4.6. Exclusion of grafting effect

A commonly used option, depending on and if justified by the research aim, may be the exclusion of grafting effects by excluding dead larvae from the experiment 24 hours after grafting. However, this is impossible if susceptibility to a certain substance or pathogen decreases with increasing age, as is the case with Paenibacillus larvae, the etiological agent of American Foulbrood. If experiments can be performed equally well with second or older instar larvae, then it might be more appropriate to graft the more robust second or older instar larvae right away instead of replacing larvae in the course of the experiment. In acute toxicity testing, sometimes a surplus of larvae is used at day four to replace larvae which died prior to treatment administration. Of course, this procedure increases the quality of the resulting honey bees and thus biases results. Whether or not to replace larvae during the experiment must be considered carefully prior to use, explicitly documented, and critically discussed when evaluating the results.