我们感谢博士。安东尼山茱萸和劳拉·诺里斯在加州大学戴维斯分校和卡塔琳娜Kreppel博士在格拉斯哥大学从坦桑尼亚提供蚊子标本。我们感谢Smita达斯女士和道格拉斯·诺里斯博士公共卫生约翰霍普金斯学院分享蚊虫样本赞比亚。我们也感谢李五米永先生在兽医遗传学实验室对实验设计培训。这项工作是支持由美国国立卫生研究院授予R01AI 078183和R21AI062929。

1. PCR扩增<ol><li>混合所有的PCR引物（见补充表S1）的1.5ml离心管。每个SNP具有两个引物（正向和反向）。确保原料浓度每种PCR引物被保持在100微米。使足够引物混合物为500-1000的反应，以减少在工作台上移液误差和时间（参见补充表S2）。如果需要的话，制备等分试样，每管并储存100-200反应在-20℃。 </li><li>制备的PCR鸡尾酒如生产商的方案（表1）中所描述。 96反应的体积包括20％的突出端。涡包含PCR鸡尾酒轻轻离心30秒，2000×g的管子。 </li><li>分配3微升的PCR鸡尾酒成的每个孔中的非裙边96孔板中。中继器吸管可以节省时间，这个过程中。 </li><li>添加2微升的蚊子的适当的基因组DNA中的每个孔中。 注:获得从米的基因组DNA的方法<html> osquito组织（头部/胸部，腹部或全身），在其他文献29-32描述。选择一个合适的DNA提取方法来区分那些与疟原虫在血液（腹部）从受感染蚊子（ 疟原虫的头部/胸部）。我们通常使用DNA从蚊子的身体部位（头部/胸部或腹部）中提取，因此从0.05-2ng /μl的DNA浓度变化。虽然总的DNA输入比制造商的建议（5-10ng / RXN）下，我们通常得自98％的DNA样本的没有全基因组扩增的SNP健壮呼叫。 </li><li>封板，轻轻拌匀或涡流，并在370×g的1分钟离心机。 </li><li>使用以下热循环协议来进行PCR扩增:（1）94℃2分钟，（2）94℃，30秒，（3）56℃，30秒，（4）72℃，1分钟，（5）重复步骤（2） - （4）45个循环，（6）72℃，1分钟，4℃保持。 </li></ol>_title"&gt; 2。SAP治疗<ol><li>准备在96孔板在SAP酶溶液（见表2）。处理在一个板96个样品。 96反应的体积包括20％的突出端。 </li><li>涡SAP酶液轻轻离心管30秒，2000×g的。分配2微升的SAP酶溶液加入到每个孔中。再次，一个转发器移液管，可以节省时间用于此过程。 </li><li>封板，轻轻拌匀或涡流，并在370×g的1分钟离心机。孵育板如下:（1）37℃下40分钟，（2）85℃下进行5分钟，以使酶失活，（3）4℃保持。在该步骤完成后，在-20存储板℃下再继续。 2个星期内处理存储的板块。 </li></ol> 3. SNP扩展<ol><li>如果样品板的SAP处理后冷冻，允许它继续之前解冻至室温。 </li><li>制备所述延伸引物混合物（补充表S2）。每个SNP有一个EXTEnsion底漆。保持股票集中每个延伸引物在500微米。任选地，对等分试样每管并储存100-200反应在-20℃。 </li><li>如表3中96的反应体积，包括20％的悬垂描述制备延伸组合。 </li><li>涡延伸鸡尾酒轻轻离心管30秒，2000×g的。分配2微升的延伸鸡尾酒到每个孔中。再次，一个转发器移液管，可以节省时间用于此过程。 </li><li>封板，轻轻拌匀或涡流，并在370×g的1分钟离心机。热循环， 如图1中的板。在这一点上，进行质谱法或板储存在-20℃达2周。 </li></ol> 4.空调的SNP扩展产品优化质谱分析<ol><li>如果样品板SNP后延冻结，允许它继续之前解冻到室温。 </li><li>从CONTA转移树脂为15mg核能研究所使用上的长勺酒窝板。使用刮刀蔓延树脂进入凹坑板的孔中。从一边扫刮板跨越酒窝板到另一边蔓延树脂。确保有树脂每口井。 </li><li>刮去多余的树脂脱使用刮板酒窝板。让树脂站在凹坑板为至少20分钟。同时让树脂站在凹坑板中，添加41微升的HPLC级水到样品板的各孔中。 </li><li>轻轻将样品板倒置，到酒窝板。确保样品板复位靠在凹坑板的小圆形交，该对齐的孔中的样品板与孔中的凹坑板。 </li><li>保持样品板和凹坑板一起，轻轻翻转它们在这样的树脂脱落凹坑板的进入样品板的孔中。点击酒窝板，使树脂脱落到样品板。确保所有的树脂在迪mple板脱落到样品板的孔中。离心板在3200 XG为30秒。 </li><li>旋转旋转器上的样品板5分钟，在室温。旋转器必须旋转样品板约长轴360°。 </li><li>离心样品板在3200×g离心5分钟。此步骤之后，进行到下一步骤之前，长达2周存储的样品板在-20℃下。 </li></ol> 5. MALDI-TOF质谱<ol><li>转移条件的SNP延伸产物到生物芯片诸如SpectroCHIP按照制造商的说明。我们使用MassArray纳米分配器用于此步骤。 </li><li>一旦转移完成，确定测定法和板如何被设置在测定数据库中。命名法和板使得每个板块都有唯一的标识符，链接到特定的试验设计（参考表三）。 </li><li>法和板已被定义后，获取利用质谱仪光谱。 </li></ol><ol><li>使用实时PCR分析数据解释软件，如打字员分析器或TyperViewer（Sequenom公司）进行数据分析。在包含样本的布局，质谱图每个井.xml格式获得的数据，该标记信息包括名称，用于对应的SNP的每种变体预期质量，并且任何错误或与每个反应相关的警告消息。参见图2所产生的质谱图。 </li><li>由该检测阈值设定（幅度或信噪比），并允许每个基因型公差/方差进行的SNP基因型的聚类分析。 图3示出了04707-KDR＃2的SNP基因型簇的例子。 </li><li>导出所得的SNP基因型的呼叫到电子表格程序。 </li></ol>

