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Improving Assisted Reproductive Technologies in the Endangered Black-Footed Ferret: Artificial Insemination and Sperm Cryopreservation

Strickler, Tara Leigh

Abstract Details

2010, Master of Science, Ohio State University, Animal Sciences.

The black-footed ferret (Mustela nigripes) is an endangered mustelid native to the grasslands of the North American western Great Plains. Recovery efforts for the species warranted the capture of the last remaining 18 black-footed ferrets for ex situ propagation. Despite vigorous attempts to equalize genetic representation, reproductive inefficiency has been documented. Black-footed ferret spermatozoa suffer significant cryodamage and a decline in percentages of normal sperm in fresh ejaculates has been observed. The decline in reproductive fitness has been attributed to the low number of founders (n = 7) that comprise the current population.

The first aim of this project was to improve genetic diversity in the black-footed ferret population by using cryopreserved spermatozoa from a genome resource bank. Black-footed ferret females (n = 4) chosen for laparoscopic artificial insemination were evaluated for estrus. Ovulation was induced with luteinizing hormone. Sires from the genome resource bank were selected based on genetic representation. Two of four (50%) females inseminated became pregnant and produced kits (mean litter size, 1.0).

The second aim of this project was to investigate three commercially available density gradient products (PureSperm, ISolate, Enhance-S Plus) to develop a technique to improve the quality of black-footed ferret frozen-thawed sperm fractions for artificial insemination. Domestic ferret ejaculates (used as a model for this study) were collected via electroejaculation and frozen according to standard procedures. Frozen- thawed spermatozoa were divided into 4 groups and either received a simple washing (control) or were subjected to one of three gradient treatments. Results indicated that PureSperm filtration produced the best sperm fractions with the highest percentages of structurally normal and motile sperm.

The third aim of this project was to investigate cholesterol-loaded cyclodextrin (CLC) technology as a means to develop a pre-freeze treatment to improve sperm cryosurvival. Domestic ferret ejaculates (chosen as the model for this study) were collected via electroejaculation, evaluated, and were divided among three groups: 1) control, 2) low dose CLC (1.5 mg CLC/ 120 x106 sperm/ml), and 3) high dose CLC (3.0 mg CLC/ 120 x106 sperm/ml). After receiving treatment, ejaculates were subjected to standard cryopreservation methods. Results indicated significant improvement in acrosomal integrity due to treatment, but there was no significant difference in this regard between low and high doses. CLC treatment appeared to have no significant effect on sperm motility or forward progression.

Offspring that resulted from this study (n = 2) are currently considered the most genetically valuable animals in the black-footed ferret population. However, further studies are warranted in these individuals and subsequent generations to determine whether increased genetic fitness improves sperm viability in this species. Moreover, future artificial inseminations using cryopreserved sperm from the black-footed ferret genome resource bank may benefit from PureSperm density gradient processing since application of this product enhanced cryopreserved sperm fractions in the domestic ferret. Although use of CLC as a pre-freeze treatment did not appear to affect sperm longevity, further investigations are warranted to confirm the degree of membrane manipulation that occurs with CLC application prior to cryopreservation, which is shown to improve acrosomal integrity in domestic ferret frozen-thawed sperm.

Joseph Ottobre, PhD (Advisor)
Douglas Danforth, PhD (Committee Member)
Mandi Vick, PhD (Committee Member)
136 p.

Recommended Citations

Citations

  • Strickler, T. L. (2010). Improving Assisted Reproductive Technologies in the Endangered Black-Footed Ferret: Artificial Insemination and Sperm Cryopreservation [Master's thesis, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1267057806

    APA Style (7th edition)

  • Strickler, Tara. Improving Assisted Reproductive Technologies in the Endangered Black-Footed Ferret: Artificial Insemination and Sperm Cryopreservation. 2010. Ohio State University, Master's thesis. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1267057806.

    MLA Style (8th edition)

  • Strickler, Tara. "Improving Assisted Reproductive Technologies in the Endangered Black-Footed Ferret: Artificial Insemination and Sperm Cryopreservation." Master's thesis, Ohio State University, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=osu1267057806

    Chicago Manual of Style (17th edition)