126.96.36.199. Artificial infestation
1. Collect mites and L5 bee larvae from natural brood combs as described in section 3.1.4.’Collecting mites from brood’.
2. Place the larva in a gelatin cell by holding its dorsum between thumb and first finger to get it stretched.
3. Insert the mite with a fine paint brush.
Gelatin cells of different diameters were tested; as a general rule, the narrower the cell diameter the higher the reproduction, but the higher the chance of injuring the larva while inserting into the cell; the best compromise is achieved with gelatin cells of 6.5 mm Æ.
4. Place in an incubator at 34.5 °C, 75 % RH.
5. Place the cells so that pupae are laying on their back.
Geotaxis is an important cue for varroa behaviour (Donzé and Guerin 1994).
6. Fix the cell to a substrate to avoid rolling and manipulate only occasionally for observations.
Pros: high percentage of fertile mites and offspring number close to natural infestations in colonies can be obtained; transparent cells allow observation of behaviour; higher sample size possible; in vitro procedure allows complete control over the infestation state of the bee since workers do not have the opportunity to remove infested brood.
Cons: tedious; non-natural infestation; non-natural cell size.
Perspectives: With the aim of developing a complete rearing method for the mite, Nazzi and Milani (unpublished data) and Dietemann, Zheng and Su (unpublished data) carried out preliminary trials aiming at obtaining several reproduction cycles in the laboratory, i.e. artificially breeding mites that were born under laboratory conditions. Attempts were discouraging and continuing efforts are needed.