The authors have nothing to disclose.

该MDA是由五个主要步骤:PCR扩增，虾碱性磷酸酶（SAP）的反应，SNP延伸，延伸产品的调理和基质辅助激光解吸/电离-飞行（MALDI-TOF）质谱33-37的时间。第一次PCR扩增步骤扩增的DNA的侧翼每个SNP，使得足够的模板DNA将是可利用的SNP的延伸步骤。在SAP反应中和未使用的dNTP其可以用下面的SNP延伸步骤干涉。 SNP扩展涉及每个​​SNP位点和大众修饰的核苷酸终止一个延伸引物。 3&#39;-末端修饰的核苷?...

阿拉伯按蚊，按蚊coluzzii和冈比亚按蚊负责非洲1疟疾传播的主要媒介。这三种物种在形态区分2，并且只能通过分子测定法3-9进行区分。此外，还有许多下游实验经常进行，以帮助流行病学和群体遗传学的研究。这些包括（1）为形态岛屿10-12基因分型测定法中，（2）一种基因分型测定法识别在1014的非同义的SNP 个 段基因的氨基酸的密码子的位置（ 击倒抗性或KDR，SNP）13 -18，（3）的寄生虫检测的PCR 19-23，和（4）筛选在蚊子中肠24,25宿主特异的DNA。 我们开发了将所有这些测定与一个目标的单个多重反应的多检测分析（MDA）的alyzing在非洲疟疾向量的流行病学重要特征。 MDA的测定法包括:多个标记，用于检测（1）种（A.阿拉伯按蚊，冈比亚按蚊，A coluzzii，或其它（所有的3）的），（2）杀虫剂中KDR的SNP表示电阻（两者L1014F 和L1014S） ，（3）存在的两个主要的疟原虫，恶性疟原虫和P.间日疟原虫 ，和（4）从一禽和六个哺乳动物宿主血液来源。 传统上，这些测定法在分开的聚合酶链式反应进行的。当使用常规PCR平台完成所有这些测定中，它需要进行8-10 PCR反应检测和伴随凝胶电泳步骤。从准备每个PCR反应，记录结果需要4-5小时，而MDA方法，这里介绍需要大约5小时的总和。这相当于节省了单独的劳动力成本的90％。在MDA这里介绍的价格为5美元每个样品基因型的所有33个SNP。这是相当便宜比单琼脂糖凝胶为基础的分析，其中的成本约为1.50美元样本。测定法检测所涵盖的所有的MDA的特性将需要至少8-10分开琼脂糖凝胶为基础的测定以$ 12-15每一个样品的成本。此外，MDA的大大减少产生假阳性或阴性通过利用至少三个标记为每个寄生虫或主机的源检测的机会。 我们所使用的平台不限于疟疾的载 ​​体，但可以在各种各样的应用，如医药，兽药和基础生物学26-28中使用。深入涉及的样品大（为了100S）的数量关联研究或群体遗传学研究需要经济高效的检测筛选同时为多个标记。利用两个或更多个单独的PCR测定大多数研究可以实现MDA的用于以较低的成本更快的结果。 </p&gt;

