Search

Preserving genetic diversity of the critically endangered Addra gazelle ( Nanger dama ruficollis ) could be enhanced through the use of frozen-thawed sperm and artificial insemination. Our aim was to characterize Addra ejaculate traits and to assess the effects of cholesterol-loaded cyclodextrin (CLC) on sperm cryosurvival. Fresh ejaculates were treated with CLC (0, 0.5, 1.5, 3.0, and 6.0 mg/ml) prior to cryopreservation. All males produced spermic ejaculates with >75% sperm motility. The mean ± SEM seminal volume, sperm concentration, percent motility, forward progression, and percent morphologically normal spermatozoa were 3.2 ± 0.3 ml, 1.2 ± 0.3 × 10 9 , 75.82 ± 2.7%, 3.2 ± 0.3 (0–5 scale; 5 = most progressive), and 57.12 ± 3.8%, respectively. More than 92% contained an intact acrosome. There was no effect of time or in vitro incubation on progression or acrosomal integrity on thawed samples ( P > 0.05). Spermatozoa pre-treated with 0.5 mg/ml CLC retained higher ( P < 0.05) motility post-thaw than aliquots treated with 0, 3.0, or 6.0 mg/ml of CLC. Spermatozoa pre-treated with 0.5, 1.5, or 3.0 mg/ml CLC exhibited greater viability than counterparts ( P < 0.05). Sperm kinetics including beat cross frequency (BCF), average path velocity (VAP), curvilinear velocity (VCL), and straight line velocity (VSL) did not differ among samples ( P > 0.05). Linearity (LIN) and straightness (STR) were different among samples after thawing. Results demonstrate treatment with CLC (0.5 mg/ml) protects Addra spermatozoa from cryo-damage. Reported advances will facilitate establishment of a frozen repository and support the genetic management of this critically endangered north African desert antelope. [ABSTRACT FROM AUTHOR]

The capacity to cryopreserve semen from captive cranes facilitates production of offspring from behaviorally incompatible or geographically separated pairs, and allows for long-term preservation of valuable genetic materials. The present study sought to develop effective cryopreservation protocols for whooping ( Grus americana ) and white-naped ( Grus vipio ) cranes, through examining the influences of two permeating (DMA and Me 2 SO) and one non-permeating (sucrose) cryoprotectants, as well as vitamin E on post-thaw sperm survival. In Study 1, ejaculates (whooping: n  = 10, white-naped: n = 8) were collected and cryopreserved in one of six cryo-diluents (crane extender with: DMA; DMA+0.1M sucrose; Me 2 SO; Me 2 SO+0.1M sucrose; 0.1M sucrose; 0.2M sucrose) using a two-step cooling method. Frozen samples were thawed and assessed for overall motility, motion characteristics, membrane integrity, morphology, and ability to bind to the inner perivitelline membrane (IPVM). In Study 2, whooping crane ejaculates ( n  = 17) were frozen in crane extender containing Me 2 SO alone or with vitamin E (5 μg/mL or 10 μg/mL). Frozen samples were thawed and assessed as in Study 1, except the binding assay. White-naped crane sperm were more tolerant to cryopreservation than whooping crane (15% vs 6% post-thawed motility). In both species, sperm cryopreserved in medium containing Me 2 SO alone displayed higher post thaw survival and ability to bind to IPVM than the other cryodiluent treatments. Vitamin E supplementation exerted no benefits to post thaw motility or membrane integrity. The findings demonstrated that there was species specificity in the susceptibility to cryopreservation. Nevertheless, Me 2 SO was a preferred cryoprotectant for sperm from both whooping and white-naped cranes. [ABSTRACT FROM AUTHOR]

To assist in genetic resource management and recovery efforts of the white-naped crane ( Antigone vipio ), we conducted two experiments to evaluate the effect of cooling condition, thawing rate, and cryoprotectant concentration on sperm survival post-thaw. Semen was collected from four mature males during breeding season (March and April) and evaluated for volume, sperm concentration, motility, and membrane integrity. In Experiment 1, ejaculates (n = 8) were diluted with Beltsville Poultry Semen Extender (BPSE) containing 10% dimethylsulfoxide (Me 2 SO) and frozen using either one (average cooling rate = 2.5 °C/min) or two step (average cooling rate = 7 and 9 °C/min, respectively) cooling method. The frozen samples were thawed using one of two thawing rates: 37 °C 30 s vs. 4 °C 1 min. In Experiment 2, samples were diluted with crane semen extender containing either 6% or 10% Me 2 SO, frozen using two-step method and then thawed at 37 °C for 30 s. Both cooling condition (two-step > one-step) and thawing rate (37 °C 30 s > 4 °C 1 min) impacted sperm motility, progression and kinetic characteristics ( P < 0.05), but did not ( P > 0.05) affect plasma membrane or acrosomal integrity. Concentration of Me 2 SO did not impact frozen-thaw survival. We conclude that white-naped crane sperm cryopreserved using a combination of two-step cooling and thawing at 37 °C 30 s was superior to other cooling and thawing combinations regarding to sustaining sperm motility with good motility kinetics. Findings represent the first steps towards the development of effective cryopreservation protocols and establishment of a genome resource bank for this threatened species. [ABSTRACT FROM AUTHOR]

Artificial insemination (AI) is a valuable tool for ex situ wildlife conservation, allowing the re-infusion and dissemination of genetic material, even after death of the donor. However, the application of AI to species conservation is still limited, due mainly to the poor survival of cryopreserved sperm. Recent work demonstrated that oviductal extracellular vesicles (oEVs) improved cat sperm motility and reduced premature acrosomal exocytosis. Here, we build on these findings by describing the protein content of dog and cat oEVs and investigating whether the incubation of cryopreserved red wolf and cheetah sperm with oEVs during thawing improves sperm function. Both red wolf and cheetah sperm thawed with dog and cat oEVs, respectively, had more intact acrosomes than the non-EV controls. Moreover, red wolf sperm thawed in the presence of dog oEVs better maintained sperm motility over time (>15%) though such an improvement was not observed in cheetah sperm. Our work demonstrates that dog and cat oEVs carry proteins important for sperm function and improve post-thaw motility and/or acrosome integrity of red wolf and cheetah sperm in vitro. The findings show how oEVs can be a valuable tool for improving the success of AI with cryopreserved sperm in threatened species. [ABSTRACT FROM AUTHOR]