הכנה מראש של microinjection: <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> להאכיל יתושים דם: להזרקה מיום שני עד יום רביעי נקבות להאכיל ביום שישי הקודם. כדי להזריק חמישי ושישי, נקבות להאכיל ביום שני של אותו שבוע. </li><li style=";text-align:right;direction:rtl"> הכינו את המחטים קוורץ באמצעות תוכנית 2 ו חיץ איזוטוני (ראה חומרים). </li><li style=";text-align:right;direction:rtl"> &quot;הנחת צינור&quot; עבור עוברי הוא בקבוקון תרבות רגיל תסיסנית. צמר גפן רטוב בתחתית, עם דיסק רטוב נייר לסנן מכסה אותו. </li><li style=";text-align:right;direction:rtl"> הכן להחליק כיסוי הפלסטיק על ידי דבק פעמיים בצד קלטת בקצה אחד. קלטת Trim כדי לכסות להחליק כך מסתיים בקצה של פליטת לכסות. </li><li style=";text-align:right;direction:rtl"> הכנת שמן התייבשות. </li><li style=";text-align:right;direction:rtl"> הכינו צלחת פטרי עם חיץ איזוטוני להעביר עוברים הבקיעה. </li></ol> הגדרת הנחת בכפייה: <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> איסוף דם 6-10 נקבות נמאס עם השימוש aspirator ולהעבירם בקבוקון התרבות תסיסנית עם צמר גפן רטוב עם נייר סינון חיץ איזוטוני. </li><li style=";text-align:right;direction:rtl"> יתושים הם ואז לשים בחזרה התנאים insectery בחושך מותר להטיל ביצים עבור 1 שעות ו 15 דקות. </li><li style=";text-align:right;direction:rtl"> בואו מבוגרים לטוס לתוך הכלוב ולהוציא את הדיסק עם נייר סינון עוברים. </li><li style=";text-align:right;direction:rtl"> בשורה ביצים, לעשות את זה מתחת למשטח הביתור. איסוף צרורות של ביצים עם מכחול דק (סייבל, לא 0000) ולהעביר לכיכר מוכן נייר Whatman 3mm ספוגה חיץ איזוטוני. שמור על נייר כל הזמן רטוב. אל תתנו ביצים לקבל מיובש. הסר את exochorion עם מברשת, זה עוזר יותר מקל ביצים לקלטת. </li><li style=";text-align:right;direction:rtl"> שימוש בסדר המלקחיים מס &#39;5 או במכחול דק, להרים עוברים אפור כהה יותר לארגן בשורה בכיכר של Whatman 3mm רטוב עם חיץ איזוטוני. הקו עד 20-30 עוברים. עוברים כל חייב להיות בכיוון זהה הזרקה יש בקוטב האחורי. סוף הקדמי של עוברים הוא קצת יותר מאשר האחורי. לאחר מכן, באמצעות פסים של נייר Whatman 3mm, לייבש את המסנן שבו ביצים מסודרים על ידי לחיצה קשה משני צידי הקו ביצה, לא נוגע את הביצים. </li><li style=";text-align:right;direction:rtl"> כדי להעביר את הביצים, להפוך את השקופית המכילה את סרט דו צדדי (רפואה), ולחץ בעדינות על הביצים. סוף האחורי של הביצים חייב להיות קרוב מאוד לקצה של הקלטת דו צדדי. הרטיבות של העיתון הוא קריטי זה, אם הוא רטוב מדי הם לא מקל. אני עושה את התייבשות של העוברים בין שניה 5-10, אבל הפעם התייבשות תלוי מן ולח והטמפרטורה בחדר microinjection. </li><li style=";text-align:right;direction:rtl"> התייבשות היא