<table border="0" cellpadding="0" cellspacing="0" width="588">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Name of Material/ Equipment</td>
			<td style="width:87px;">Company</td>
			<td style="width:119px;">Catalog Number</td>
			<td style="width:167px;">Comments/Description</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">MassARRAY Analyzer Compact</td>
			<td style="width:87px;">Sequenom</td>
			<td style="width:119px;">MT9</td>
			<td>MALDI-TOF mass spectrometry for genomic applications to analyze nucleic acids.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">MassARRAY Nanodispenser</td>
			<td style="width:87px;">Sequenom</td>
			<td style="width:119px;">RS1000</td>
			<td>Transfers completed iPLEX reaction products to the SpectroCHIP</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">iPLEX Gold Genotyping Reagent Set</td>
			<td style="width:87px;">Sequenom</td>
			<td style="width:119px;">10158</td>
			<td>Reagents used for iPLEX assay including SAP kit.</td>
		</tr>
	</tbody>
</table>

a multi-detection assay for malaria transmitting mosquitoes

在冈比亚按蚊种类复杂，包括主要的疟疾在非洲发射蚊子。由于这些品种都是这样的医疗重要性，一些特质的典型特点采用分子检测的流行病学研究提供帮助。这些特征包括物种鉴定，抗药性，寄生虫感染状态，和主机偏好。由于冈比亚按蚊复杂的群体在形态上没有区别，聚合酶链式反应（PCR）是传统上用于识别物种。一旦种是已知的，一些下游的测定是进行例行阐明进一步的特点。例如，在一对位基因被称为KDR突变赋予对DDT和拟除虫菊酯类杀虫剂抗性。此外，酶联免疫测定（ELISA）或疟原虫寄生虫的DNA检测的PCR测定法是用于检测存在于蚊子组织寄生虫。最后，一​​个combinatioÑPCR和限制酶消化的可用于阐明宿主偏好（ 例如 ，人类与动物的血液）通过筛选蚊子吸血寄宿特异性DNA。我们已经开发了在一个时刻将所有的上述分析成单个多重反应的基因分型33SNPs为96或384个样品的多检测分析（MDA）。因为MDA包括多个标记的物种， 疟原虫的检测，和主机血液鉴定，产生假阳性或阴性的可能性大大从以前的测定法，其中包括每性状只有一个标记降低。这个强大而简单的分析可以经济高效地在现有的实验的一小部分时间检测到这些关键的蚊子特点。

物种鉴定: 以下5个SNP一起识别3种（A.阿拉伯按蚊，A coluzzii和冈比亚按蚊 ）（ 见表4）。如果样品是不是3种之一，三个单核苷酸多态性（01073-213，04679-157和10313 -052）无法放大。 KDR基因型推断抗药性: 对位电压-门控钠通道的1014 个密码子对应于KDR。两个单核苷酸多态...

Watch this Scientific Journal Video about A Multi-detection Assay for Malaria Transmitting Mosquitoes at JoVE.com

传播疟疾的蚊子的多重检测分析|生物学 |视频

A Multi-detection Assay for Malaria Transmitting Mosquitoes

Malaria transmitting mosquitoes have a number of epidemiologically important characteristics that can only be detected using molecular techniques. Utilizing a MALDI-TOF based SNP genotyping platform, we developed an assay for simultaneously detecting multiple key traits (species, insecticide resistance, parasite infection and host choice) of malaria vectors.

一种多检测试验为疟疾蚊子发射

The Anopheles gambiae species complex includes the major malaria transmitting mosquitoes in Africa. Because these species are of such medical importance, ...

