220.127.116.11.2. Identifying and replicating the treatment modalities
The number of modalities is defined by the objectives of the study and includes at least the following groups:
1) A control - untreated sugar water, often containing the solvent used to dissolve the test compound in the treatment doses. The control provides the evaluation standard in the assessments.
2) The toxic reference - This reference verifies bee sensitivity to toxic compounds. The toxic standard validates the test. Dimethoate is the main toxic standard used and provides a high subsequent mortality at known doses. It is usually administered at 2-3 doses to cover the expected LD50 value. The expected oral LD50 for dimethoate ranges from 0.10 to 0.35 µg AI/bee.
3) The test compound at five doses.
Consequently, there are at least 9 “groups” for each study (the control, the toxic standard administered at 3 doses, and the test compound administered at 5 doses). Each group should be replicated three times (i.e. with 3 hoarding cages of 10 to 20 bees) (EPPO 2010a; OECD, 1998a, 1998b). The CEB (2011) guideline requires three “runs” of three replicates/run (3 x 3).