title: Survival of emu (Dromaius novaehollandiae) sperm preserved at subzero temperatures and different cryoprotectant concentrations.
abstract: Three experiments were conducted to optimize the protocol for cryopreservation of emu sperm. Ejaculates were collected from trained male emus then diluted 1:1 and pooled before allocation to treatments and measured for sperm viability, motility, egg membrane penetration ability, membrane stability, and morphology. In Experiment 1, semen was either cooled to 5 °C after dilution or diluted with a precooled to 5 °C diluent before cooling to 5 °C and then frozen at liquid nitrogen vapor temperatures of -140 °C and -35 °C, with 6% or 9% dimethylacetmide (DMA; a permeating cryoprotectant) and compared for sperm functions. The percentages of viable (42.8 ± 1.1%), normal (39.0 ± 1.3%), and motile (29.8 ± 1.3%) sperm were higher (P < 0.001) for semen frozen at -14 °C with 9% DMA (path 2) than for all other combinations. In Experiment 2, we assessed the value of combining DMA and trehalose in the diluent. Combining trehalose (3% to 9%) with DMA (3% to 9%) prior to freezing reduced (P < 0.001) the percentages of postthaw viable (by 4 to 9 ± 1.2%), normal (by 5 to 11 ± 1.3%), and motile sperm (by 13 to 17 ± 2.5%) and the number of holes on the perivitelline layer (by 27 to 29 holes/mm(2)). Postthaw function was best preserved with 9% DMA alone. In experiment 3, we investigated the possibility of increasing DMA concentrations from 6% to 24%. Postthaw sperm viability (52 to 55 ± 2.3%) and morphology (48 to 51 ± 1.7%) were higher (P < 0.05) with 18% and 24% than with 6% to 12% DMA and did not differ between 18% and 24% DMA. However, sperm motility (36 to 43 ± 2.9%) and the number of perivitelline holes were similar (P > 0.05) for 9% to 18% DMA (36 to 55 ± 12%). We concluded that adding 6% to 9% trehalose to the diluent offered no advantage, and that the current best practice for preserving postthaw function in emu sperm is to dilute semen with a precooled to 5 °C diluent and use 18% DMA.
authors.affiliation: School of Animal Biology, Faculty of Natural and Agricultural Sciences, the University of Western Australia, Crawley, WA, Australia. sushilsood@gmail.com
authors.forename: S
authors.initials: S
authors.lastname: Sood
authors.forename: I A
authors.initials: IA
authors.lastname: Malecki
authors.forename: A
authors.initials: A
authors.lastname: Tawang
authors.forename: G B
authors.initials: GB
authors.lastname: Martin
mesh_terms: D000081:Acetamides / Q000008:administration & dosage; D000818:Animals; D002470:Cell Survival; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D003451:Cryoprotective Agents / Q000008:administration & dosage*; D020312:Dromaiidae; D005260:Female; D006358:Hot Temperature; D008297:Male; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary*; D013081:Sperm Motility; D013084:Sperm-Ovum Interactions; D013094:Spermatozoa / Q000002:abnormalities / Q000502:physiology*; D013696:Temperature; D014199:Trehalose / Q000008:administration & dosage
chemical_list: D000081:Acetamides; D003451:Cryoprotective Agents; D014199:Trehalose; C013959:dimethylacetamide