קריטית עבור ביצים: קצת יותר מדי עוברי לא יבקעו. ללא ה-DNA יובש לא הולך לעובר. </li><li style=";text-align:right;direction:rtl"> כיסוי עוברים מיובש עם שמן halocarbon כדי למנוע התייבשות נוספת. </li></ol> Microinjection של העוברים: ההיבט החשוב ביותר של זריקה טובה הוא איכות המחט (ראה הערה). <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> מלא מחט עם פתרון ה-DNA להיות מוזרק באמצעות microloader (Eppendorf # 5242 956.003). מאוד soliution הזרקה קטן יש צורך, 1-2 מ&quot;ל. </li><li style=";text-align:right;direction:rtl"> חבר את המחט אל Transjector Eppendorf, השולטת זמן הזרקה בלחץ, כמו גם את backpressure. Microinjection מבוצע באמצעות מיקרוסקופ עם הבמה נע (לייקה) בהגדלה של 10 x ו micromanipulator (לייקה). אם יש צורך, בשלב מיקרוסקופ העלתה עשוי להיות מוכן על ידי הערמה שקופיות מיקרוסקופ 4-5. מניחים את תלוש לכסות נושאת את העוברים על הבמה המוגבהת. עוברים מוזרקים בזווית 150 °; חדירה צריך להיות בקוטב האחורי. עבור זריקה, אני ממשיך המחט נייחים להזיז את הבמה של המיקרוסקופ. כאשר המחט חדשה, קצה אטום ובדרך כלל הפסקות הזריקה הראשונה. אני תמיד לחשב את הלחץ הדרוש כדי לגרש טיפה קטנה של דנ&quot;א לתוך השמן בין כל הזרקה. </li><li style=";text-align:right;direction:rtl"> אני קובע את זמן ההזרקה כדי 0.2-0.9 שניות ו 1000 hPa כאשר המחט חדשה. אני להשתנות את הלחץ ואת הזמן עד טיפה קטנה נראה יוצא מן המחט לתוך השמן. Backpressure צריך להיות מוגדר סביב 100. כאשר קצה המחט מקבל שחוקים, הלחץ הזרקת צריך להיות מופחת כדי לשמור על נפח הזרקה נמוך (כ 300 hPa). אחרי זה, קצה המחט גדולה מדי, והוא הורג את מרבית העוברים. </li><li style=";text-align:right;direction:rtl"> לאחר ההזרקה, להסיר את השמן עם נייר ומכסים את השקופית עם עוברי ומניחים בצלחת פטרי עם חיץ איזוטוני. מניחים את הכלים insectory ולהשאיר אותם שם עד עוברים את הפתח. הם מתחילים לבקוע לאחר 2-6 ימים. מעבירים את הזחלים למים מזוקקים עם קמצוץ של מזון דגים הקרקע. </li></ol> הערות ה-DNA להזרקה: פתרון ה-DNA פלסמיד להזרקה היה מבודד באמצעות ערכת פלסמיד EndoFree.Construct פלסמיד ומערבבים Helper פלסמיד יחד בריכוז הנכון, זירז עם isopropanol. גלולה נשטף עם אתנול 70% לפני resuspending במאגר ההזרקה. לפני הזריקה, DNA נקי באמצעות Millex-GV העמודה. Izotonic חיץ: <table border="1" cellpadding="8" style=";text-align:right;direction:rtl"><tbody><tr><td> 1.68 M NaCl - 88.3 מ&quot;ל 1.68 M NaCl - 2.9 מ &#39; 1 M Hepes -10.7 מ&quot;ל M 1.12 CaCl 2-2.2 מ&quot;ל DH 2 O - 896 מ&quot;ל </td><td> או </td><td> 5 M NaCl - 29.67 מ&quot;ל 1 M KCl - 4.87 מ&quot;ל 1 M Hepes - 10.7 מ&quot;ל M 1.12 CaCl 2-2.2 מ&quot;ל DH 2 O - 952.56 מ&quot;ל </td></tr></tbody></table> התאם ל-pH 7.2