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SCIENTISTS IN ACTION

13.0K 次观看.  School of Veterinary Medicine, University of California - Davis. Malaria transmitting mosquitoes have a number of epidemiologically important characteristics that can only be detected using molecular techniques. Utilizing a MALDI-TOF based SNP genotyping platform, we developed an assay for simultaneously detecting multiple key traits (species, insecticide resistance, parasite infection and host choice) of malaria vectors.

视频: A Multi-detection Assay for Malaria Transmitting Mosquitoes

Injection of An. stephensi Embryos to Generate Malaria-resistant Mosquitoes

The introduction of exogenous genes into the genomes of mosquitoes requires microinjection techniques tailored to the specific species of interest. This video protocol demonstrates a method used by the James laboratory to microinject DNA constructs into Anopheles stephensi embryos for the generation of transformed mosquitoes. Techniques for preparing microinjection needles, collecting and preparing embryos and performing the microinjection are illustrated.

Anopheles stephensi mosquitoes are vectors for malaria inhabiting India and throughout Asia. This video demonstrates the technique for performing microinjections of this species with transgenes that will confer resistance to the malaria to the mosquito. Much of the methodology demonstrated in this video is applicable to microinjection techniques of other mosquito species.

注射的。 stephensi胚胎生成抗药性疟疾的蚊子

The introduction of exogenous genes into the genomes of mosquitoes requires microinjection techniques tailored to the specific species of interest. This ...

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Protocol for Dengue Infections in Mosquitoes (A. aegypti) and Infection Phenotype Determination

The purpose of this procedure is to infect the Aedes mosquito with dengue virus in a laboratory condition and examine the infection level and dynamic of the virus in the mosquito tissues. This protocol is routinely used for studying mosquito-virus interactions, especially for identification of novel host factors that are able to determine vector competence. The entire experiment must be conducted in a BSL2 laboratory. Similar to Plasmodium falciparum infections, proper attire including gloves and lab coat must be worn at all times. After the experiment, all the materials that came in contact with the virus need to be treated with 75% ethanol and bleached before proceeding with normal washing. All other materials need to be autoclaved before discarding them.

Protocol for Dengue Infections in Mosquitoes A. aegypti and Infection Phenotype Determination

Once a gene is identified as potentially refractory for the dengue virus, it must be evaluated for it's role in preventing viral infections within the mosquito. This protocol illustrates how the extent of dengue infections of mosquitoes can be assayed. The techniques for growing up the virus in culture, membrane feeding mosquitoes human blood, and assaying viral titers in the mosquito midgut are demonstrated.

在蚊子（埃及斑蚊）感染引起的表型测定为登革热感染病例的议定书“

The purpose of this procedure is to infect the Aedes mosquito with dengue virus in a laboratory condition and examine the infection level and dynamic of ...

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.

蚊子养育的议定书“（冈比亚按蚊）

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 ...

Protocol for Plasmodium falciparum Infections in Mosquitoes and Infection Phenotype Determination

Once a gene is identified as potentially refractory for malaria, it must be evaluated for its role in preventing Plasmodium infections within the mosquito.    This protocol illustrates how the extent of plasmodium infections of mosquitoes can be assayed.  The techniques for preparing the gametocyte culture, membrane feeding mosquitoes human blood, and assaying viral titers in the mosquito midgut are demonstrated.

Once a gene is identified as potentially refractory for malaria, it must be evaluated for its role in preventing Plasmodium infections within the mosquito. This protocol illustrates how the extent of plasmodium infections of mosquitoes can be assayed. The techniques for preparing the gametocyte culture, membrane feeding mosquitoes human blood, and assaying viral titers in the mosquito midgut are demonstrated.

恶性疟原虫在蚊子和病毒感染的表型测定感染的议定书

Once a gene is identified as potentially refractory for malaria, it must be evaluated for its role in preventing Plasmodium infections within the ...

Population Replacement Strategies for Controlling Vector Populations and the Use of Wolbachia pipientis for Genetic Drive

In this video, Jason Rasgon discusses population replacement strategies to control vector-borne diseases such as malaria and dengue. "Population replacement" is the replacement of wild vector populations (that are competent to transmit pathogens) with those that are not competent to transmit pathogens. There are several theoretical strategies to accomplish this. One is to exploit the maternally-inherited symbiotic bacteria Wolbachia pipientis. Wolbachia is a widespread reproductive parasite that spreads in a selfish manner at the extent of its host's fitness. Jason Rasgon discusses, in detail, the basic biology of this bacterial symbiont and various ways to use it for control of vector-borne diseases.

