作者非常感谢来自美国林务局森林健康保护评估监测和特科技发展计划提供支持的开发，修改和应用在这里描述识别树木，抗性和敏感的山毛榉介壳虫的协议提供资金。我们也感谢霍顿植物园，柯特兰，俄亥俄州，为我们的美国榉木抗性筛选程序的继续合作和支持。

1植物材料：成熟的领域树木，盆栽苗木，盆栽或移植<ol><li>现场测试中，选择显示比例虫蛀或疾病进行检测的可能的阻力的迹象成熟健康的美国山毛榉树。明显地容易树木也将需要确定被用作对照（ 图1）。 </li><li>为了测试盆栽苗或移植物，收集和在科赫和凯里，2004或在凯里等人 ，2013描述的接穗嫁接描述发芽山毛榉。 </li><li>育苗移植或在盆栽土壤混合修订，47克微量营养素，477克缓释肥料15N-3.9P-9.9K，700克粗珍珠岩和每2.8立方米75克硫酸铝。英尺袋。如果在生长季节后需要，植物施肥每周用可溶性17N-1.3P-14.1K在200 ppm的氮。 </li><li>在生长季节，保持在阴凉处室内植物。让植物外面在秋天之前，移动它们进入休眠状态要控制温度储存设施（〜4℃）从十一月至四月。 </li></ol>榉木秤鸡蛋2个。收藏<ol><li>在一个BBD出没的展台，检查严重感染的树木（通过他们的“粉刷”的出现， 图1A易于识别）用一只手放大镜来确认鸡蛋，通常从丰富七月中旬到八月中旬的存在。 </li><li>使用画笔轻轻刷成人介壳虫，鸡蛋等杂物的白色蜡状团块，成塑料密封1加仑收集袋（ 图1B）。从至少三个不同的树木收集至少12米外。 </li><li>如果需要，在密封的收集袋长达两个星期的鸡蛋存放在4°C。带的湿的聚乙烯泡沫的袋子，以防止鸡蛋从晒出的内部的小片（2.5厘米见方）。 </li><li>从成年人（0.60毫米）和debri分开规模鸡蛋（0.15×0.25毫米）秒，用一段短的2&#39;PVC管和一个耦合到支持一块方形的250微米的尼龙筛（ 图2）构建一个筛子。 </li><li>清空成人，蜡，鸡蛋，并从收集袋碎片的混合物到筛子并用一个小漆刷轻轻地鼓励卵穿过网眼进入下面的玻璃培养皿中。应避免塑料培养皿中，因为它们容纳更多的静电，从而难以移动的鸡蛋。鸡蛋之前和筛分后示于图3。 </li><li>纯化的蛋可以存储在培养皿中，在4℃下至少一周。为了防止鸡蛋变干，大盘潮湿的一块泡沫的盖子，用封口膜密封。 </li></ol> 3，卵子活力测定<ol><li>评估卵存活率，使用10毫升注射器，以涂上薄薄的凡士林的环围绕60mm的玻璃培养皿的底部（ 图4A中的圆周</stroNG&gt;）。 </li><li>转让约100个​​鸡蛋到环的中心，将盖子上的培养皿中，并用封口膜密封。允许所述密封培养皿，以保持原状在室温下放置2周，3周的时间，如果已存储的蛋在4℃下开始测定之前。 </li><li>孵化的若虫会卡在凡士林，并且可以很容易地计算，并且空的鸡蛋很容易从未孵化的卵通过它们的颜色和光泽（ 图4B）不同。除以若虫的数量通过的空鸡蛋加余下的全蛋的总和计算存活率。良好的生存力应该在75％到90％孵出的卵范围。 </li></ol> 4，规模性的现场大型成熟的山毛榉树的筛选<ol><li>对于定量检测，计数出500个卵用解剖显微镜和使用小刮勺跨predampened，开孔10×15×1.3厘米聚乙烯泡​​沫塑料的矩形的中心轻轻地撒它们。至抑制泡沫，润湿它，然后挤压出尽可能多的水可能。对于定性试验，500鸡蛋可以算出来，放在一个小玻璃小瓶和一个“填充”行画，可用于测量约500个卵额外的批次。 </li><li>将泡沫垫到测试树鸡蛋一边对树皮面对的问题。用绳，带，细绳，或涂塑丝硬件握住垫到位。塑料或金属基材料，应使用而非天然纤维材料，这些材料由野生动物更易于清理。 </li><li>线的顶部和预切割23×30厘米的一块透蒸汽防水屋保鲜用乙硅氧烷基粘合剂的两侧，并将其放置在泡沫测试焊盘。请按殿裹边的树来创建一个防水密封。将尼龙绳或塑料涂层的硬线周围的树和房子包起来，使其保持在一定的地方，而粘台（ 图5）。 </li><li>放置至少两测试焊盘上的每个测试树，最好是在树干的两侧。在每个站点，放置测试点上至少有2棵敏感（有明显的天然鳞片虫蛀）作为对照。在此之前放置在测试片上的控制树，取出用坚定的毛刷任何自然发生介壳虫或鸡蛋。 注意：泡沫垫用鸡蛋，也可放置在树上的饲养规模鸡蛋，这可以是特别有帮助的地区为患电平为低的目的。 </li></ol> 5，规模性盆栽幼苗或移植物的筛选<ol><li>为了进行测试，选择1厘米（卡尺5厘米以上的土壤线）是足够高的至少有2个独立的测试焊盘放置在它们的最小直径盆栽树木。必要时，以腾出空间供试片修剪小偏科。 </li><li>用解剖显微镜来计算出150鸡蛋和撒在预切割和挫伤2.5×7.6×1.3厘米开孔polyethy溶解乙炔泡沫垫。 </li><li>加盖泡沫垫与苗对树皮鸡蛋的一面，用涂塑丝。缠绕在幼苗只是泡沫上面有个小平米的房子保鲜膜和醋酸基有机硅密封（ 见图6A）。 </li><li>包括已知的易感苗家庭或移植作为对照。 </li></ol> 6，数据采集注意：将所述泡沫垫和鸡蛋在测试树后约52至57周后，数据可以被收集。重要的是，不这样做，大人也开始产卵，使他们的繁殖能力才能确定之后。在一些树木耐它是很平常看到少数的成年人建立，但没有再生产发生。 <ol><li>小心地取下泡沫垫和计数的使用手镜或放大镜（10倍），在树皮上建立成人介壳虫的数量。 </li><li>当泡沫垫被去除，它是非uncommon一些成人和最蛋同一簇的泡沫要被拉断树（参见图7）。使用解剖显微镜，算上卵块，而仍然附着在泡沫垫成年人。 </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. 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作者宣称，他们有没有竞争的财务权益。

