Begin by transferring all the collected Noctuidae insects from the net bag to a net cage. Next, provide a Petri dish containing a sterilized 10%honey water solution. Set the cage in a stable environment.
Choose wild type female insects that have ventured inside the cage that same day. Carefully transfer these insects into individual vial tubes, ensuring each tube is sealed with a cotton lid. For insect dissection, first place a freshly paralyzed female insect into a dissecting Petri dish containing absolute ethanol.
To ensure a clean dissection, infiltrate the insect with absolute alcohol and rinse it in clean water. Next, using forceps, separate the dorsal wings at the junction of the chest and abdomen. Transfer the abdomen into a new disposable dish filled with two to five millimeters of water.
Using dissecting forceps, gently peel the abdominal exoskeleton, starting from the pointed mouth dorsoventrally to the tail. Replicate these steps on the other side and place it in clean water to disperse the intact tissues. Now delicately peel off the epidermis fat tissues using forceps and gently pull and release the ovaries.
With a pair of dissecting forceps, gently remove fat particles and any surrounding organs from the ovaries. Gradually peel off the mating sac from the middle and eliminate fat particles attached to the ovarian tubes. Gently hold onto the ovary and mating sac from the vertical posterior end, then slowly unfold it downwards.
To prevent damage, transfer the ovary into a new or clean Petri dish filled with water. Hold the ovarian tip and unfold the ovary inwardly. Assess the egg maturity for the insect using color and size indicators.
Then judge the ovarian grade according to the egg development. After dissection, make sure to separate the female mating sac tissues from the intact ovaries. Examine the morphology as mating sac anatomical features differ from species to species.
Next, analyze the ovarian anatomy by dividing the ovarian tissues into five distinct grades. Lastly, capture images of the ovary using a digital camera as per the requirements of the experiment. Four different stages of ovarian development were observed, including the milky white transparent stage with underdeveloped ovaries, the yolk deposition stage with large slightly immature eggs, the ripening stage with bright yellow bead-shaped eggs and the end spawning stage with few or no egg grains.
The mating sac of a wild type insect Helicoverpa armigera had a columnar right helix with an inner lumen. Mythimna separata had a G-shaped mating sac with dark brown bands. The mating sac of Agrotis ipsilon was linear with large coils outer back cysts.
Spaelotis valida had a slightly brownish thin J-shaped mating sac, while Spodoptera exigua showed a white translucent dumbbell-shaped sac with a bubble cyst in the middle. The mating sac of Spodoptera litura was red with an upper milky white transparent mouth and a faintly visible internal cystic cavity. Pseudoptera lepigone mating sac was small and coiled with closed outer cysts.
Mamestra brassicae had olive colored double coiled sacs with outer cysts. Old worm ovarian grading showed transparent and elastic ovarian ducts with grade one visible oocytes. At grade two, the ovarian duct became transparent with tender yellow eggs.
At grade three, large fat bodies were absent with the duct being thickest at the ovipositor. Grades four and five showed clusters and fewer eggs in the ovaries with pale yellow and greenish color. All five ovarian grades were also observed in the armyworm, in the tarot caterpillar, and in the bollworm.
