5.2. Injection of test substances into the hemocoel
Potential wax moth control agents can be injected directly into the larval hemocoel (West and Briggs, 1968). Possible treatment compounds include bacterial toxins (such as Bacillus thuringiensis), fungal toxins (i.e. Vilcinskas et al., 1997), insecticides, plant resins, etc. This procedure also can be used to initiate immune responses in wax moths and for other purposes beyond simple pest control.
- Raise larvae per Section 3 to 100-200 mg/individual.
- Prepare solutions (treatment and control) per the needs and conditions of the experiment.
- Using a calibrated microinjection apparatus
with a 27-gauge needle, insert the needle into the lateral integument about
halfway down the body (be careful not to damage internal organs).
Note: Alternatively, microliter cemented needle syringes fitted with a 26 gauge needle may be used for microinjections.
- Inject a consistent, desired volume into each larva.
- Repeated injections are discouraged because of the size of the insect and possible associated damage (Stephens, 1959).
- Observe specimens for desired change (see Section 5.1. for parameters).
Considerations: In microinjection experiments, care should be taken to maintain a clean workspace and equipment to limit physiological change due to contamination rather than the experimental treatment. One should also include controls for the study which include moths injected with Ringers solution. Solutions should be prepared so they are physiologically compatible with the larval hemocoel. It is possible for large injection volumes to cause non-treatment associated effects. West and Briggs (1968) had successful results injecting 20 ml bolus volumes though a range of injection volumes are reported in the literature.