa over the last decade, becoming a devastating pest of various soft-skinned fruits in its invaded regions. Biological control, especially by means of self-perpetuating and specialized parasitoids, is expected to be a viable option for sustainable area-wide management of this highly mobile and polyphagous pest. Ganaspis brasiliensis Ihering (Hymenoptera: Figitidae) is a larval parasitoid that is widely distributed in East Asia, and has been found to be one of the most effective parasitoids of D. suzukii.
Following rigorous pre-introduction evaluations of its efficacy and potential non-target risks, one of the more host-specific genetic groups of this species (G1 G. brasiliensis) has been approved recently for introduction and field release in the United States and Italy. Another genetic group (G3 G. brasiliensis), which was also commonly found to attack D. suzukii in East Asia, may be considered for introduction in the near future. There is currently enormous interest in rearing G. brasiliensis for research or in mass-production for field release against D. suzukii. This protocol and associated video article describe effective rearing methods for this parasitoid, both on a small scale for research and a large scale for mass-production and field release. These methods may benefit further long-term research and use of this Asian-native parasitoid as a promising biological control agent for this global invasive pest.

Methods for Rearing the Parasitoid Ganaspis brasiliensis, a Promising Biological Control Agent for the Invasive Drosophila suzukii

Ganaspis brasiliensis-a larval parasitoid of Drosophila suzukii (a global invasive fruit crop pest)-has been approved or is considered for introduction into Europe and the United States for biological control of this pest. This article provides protocols for both small-scale and large-scale rearing of this parasitoid.

Métodos de Criação do Parasitoide Ganaspis brasiliensis, um Agente promissor de controle biológico para a Drosophila suzukii invasiva

Native to East Asia, the spotted-wing drosophila, Drosophila suzukii (Matsumura) (Diptera: Drosophilidae), has established widely in the Americas, Europe, ...

Criação de vespas Nasonia livres de germes

Optimizing the Rearing Procedure of Germ-Free Wasps

Aseptic rearing technology is a method of culturing insects under sterile or almost sterile conditions, which can effectively eliminate the influence of external microorganisms on insect microbiota and thus promote the rapid development of insect microbiota research. Nasonia (wasp genus) is a parasitic wasp insect that has many advantages, such as a short lifespan, high genetic variation, easy operation, etc., and is widely used as an insect model system. Unlike antibiotic treatment, which can only reduce the number of microorganisms in animals, aseptic rearing techniques can control both the composition and quantity of microorganisms in animals, further facilitating the study of host-microbe interactions. However, previous versions of Nasonia rearing medium (NRM) have some defects and problems, such as a complex and time-consuming preparation process, easy contamination by bacteria or fungi, and short storage time. Therefore, this study solves these problems by optimizing the tools used in the NRM preparation process, storage conditions, and component ratios. The optimized medium could allow storage at -20 &#176;C for at least 3 months and eliminate the possibility of NRM contamination during feeding sterile wasps. This further improves the survival rate and health level of aseptic Nasonia, which is important for using Nasonia as a model for microbial research.

Autor em destaque: Otimizando o procedimento de criação de vespas livres de germes  

Nasonia wasp embryos were dissected from Lucillia sericata pupae after parasitization for 12-24 h and washed with alcohol and 10% sodium hypochlorite solution to obtain germ-free embryos. After rearing the germ-free embryos&#160;and supplying them with Nasonia rearing medium to grow and develop in vitro, germ-free Nasonia adults were obtained.

Otimizando o procedimento de criação de vespas livres de germes

Aseptic rearing technology is a method of culturing insects under sterile or almost sterile conditions, which can effectively eliminate the influence of ...