<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN" "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en" lang="en">	
		
		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Injection Quartz needles</td>
<td>&nbsp;</td>
<td>Sutter Instrument</td>
<td>&nbsp;</td>
<td>O.D.- 1.0 mm, I.D.-0.70, 10 cm length, are prepared by drawing capillary tubing into a fine 3-8 um with a shaft of approximately 100-300 mm micropipette puller. </td>
</tr>
<tr>
<td>Quartz puller</td>
<td>Tool</td>
<td>Sutter Instrument Co</td>
<td>P-2000</td>
<td>Set-up: Heat-275, Fil-3, Vel-38, Del-250, Pull-141</td>
</tr>
<tr>
<td>EndoFree plasmid kit</td>
<td>&nbsp;</td>
<td>Quiagen</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Isopropanol</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Millex-GV column</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>SLGV RO4 NL</td>
<td>for DNA cleaning</td>
</tr>
<tr>
<td>Construct DNA</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>0.5 mg/ml </td>
</tr>
<tr>
<td>Helper plasmid</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>0.3 mg/ml </td>
</tr>
<tr>
<td>Injection solution</td>
<td>Buffer</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>1x solution: 5mM KCl, 0.1mM sodium phosphate, ph 6. 8</td>
</tr>
<tr>
<td>Isotonic buffer</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>see recipe in the protocol section</td>
</tr>
<tr>
<td>Dessication oil</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>Halocarbon oil 700:Halocarbon oil 27(1:1). Mix well. Prepare in advance of microinjection.</td>
</tr>
<tr>
<td>Blood</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>to feed mosquitos</td>
</tr>
<tr>
<td>Anopheles stephensi</td>
<td>Animal</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>Mosquitos, male and female.</td>
</tr>
<tr>
<td>Drosophila culture vial</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>for egg laying</td>
</tr>
<tr>
<td>Petri dishes</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>with isotonic buffer, for embryo hatching</td>
</tr>
<tr>
<td>Fine paintbrush</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>Sable # 0000</td>
</tr>
<tr>
<td>3MM Whatman paper</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Forceps #5</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>double sided tape</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Microloader</td>
<td>&nbsp;</td>
<td>Eppendorf</td>
<td>5242 956.003</td>
<td>&nbsp;</td>
<tr>
<td>Transjector</td>
<td>&nbsp;</td>
<td>Eppendorf</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Microscope</td>
<td>&nbsp;</td>
<td>Leica</td>
<td>&nbsp;</td>
<td>with moving stage and micromanipulator, set at 10x magnification </td>
</tr>		</tbody>
		</table>
		</div>
			<div>
					</div>

injection of an. stephensi embryos to generate malaria-resistant mosquitoes

המבוא של גנים חיצוניים לתוך הגנום של יתושים דורש טכניקות microinjection המותאמים למין ספציפי של עניין. זה פרוטוקול וידאו מדגים שיטה המשמשת את המעבדה ג&#39;יימס microinject בונה DNA לעוברי אנופלס stephensi עבור הדור של יתושים טרנספורמציה. טכניקות להכנת מחטים microinjection, איסוף והכנת עוברי וביצוע microinjection מומחשים.

Watch this Scientific Journal Video about Injection of An. stephensi Embryos to Generate Malaria-resistant Mosquitoes at JoVE.com

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

אנופלס יתושים stephensi הם וקטורים עבור מלריה בהודו המאכלסים וברחבי אסיה. וידאו זה מדגים את הטכניקה לביצוע microinjections של מין זה עם transgenes כי יעניק התנגדות מלריה אל היתוש. הרבה המתודולוגיה הפגינו וידאו זה הוא ישים טכניקות microinjection של מינים יתושים אחרים.

הזרקת. עוברים stephensi ליצור מלריה עמידים יתושים

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

injection-an-stephensi-embryos-to-generate-malaria-resistant

Research

JoVE Journal

Biology

12.9K Views.  University of California, Irvine (UCI). אנופלס יתושים stephensi הם וקטורים עבור מלריה בהודו המאכלסים וברחבי אסיה. וידאו זה מדגים את הטכניקה לביצוע microinjections של מין זה עם transgenes כי יעניק התנגדות מלריה אל היתוש. הרבה המתודולוגיה הפגינו וידאו זה הוא ישים טכניקות microinjection של מינים יתושים אחרים.