In this interview, Jason Rasgon explains the concept of genetic drive and the characteristics of an effective gene drive system. The use of the endosymbiotic bacterium, Wolbachia pipientis, as a means to spread genes through mosquito populations, is hypothesized.

人口控制媒介种群和遗传的驱动器使用的Wolbachia pipientis替换策略

In this video, Jason Rasgon discusses population replacement strategies to control vector-borne diseases such as malaria and dengue. "Population ...

Protocol for RNAi Assays in Adult Mosquitoes (A. gambiae)

Reverse genetic approaches have proven extremely useful for determining  which genes underly resistance to vector pathogens in mosquitoes.  This video protocol illustrates a method used by the Dimopoulos lab to inject dsRNA into Anopheles gambiae mosquitoes, which harbor the malaria parasite.  The technique manipulating the injection setup and injecting dsRNA into the thorax is illustrated.

Protocol for RNAi Assays in Adult Mosquitoes A. gambiae

Reverse genetic approaches have proven extremely useful for determining which genes underly resistance to vector pathogens in mosquitoes. This video protocol illustrates a method used by the Dimopoulos lab to inject dsRNA into Anopheles gambiae mosquitoes, which harbor the malaria parasite. The technique manipulating the injection setup and injecting dsRNA into the thorax is illustrated.

成蚊（冈比亚按蚊的RNAi实验议定书）

Reverse genetic approaches have proven extremely useful for determining  which genes underly resistance to vector pathogens in mosquitoes.  This video ...

Preventing the Spread of Malaria and Dengue Fever Using Genetically Modified Mosquitoes

In this candid interview, Anthony A. James explains how mosquito genetics can be exploited to control malaria and dengue transmission.   Population replacement strategy, the idea that transgenic mosquitoes can be released into the wild to control disease transmission, is introduced, as well as the concept of genetic drive and the design criterion for an effective genetic drive system. The ethical considerations of releasing genetically-modified organisms into the wild are also discussed.

In this candid interview, Anthony A. James explains how mosquito genetics can be exploited to control malaria and dengue transmission. Population replacement strategy, the idea that transgenic mosquitoes can be released into the wild to control disease transmission, is introduced as well as the concept of genetic drive and the design criterion for an effective genetic drive system. The ethical considerations of releasing genetically-modified organisms into the wild are also discussed.

防止疟疾的传播和使用基因改造的蚊子登革热

In this candid interview, Anthony A. James explains how mosquito genetics can be exploited to control malaria and dengue transmission.   Population ...

Building a Better Mosquito: Identifying the Genes Enabling Malaria and Dengue Fever Resistance in A. gambiae and A. aegypti Mosquitoes

In this interview, George Dimopoulos focuses on the physiological mechanisms used by mosquitoes to combat Plasmodium falciparum and dengue virus infections.   Explanation is given for how key refractory genes, those genes conferring resistance to vector pathogens, are identified in the mosquito and how this knowledge can be used to generate transgenic mosquitoes that are unable to carry the malaria parasite or dengue virus.

In this interview, George Dimopoulos focuses on the physiological mechanisms used by mosquitoes to combat Plasmodium falciparum and dengue virus infections. Explanation is given for how key refractory genes, those genes conferring resistance to vector pathogens, are identified in the mosquito and how this knowledge can be used to generate transgenic mosquitoes that are unable to carry the malaria parasite or dengue virus.

建设一个更美好的蚊子：确定基因在冈比亚按蚊和埃及斑蚊蚊子启用疟疾和登革热的电阻

In this interview, George Dimopoulos focuses on the physiological mechanisms used by mosquitoes to combat Plasmodium falciparum and dengue virus ...

