title: In vivo validation of in vitro quality tests for cryopreserved honey bee semen.
abstract: Development of cryopreservation protocols for honey bee semen is hampered by the lack of validated laboratory tests that allow the prediction of in vivo performance of frozen-thawed semen. Here we analyzed correlations between seven in vitro tests and indicators of semen performance after insemination. These tests included measures of motility, cell conformation, and membrane permeability before and after exposure to physiochemical stress. We show that the proposed protocol for motility measurement yields results that correlate well with the number of sperm reaching the storage organ of queens (correlation coefficient ρ=0.67) and the proportion of viable eggs in inseminated queens (ρ=0.48). The conventional live/dead assay of membrane permeability by dual fluorescent staining and a new test based on the leakage of the glycolytic enzyme glucose-phosphate-isomerase (GPI) from damaged cells were also correlated to the number of sperm reaching the spermatheca (ρ=0.54 and -0.61, respectively). We conclude that motility, live/dead-staining and the assay for GPI-leakage are valuable tools for the improvement of cryopreservation of honey bee semen.
authors.affiliation: Institute for Bee Research, Friedrich-Engels-Strasse 32, 16540 Hohen Neuendorf, Germany. jakob.wegener@rz.hu-berlin.de
authors.forename: Jakob
authors.initials: J
authors.lastname: Wegener
authors.forename: Tanja
authors.initials: T
authors.lastname: May
authors.forename: Ulrich
authors.initials: U
authors.lastname: Knollmann
authors.forename: Günter
authors.initials: G
authors.lastname: Kamp
authors.forename: Karin
authors.initials: K
authors.lastname: Müller
authors.forename: Kaspar
authors.initials: K
authors.lastname: Bienefeld
mesh_terms: D000818:Animals; D001516:Bees / Q000166:cytology / Q000502:physiology*; D002463:Cell Membrane Permeability; D002470:Cell Survival; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D005260:Female; D007314:Insemination; D008297:Male; D012661:Semen / Q000166:cytology* / Q000502:physiology; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary*; D013081:Sperm Motility