需要这种测定法的成功关键步骤包括对所述蛋的可行性测试和在每个站点测试使用容易控制的，并与盆栽材料。同样重要的是使用每树或幼苗多个测试垫。我们发现，无论是在现场，并于盆栽多控制测试中，错误的一个常见来源可以是键盘故障。例如，在现场，即使我们的覆盖测试垫与房子的包裹，以防止多余的水分，水向下流动阀杆可以偏向于树的一侧，并且产生一个测试垫的损失的修改。 </p...

山毛榉树皮病（BBD）自19世纪90年代后期3引入了入侵的山毛榉介壳虫， 隐球菌fagisuga，在新斯科舍省的加拿大省已在北美地区于美国山毛榉产生了不利影响。当山毛榉介壳虫其插入喂食管芯到树皮制作的小裂缝，提供入口感染的Neonectria种真菌（Neonectria ditissima或Neonectria faginata）中的一本昆虫病情复杂启动。由于真菌菌丝体生长，大面积的组织可能会死，最终完全环剥树。从疾病的伤害削弱了树，使它容易出现抢购的大风4。在疾病的第一波死亡率水平已报道高达50％5。幸存的树木往往严重变形，溃疡形成降低了树的值作为木材产品。0;这种树有一个倾向根萌这导致了“山毛榉树丛”，以防止其他更理想的物种建立，减少了支架6的经济和生态价值的形成。虽然山毛榉树皮病是不是很可能导致美国山毛榉灭绝，它改变立场的组成和健康导致食物和栖息地减少对野生动物的7,8。 在受BBD多年看台上，有报道称，依然无任何疾病症状的树木。人工接种试验表明，这些树是耐介壳虫2。如果没有规模虫蛀，有一个Neonectria感染的机会很少，尽量减少真菌的影响。大规模死亡的美国榉木，由于Neonectria感染在没有事先规模为患从未据报道，所以抗阻为BBD山毛榉介壳虫结果。 对BBD的管理最近的研究集中在识别，繁殖，饲养，并保留美国山毛榉树与阻力山毛榉介壳虫的。遗传研究表明，抗介壳虫是可以遗传的，耐树木的精心选育可能会导致在一个世代9显著改善。这一发现得益于在美国的州和国家森林经理努力建立抵抗美国榉木的区域种子园提供遗传多样性BBD性种子的来源进行恢复种植10,11。研究还表明立场遗传学造林操纵去除易感树木和保留性的树木可能会导致林分改造9,12。 &lt;/ P&gt; BBD通过树木改良活动，或通过开展造林处方管理需要来选择和山毛榉规模性和易感树木区分的能力。这里介绍的方法已经改编自一个方法首先由戴维·休斯顿引入人工接种苗木榉规模鸡蛋1。该方法可以作为一种筛选工具，以确定数量性状基因座（QTL）与电阻或在遗传研究抗性和易感盆栽幼苗或接枝分株区分相关联。另外，它可用于筛选成熟的树木在现场以确定种子园的发展，或保留在现场耐树木。易感树木可以识别并删除，以减少疾病的影响。

