New Methods and Media for the Centrifugation of Honey Bee (Hymenoptera: Apidae) Drone Semen

ABSTRACT Centrifugation of Apis mellifera L. drone semen is a necessary step in the homogenization of semen pools for the enlargement of the effective breeding population, as well as in the collection of semen by the so-called washing technique. It is also of interest for the removal of cryoprotectants after cryopreservation. The adoption of methods involving semen centrifugation has been hampered by their damaging effect to sperm. Here, we tested four new diluents as well as three additives (catalase, hen egg yolk, and a protease inhibitor), using sperm motility and dual fluorescent staining as indicators of semen quality. Three of the new diluents significantly reduced motility losses after centrifugation, as compared with the literature standard. Values of motility and propidium iodide negativity obtained with two of these diluents were not different from those measured with untreated semen. The least damaging diluent, a citrate-HEPES buffer containing trehalose, was then tested in an insemination expe...