184.108.40.206.2. Preparation of the colonies
1) The OECD 75 recommends using small healthy honey bee colonies (e.g. Mini Plus, nuclei, etc.) for the test, but it is also possible to use small commercial bee colonies. However, the size of the colonies should be adapted to the size of the crop area within the tunnels.
2) All colonies of one set or study have to be produced at the same time from colonies headed by sister queens to guarantee that the colonies in all variants are uniform as far as possible (Delaplane et al., 2013b). The colonies must be headed by sister queens which are the progeny of the same queen and mated at the same place in order to minimise genetic variability.
3) The bee colonies should be free of clinical symptoms of disease (e.g. nosema, Amoeba, chalkbrood, sacbrood, and American or European foulbrood) or pests (Varroa destructor): see BEEBOOK Volume II. The colonies should be free of unusual occurrences (e.g. presence of dead bees, dark-“bald”-bees, “crawlers” or flightless bees, unusual brood distribution patterns or brood age structure).
4) After establishment of the colonies within the tunnels, all hives are equipped with a dead bee trap at the entrance to count the number of dead bees (Human et al., 2013).
5) The colonies should be established in the tunnels shortly before full flowering of the crop and at least three days before application in order to allow the bees to adapt to the conditions in the tunnels.
6) The colonies should be exposed to the treated crop in the tunnels for a period of at least 7 days after the application.