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

山毛榉树皮病（BBD）的结果在高水平的初始死亡率，留下幸存者的树木被大大削弱和变形。本病是由喂养侵入山毛榉介壳虫， 隐球菌fagisuga，它由Neonectria种真菌中的一种创造切入点感染活动启动。如果没有规模为患，对于真菌感染的机会很少。使用规模鸡蛋人为侵扰的严重影响BBD看台的健康树木证明，这些树是耐病情复杂1的介壳虫部分。在这里，我们提出，我们已经开发的基础上，由休斯敦2人为侵扰的技术，它可以用于筛查的领域和更小的盆栽苗木移植和规模性树木的协议。规模性树木识别是对BBD的管理，通过树的重要组成部分改进方案和造林操纵。

图6显示了抗苗（C）和两个易感苗（D，E）表现出不同程度的敏感性。一个什么样的易感成熟的树看起来像59周人工接虫试验成立后的一个例子如图5B所示。当泡沫被剥离的树，它是不寻常的介壳虫及其卵簇保持粘在泡沫如示于图7，这就是为什么它是必不可少的，以收集从树和泡沫衬垫的数据。 图8?...

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.

山毛榉树皮病是由喂养，在树皮真菌创建入口点的山毛榉介壳虫活动启动。树木能够抵抗介壳虫也是疾病的抵抗力。在这里，我们提出，我们已经发展到筛选山毛榉规模抵抗个别山毛榉树的协议。

一种技术，屏幕美国榉木为抵抗山毛榉介壳虫（<em&gt;隐球菌fagisuga</em&gt;林德。）

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

a-technique-to-screen-american-beech-for-resistance-to-beech-scale

Research

JoVE Journal

Environment

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

9.5K 次观看.  US Forest Service. 山毛榉树皮病是由喂养，在树皮真菌创建入口点的山毛榉介壳虫活动启动。树木能够抵抗介壳虫也是疾病的抵抗力。在这里，我们提出，我们已经发展到筛选山毛榉规模抵抗个别山毛榉树的协议。 

视频: 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.

一种非侵入性的方法<em&gt;原位</em&gt;交配成功的女美国龙虾​​（测定<em&gt;美洲螯龙虾</em&gt;）

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

科研

环境科学

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.

用于蜂蜜蜜蜂筛选化学成分的视频跟踪协议

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.

用实验室尺度模型评价深层包粪排放的恶臭和气体浓度

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.

双链 rna 口服法诱导韧皮部和植物-sap 喂养 Hemipteran 昆虫 rna 干扰

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.

藤壶Balanus improvisus作为海洋模型的培养和基因表达

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.

检测幼虫行为反应剖面的实验协议: 在神经刺激咖啡因中的应用

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.

一种精确、自主的昆虫产生模式检测系统

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.

不同环境条件下生物气溶胶的组成及分布分析

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.

分离叶测定,以简化马铃薯中基因表达研究,在咀嚼昆虫曼杜卡性塔感染

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.

一种新的微声学标记在少年太平洋兰普瑞和美洲鳗鱼中的植入

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