Transmitting Plant Viruses Using Whiteflies

Whiteflies, Hemiptera: Aleyrodidae, Bemisia tabaci, a complex of morphologically indistinquishable species5, are vectors of many plant viruses. Several genera of these whitefly-transmitted plant viruses (Begomovirus, Carlavirus, Crinivirus, Ipomovirus, Torradovirus) include several hundred species of emerging and economically significant pathogens of important food and fiber crops (reviewed by9,10,16). These viruses do not replicate in their vector but nevertheless are moved readily from plant to plant by the adult whitefly by various means (reviewed by2,6,7,9,10,11,17). For most of these viruses whitefly feeding is required for acquisition and inoculation, while for others only probing is required. Many of these viruses are unable or cannot be easily transmitted by other means. Therefore maintenance of virus cultures, biological and molecular characterization (identification of host range and symptoms)3,13, ecology2,12, require that the viruses be transmitted to experimental hosts using the whitefly vector. In addition the development of new approaches to management, such as evaluation of new chemicals14 or compounds15, new cultural approaches1,4,19, or the selection and development of resistant cultivars7,8,18, requires the use of whiteflies for virus transmission. The use of whitefly transmission of plant viruses for the selection and development of resistant cultivars in breeding programs is particularly challenging7. Effective selection and screening for resistance employs large numbers of plants and there is a need for 100% of the plants to be inoculated in order to find the few genotypes which possess resistance genes. These studies use very large numbers of viruliferous whiteflies, often several times per year.
Whitefly maintenance described here can generate hundreds or thousands of adult whiteflies on plants each week, year round, without the contamination of other plant viruses. Plants free of both whiteflies and virus must be produced to introduce into the whitefly colony each week. Whitefly cultures must be kept free of whitefly pathogens, parasites, and parasitoids that can reduce whitefly populations and/or reduce the transmission efficiency of the virus. Colonies produced in the manner described can be quickly scaled to increase or decrease population numbers as needed, and can be adjusted to accommodate the feeding preferences of the whitefly based on the plant host of the virus.
There are two basic types of whitefly colonies that can be maintained: a nonviruliferous and a viruliferous whitefly colony. The nonviruliferous colony is composed of whiteflies reared on virus-free plants and allows the weekly availability of whiteflies which can be used to transmit viruses from different cultures. The viruliferous whitefly colony, composed of whiteflies reared on virus-infected plants, allows weekly availability of whiteflies which have acquired the virus thus omitting one step in the virus transmission process.

Improving our understanding of and our ability to manage many of the insect-transmitted plant viruses requires the use of the vector. Insect transmission of plant viruses is a tritrophic interaction, and as such requires the manipulation of insects, virus, and plant. Vectors must be reared in large numbers and manipulated in such a way as to insure high rates of transmission to test plants. The basics of rearing and manipulating the whitefly, Bemisa tabaci (Hemiptera: Aleyrodidae), a vector of plant viruses belonging to several emerging plant virus genera, representing a large number of economically significant viruses is presented.

发射使用粉虱的植物病毒

Whiteflies, Hemiptera: Aleyrodidae, Bemisia tabaci, a complex of morphologically indistinquishable species5, are vectors of many plant viruses. Several ...

Toxicological Assays for Testing Effects of an Epigenetic Drug on Development, Fecundity and Survivorship of Malaria Mosquitoes

Insecticidal resistance poses a major problem for malaria control programs. Mosquitoes adapt to a wide range of changes in the environment quickly, making malaria control an omnipresent problem in tropical countries. The emergence of insecticide resistant populations warrants the exploration of novel drug target pathways and compounds for vector mosquito control. Epigenetic drugs are well established in cancer research, however not much is known about their effects on insects. This study provides a simple protocol for examining the toxicological effects of 3-Deazaneplanocin A (DZNep), an experimental epigenetic drug for cancer therapy, on the malaria vector, Anopheles gambiae. A concentration-dependent increase in mortality and decrease in size was observed in immature mosquitoes exposed to DZNep, whereas the compound reduced the fecundity of adult mosquitoes relative to control treatments. In addition, there was a drug-dependent decrease in S-adenosylhomocysteine (SAH) hydrolase activity in mosquitoes following exposure to DZNep relative to control treatments. These protocols provide the researcher with a simple, step-by-step procedure to assess multiple toxicological endpoints for an experimental drug and, in turn, demonstrate a unique multi-prong approach for exploring the toxicological effects of water-soluble epigenetic drugs or compounds of interest against vector mosquitoes and other insects.

A protocol is developed to examine the effects of an epigenetic drug DZNep on the development, fecundity and survivorship of mosquitoes. Here we describe procedures for the aqueous exposure of DZNep to immature mosquitoes and a blood-based exposure of DZNep to adult mosquitoes in addition to measuring SAH hydrolase inhibition.

毒理学化验一个后生药物对疟蚊的发育，繁殖和存活率测试效果

Insecticidal resistance poses a major problem for malaria control programs. Mosquitoes adapt to a wide range of changes in the environment quickly, making ...