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

Microinjection of A. aegypti Embryos to Obtain Transgenic Mosquitoes

In this video, Nijole Jasinskiene demonstrates the methodology employed to generate transgenic Aedes aegypti mosquitoes, which are vectors for dengue fever.  The techniques for correctly preparing microinjection needles, dessicating embryos, and performing microinjection are demonstrated.

In this video, Nijole Jasinskiene demonstrates the methodology employed to generate transgenic Aedes aegypti mosquitoes, which are vectors for dengue fever. The techniques for correctly preparing microinjection needles, dessicating embryos, and performing microinjection are demonstrated.

Microinjection עוברים א aegypti לקבל יתושים טראנסגנטי

In this video, Nijole Jasinskiene demonstrates the methodology employed to generate transgenic Aedes aegypti mosquitoes, which are vectors for dengue ...

In Utero Intraventricular Injection and Electroporation of E16 Rat Embryos

In-utero in-vivo injection and electroporation of the embryonic rat neocortex provides a powerful tool for the manipulation of individual progenitors lining the walls of the lateral ventricle. This technique is now widely used to study the processes involved in corticogenesis by over-expressing or knocking down genes and observing the effects on cellular proliferation, migration, and differentiation. In comparison to traditional knockout strategies, in-utero electroporation provides a rapid means to manipulate a population of cells during a specific temporal window. In this video protocol, we outline the experimental methodology for preparing rats for surgery, exposing the uterine horns through laporatomy, injecting DNA into the lateral ventricles of the developing embryo, electroporating DNA into the progenitors lining the lateral wall, and caring for animals post-surgery. Our laboratory uses this protocol for surgeries on E15-E21 rats, however it is most commonly performed at E16 as shown in this video.

בשנת הזרקה תוך חדרי ברחם ו electroporation של E16 עוברים עכברוש

In-utero in-vivo injection and electroporation of the embryonic rat neocortex provides a powerful tool for the manipulation of individual progenitors ...

In Utero Intraventricular Injection and Electroporation of E15 Mouse Embryos

In-utero in-vivo injection and electroporation of the embryonic mouse neocortex provides a powerful tool for the manipulation of individual progenitors lining the walls of the lateral ventricle. This technique is now widely used to study the processes involved in corticogenesis by over-expressing or knocking down genes and observing the effects on cellular proliferation, migration, and differentiation. In comparison to traditional knockout strategies, in-utero electroporation provides a rapid means to manipulate a population of cells during a specific temporal window. In this video protocol we outline the experimental methodology for preparing mice for surgery, exposing the uterine horns through laporatomy, injecting DNA into the lateral ventricles of the developing embryo, electroporating DNA into the progenitors lining the lateral wall, and caring for animals post-surgery. Our laboratory uses this protocol for surgeries on E13-E16 mice, however, it is most commonly performed at E15, as shown in this video. 

בשנת הזרקה תוך חדרי ברחם ו electroporation E15 של עוברי עכברים

In-utero in-vivo injection and electroporation of the embryonic mouse neocortex provides a powerful tool for the manipulation of individual progenitors ...

Microinjection of Zebrafish Embryos to Analyze Gene Function

One of the advantages of studying zebrafish is the ease and speed of manipulating protein levels in the embryo.  Morpholinos, which are synthetic oligonucleotides with antisense complementarity to target RNAs, can be added to the embryo to reduce the expression of a particular gene product.  Conversely, processed mRNA can be added to the embryo to increase levels of a gene product.  The vehicle for adding either mRNA or morpholino to an embryo is microinjection.  Microinjection is efficient and rapid, allowing for the injection of hundreds of embryos per hour.  This video shows all the steps involved in microinjection.  Briefly, eggs are collected immediately after being laid and lined up against a microscope slide in a Petri dish.  Next, a fine-tipped needle loaded with injection material is connected to a microinjector and an air source, and the microinjector controls are adjusted to produce a desirable injection volume.  Finally, the needle is plunged into the embryo's yolk and the morpholino or mRNA is expelled.

This video shows how morpholino or mRNA can be injected into zebrafish embryos at the one-cell stage to decrease or increase the level of specific gene products during subsequent development.

