Thesis
Effectiveness of vitrification and vapor immersion for honey bee semen cryopreservation
Washington State University
Master of Science (MS), Washington State University
05/2021
DOI:
https://doi.org/10.7273/000004282
Handle:
https://hdl.handle.net/2376/124650
Abstract
Semen cryopreservation is a valuable conservation tool and is often used in livestock species to accelerate artificial selection of desirable traits. Recently, semen cryopreservation has been successfully introduced to honey bees, bolstering trait selection for breeders and aiding conservation efforts for threatened bee populations. Current cryopreservation methods use slow-programmable freezing to achieve long-term storage of honey bee germplasm. However, this method is costly and time consuming, making it less accessible to breeders and researchers. Additionally, the success of this method is inconsistent, producing inseminated queens with variable fertilization success, prompting the need for an alternative freezing method. Here, I tested vitrification and vapor immersion as two potential alternatives to slow-programmable freezing. With each freezing method, I frozen semen in either a CryoLock or microdialysis tube using either 20, 40, or 60% dimethyl sulfoxide as a cryoprotectant. I found that freezing honey bee semen in microdialysis tubes with vapor immersion and 20% dimethyl sulfoxide was the only combination of treatments to produce similar post-thaw sperm quality to semen frozen with slow-programmable freezing. I then instrumentally inseminated queens with this freezing method (n = 8) and compared the percentage of fertilized offspring to that produced by queens inseminated with semen frozen with standard slow-programmable freezing (n = 8), semen frozen at -1.5°C/min (half speed, n = 8), and non-frozen semen (n = 6). Several queens died prior to egg laying resulting in smaller sample sizes (n = 4, 4, 3, 2, respectively). The percentage of fertilized offspring was highest with the standard slow-programmable freezing method, but all treatments were statistically similar. No definitive conclusion can be made regarding the success of vapor immersion from the results of the queen insemination experiment; however, using vapor immersion to freeze honey bee semen in microdialysis tubes with 20% dimethyl sulfoxide did produce appreciable post-thaw sperm quality. With proper optimization of the dimethyl sulfoxide exposure and freezing rate, this freezing method may be used to provide an accessible honey bee semen cryopreservation method to slow programmable freezing.
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Details
- Title
- Effectiveness of vitrification and vapor immersion for honey bee semen cryopreservation
- Creators
- Connor Anthony Auth
- Contributors
- BRANDON KINGSLEY HOPKINS (Advisor) - Washington State University, Entomology, Department of
- Awarding Institution
- Washington State University
- Academic Unit
- Entomology, Department of
- Theses and Dissertations
- Master of Science (MS), Washington State University
- Publisher
- Washington State University
- Identifiers
- 99900896412501842
- Language
- English
- Resource Type
- Thesis