This Test Guideline (TG) describes a honey bee brood acute toxicity test under laboratory conditions. The method aims at the determination of the lethal dose (72-h LD50) following single exposure of larvae to a chemical.

On day 1 (D1) of the study, first instar synchronised larvae are taken from the comb of three colonies (thirty-six larvae in total per group) and individually placed into 48 well-plates where they are fed a standardized amount of artificial diet. On day 4 (D4) of the test, a single dose of the test chemical is administered to the larvae with the diet. Each group of thirty-six larvae is administered a given dose in a range of five increasing doses. Mortalities are recorded on D5, D6, and D7 of the test. The 72-hr LD50 is calculated for larvae (cumulative mortality at D7). The study report also includes a number of important elements, in particular regarding test conditions (e.g., temperature and humidity).