Microinjection של עוברי דג הזברה לנתח פונקציה ג&#39;ין

One of the advantages of studying zebrafish is the ease and speed of manipulating protein levels in the embryo.  Morpholinos, which are synthetic ...

RNAi Interference by dsRNA Injection into Drosophila Embryos

Genetic screening is one of the most powerful methods available for gaining insights into complex biological process 1. Over the years many improvements and tools for genetic manipulation have become available in Drosophila 2. Soon after the initial discovery by Frie and Mello 3 that double stranded RNA can be used to knockdown the activity of individual genes in Caenorhabditis elegans, RNA interference (RNAi) was shown to provide a powerful reverse genetic approach to analyze gene functions in Drosophila organ development 4, 5.
Many organs, including lung, kidney, liver, and vascular system, are composed of branched tubular networks that transport vital fluids or gases 6, 7. The analysis of Drosophila tracheal formation provides an excellent model system to study the morphogenesis of other tubular organs 8. The Berkeley Drosophila genome project has revealed hundreds of genes that are expressed in the tracheal system. To study the molecular and cellular mechanism of tube formation, the challenge is to understand the roles of these genes in tracheal development. Here, we described a detailed method of dsRNA injection into Drosophila embryo to knockdown individual gene expression. We successfully knocked down endogenous dysfusion(dys) gene expression by dsRNA injection. Dys is a bHLH-PAS protein expressed in tracheal fusion cells, and it is required for tracheal branch fusion 9, 10. dys-RNAi completely eliminated dys expression and resulted in tracheal fusion defect. This relatively simple method provides a tool to identify genes requried for tissure and organ development in Drosophila.

RNAi Interference by dsRNA Injection into Drosophila Embryos

RNA interference has been proven very effective to analyze gene function in Drosophila tracheal development. A detailed protocol used by Jiang lab to inject dsRNA into fly embryos to knockdown gene expression is illustrated. This technique has the potential for screening genes required for tissue and organ development in Drosophila.

RNAi הפרעה על ידי הזרקה לתוך dsRNA תסיסנית עוברים

Genetic screening is one of the most powerful methods available for gaining insights into complex biological process 1. Over the years many improvements ...

An in vivo Crosslinking Approach to Isolate Protein Complexes From Drosophila Embryos

Many cellular processes are controlled by multisubunit protein complexes. Frequently these complexes form transiently and require native environment to assemble. Therefore, to identify these functional protein complexes, it is important to stabilize them in vivo before cell lysis and subsequent purification. Here we describe a method used to isolate large bona fide protein complexes from Drosophila embryos. This method is based on embryo permeabilization and stabilization of the complexes inside the embryos by in vivo crosslinking using a low concentration of formaldehyde, which can easily cross the cell membrane. Subsequently, the protein complex of interest is immunopurified followed by gel purification and analyzed by mass spectrometry. We illustrate this method using purification of a Tudor protein complex, which is essential for germline development. Tudor is a large protein, which contains multiple Tudor domains - small modules that interact with methylated arginines or lysines of target proteins. This method can be adapted for isolation of native protein complexes from different organisms and tissues.

An in vivo Crosslinking Approach to Isolate Protein Complexes From Drosophila Embryos

Multi-component protein complexes play crucial roles during cellular function and development. Here we describe a method used to isolate native protein complexes from Drosophila embryos after in vivo crosslinking followed by purification of the crosslinked complexes for subsequent structure-function analysis.

In vivo גישת Crosslinking לבודד חלבון מן מכלולים דרוזופילה עוברים

Many cellular processes are controlled by multisubunit protein complexes. Frequently these complexes form transiently and require native environment to ...

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.

