4.5.4.2.6.2. Assessment of safety of product for honeybees

1. Record bee mortality inside and adjacent to the hive daily or at least three times a week throughout the three stages of the experiment

The use of dead-bee traps is recommended (see section 2. ‘Estimating the number of dead bees expelled from a colony’ of the BEEBOOK paper on miscellaneous methods (Human et al., 2013)).

2. Monitor the morbidity, mortality, as well as the size and development of surviving colonies at the time of the first flight in spring and thereafter (see the BEEBOOK paper on estimating colony strength (Delaplane et al., 2013)) if applicable (envisaged therapeutic use in autumn or winter).

3. Measure flight activity of bees during the trial (see the BEEBOOK paper on behavioural methods (Scheiner et al., 2013)).

This verifies whether the product influences foraging activity of the treated colonies.

4. Measure honey production.

This verifies whether the product influences the productivity of the treated colonies

5. Quantify brood area of test colonies during the three phases and compare to the control group (see the BEEBOOK paper on estimating colony strength (Delaplane et al., 2013)).

In cases in which the product is intended for use in colonies with brood, the demonstration of safety for all stages of brood should be carried out (see the BEEBOOK article on toxicology methods by Medrzycki et al. (2013)). An additional method to determine effect of the product tested on brood is to determine which of feeding incompetence of worker bees and direct adverse effects on eggs and larvae occurred. For this:

1. Leave frames with eggs and larvae to develop in the hive until a chosen stage of the larval stage after applying therapeutic doses of the test product.

2. Monitor feeding behaviour of these larvae by measuring the amount of food found in their cells.

By comparing development of brood and amount of larval food and taking into account the ratio between quantity of brood and number of worker bees between control and test groups, it is possible to differentiate between effects of feeding incompetence of worker bees and direct adverse effects on eggs and larvae after application of the product.

3. Verify the presence of the live queen at the end of the experiment.

A significant difference in queen survival between test and control groups indicates an effect of the treatment.