Methods in Anopheles Research - MR4

Chapter 3 : Specific Anopheles Techniques3.7 Anopheles Mating3.7.2 Forced CopulationPage 3 of 6Figure 3.7.2.3. Holding male with vacuumpressure, gently stab the male through thethorax taking care to not crush the head.Figure 3.7.2.4. Properly pinned male.6. Place one container of 10 females into the anesthetizing chamber and leave it for 6-10 seconds(depending on the species and strength of ether).7. Watch the females closely, and once they have all fallen from the sides, remove them from thechamber. Do not leave females in the ether too long or they will not recover from the treatment.8. Gently disperse the females onto a piece of filter paper and position ventral side up. Take a mountedmale, pinch off the head and hind-tarsi (Figure 3.7.2.5), and then gently stroke the abdomen of themale over the female’s abdomen to stimulate the claspers to open (Figure 3.7.2.6).9. Place the male at a 45-90° angle venter-to-venter with the female until the male clasps the female(Figure 3.7.2.7). Leave clasped for 1-2 seconds then pick up both using the male on the pin. If matedsuccessfully, they will remain attached for several seconds. 2 Successful mating has usually occurredif they remain attached for 3-5 seconds (Figure 3.7.2.8).10. Place male and female together into a new cage to allow female to recover from the anesthesia.11. Ensure that the females are recovering from the anesthesia by gently blowing into the recovery cage.If females are not waking up within 10 minutes, anesthetize more lightly.2 It is possible to reuse males to mate more than one female.