Infecting Flies with Bacterial Suspension Using a Nanoinjector

To infect flies, begin with a narrow-diameter glass needle filled with mineral oil.

Secure the needle onto the nanoinjector plunger with an O-ring and a spacer for precise and controlled injection, ensuring minimal damage to the flies.

Release the mineral oil, leaving a small amount in the needle as a barrier between the bacterial solution and the injector. Load the needle with the bacterial suspension. 

Position an anesthetized Drosophila melanogaster ventrolateral side-up.

Inject the needle into the fly's anterior abdomen, piercing through the cuticle and epithelial layer.  Deliver the bacterial suspension into the hemocoel — a body cavity containing hemolymph — a nutrient-rich circulatory fluid. 

Within the hemocoel, immune cells called hemocytes, circulate alongside the fat body, a specialized tissue equipped with pattern recognition receptors.

Allow the injected flies to recover in a vial containing medium. Injected bacteria utilize the nutrients from hemolymph and multiply, increasing the bacterial load, and establish a systemic infection.

Fat body cell receptors interact with bacterial antigens, triggering an immune response to produce antimicrobial peptides, ultimately eliminating some bacteria.

Circulating hemocytes perform phagocytic activity, engulfing and degrading a few bacteria. However, some bacteria evade the host's immune response and survive within the flies, establishing a bacterial infection.