מבחני לבדיקת השפעות רעילים של תרופות אפיגנטיים בפיתוח, פוריות והישרדות של יתושים מלריה

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

Protocols for Testing the Toxicity of Novel Insecticidal Chemistries to Mosquitoes

New classes of insecticides with novel modes of action are needed to control insecticide resistant populations of mosquitoes that transmit diseases such as Zika, dengue and malaria. Assays for rapid, high-throughput analyses of unformulated novel chemistries against mosquito larvae and adults are presented. We describe protocols for single point-dose and dose response assays to evaluate the toxicity of small molecule chemistries to the Aedes aegypti vector of Zika, dengue and yellow fever,&#160;the malaria vector, Anopheles gambiae and the northern house mosquito, Culex quinquefasciatus, on contact and via ingestion. As an example, we evaluated the toxicity of amitriptyline, a small molecule antagonist of G protein-coupled receptors, via larval, adult topical and adult blood-feeding assay. The protocols provide a starting point to investigate insecticide potential. Results are discussed in the context of additional experiments to explore product applications and mechanisms for delivery.

Protocols are described for assessing the toxicity of chemistries to immature and adult mosquitoes for development as new classes of larvicides, adulticides and endectocides. The protocols enable high-throughput testing of multiple chemistries at single-point dose and subsequent evaluation via dose response assay to determine toxicity on contact or via ingestion.

פרוטוקולים לבדיקת הרעילות של הרומן בדיקות, חרקים הביוכימיה על היתושים

New classes of insecticides with novel modes of action are needed to control insecticide resistant populations of mosquitoes that transmit diseases such ...

Forced Salivation As a Method to Analyze Vector Competence of Mosquitoes

Vector competence is defined as the potential of a mosquito species to transmit a mosquito-borne virus (mobovirus) to a vertebrate host. Viable virus particles are transmitted during a blood meal via the saliva of an infected mosquito. Forced salivation assays allow determining the vector potential on the basis of single mosquitoes, avoiding the use of animal experiments. The method is suitable to analyze a large number of mosquitoes in one experiment within a short period of time. Forced salivation assays were used to analyze 856 individual mosquitoes trapped in Germany, including two different Culex pipiens pipiens biotypes, Culex torrentium as well as Aedes albopictus, which were experimentally infected with Zika virus (ZIKV) and incubated at 18 &#176;C or 27 &#176;C for two and three weeks. The results indicated the lack of vector competence of the different Culex taxa for ZIKV. In contrast, Aedes albopictus was susceptible to ZIKV, but only at 27 &#176;C, with transmission rates similar to an Aedes aegypti laboratory colony tested in parallel.

For efficient control of mosquito borne virus transmission, the knowledge of the vector potential of respective mosquitoes is of particular interest. We describe forced salivation as a method to analyze vector competence of Aedes albopictus and three different Culex taxa for the transmission of Zika virus.

הכריחו ריור כשיטה לנתח מיומנות וקטור של יתושים

Vector competence is defined as the potential of a mosquito species to transmit a mosquito-borne virus (mobovirus) to a vertebrate host. Viable virus ...

A Blood-Free Diet to Rear Anopheline Mosquitoes

Malaria research requires large-scale breeding and production conditions for mosquitoes (Anopheles spp.) in captivity. The sustainable and reliable production of mosquitoes is currently inhibited by the supply of fresh vertebrate blood. Alternatives to blood are required to promote efficient control strategies for malaria and other vector borne diseases that are transmitted by blood feeding insects. With this in mind, artificial liquid diets were formulated as substitutes for fresh vertebrate blood. Herein we report a blood-free artificial liquid diet that delivers feeding rates similar to blood and mimics the physiological effects of a fresh vertebrate blood meal. The diet induces ovarian and egg maturation of Anopheles mosquitoes and also produces good larval survival and development of functional adults. The formulated blood-free liquid diet is an important advance towards sustainable mosquito breeding in captivity and will reduce the maintenance costs of mosquito colonies and eliminate the need for fresh vertebrate blood.

A protocol is presented for formulation of a blood-free artificial diet to feed Anopheles mosquitoes in captivity. This diet has a similar performance to vertebrate blood and triggers oogenesis and egg maturation and produces viable adult progeny.

דיאטה ללא דם ליתושים Anopheline אחוריים

Malaria research requires large-scale breeding and production conditions for mosquitoes (Anopheles spp.) in captivity. The sustainable and reliable ...