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Refinement of Southern white rhinoceros semen sample processing protocols to optimize post-thaw sperm viability and integrity
Thesis (MScAgric)--Stellenbosch University, 2025.
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Stellenbosch : Stellenbosch University
2025
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| _version_ | 1867614092477333504 |
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| access_status_str | Open Access |
| author | Ferreira, Jana |
| author2 | Lambrechts, Helet |
| author_browse | Ferreira, Jana Lambrechts, Helet |
| author_facet | Lambrechts, Helet Ferreira, Jana |
| author_sort | Ferreira, Jana |
| collection | Thesis |
| dc_rights_str_mv | Stellenbosch University |
| description | Thesis (MScAgric)--Stellenbosch University, 2025. |
| format | Thesis |
| id | oai:scholar.sun.ac.za:10019.1/132351 |
| institution | Stellenbosch University (South Africa) |
| last_indexed | 2026-06-10T12:46:32.674Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from SUNScholar — Stellenbosch University Repository |
| publishDate | 2025 |
| publishDateRange | 2025 |
| publishDateSort | 2025 |
| publisher | Stellenbosch : Stellenbosch University |
| publisherStr | Stellenbosch : Stellenbosch University |
| record_format | dspace |
| source_str | SUNScholar — Stellenbosch University Repository |
| spelling | oai:scholar.sun.ac.za:10019.1/132351 Refinement of Southern white rhinoceros semen sample processing protocols to optimize post-thaw sperm viability and integrity Ferreira, Jana Lambrechts, Helet De la Rey, Morne Stellenbosch University. Faculty of AgriSciences. Dept. of Animal Sciences. White rhinoceros -- Breeding Semen Semen -- Evaluation White rhinoceros -- Reproduction -- Endocrine aspects White rhinoceros -- Behavior Endangered species White rhinoceros -- Conservation -- Management Semen -- Transportation UCTD Thesis (MScAgric)--Stellenbosch University, 2025. Ferreira, J. 2025. Refinement of Southern white rhinoceros semen sample processing protocols to optimize post-thaw sperm viability and integrity. Unpublished masters thesis. Stellenbosch: Stellenbosch University [online]. Available: https://scholar.sun.ac.za/items/da6df263-2f8b-40e9-bb11-a0baff9d1914 ENGLISH ABSTRACT: The aim of this study was to establish a baseline for future studies to characterize Southern white rhinoceros sperm samples. The effect of age, mating history, and social status of Southern white rhinoceros bulls on sperm parameters of samples collected using electroejaculation was described. The optimal duration of transport and equilibration, respectively, and the thawing protocol were determined with INRA- and Biladyl as extenders. The membrane-stabilizing capacity of trehalose during cooling and equilibration, to minimize morphological damage, was tested by respectively supplementing INRA Freeze® and Biladyl A with 50mM trehalose. Semen samples were collected from eight free-roaming Southern white rhinoceros bulls by means of electroejaculation. The fresh ejaculates were described by recording macroscopic parameters, sample volume, colour, viscosity, mass motility, progressive individual motility, and sperm concentration. The microscopic parameters measured included sperm viability, morphology, and acrosome integrity. The most optimal sperm samples were collected from a mature (older than 10 years) Southern white rhinoceros bull that was roaming with multiple cows and being able to exert dominance over subordinate bulls by practicing regular mating. Processing protocols using INRA was tested by transporting sperm samples at 4°C in INRA96® for 3, 4, 5, and 6 hours respectively, followed by equilibration periods of 0.5, 1, 1.5, and 2 hours in INRA Freeze®. The effect of two different thawing rates were evaluated by thawing the cryopreserved sperm samples at 35°C for 30 seconds, and at 45°C for 15 seconds, respectively. INRA Freeze® was supplemented with 50mM trehalose to determine its protective capacity during equilibration and cryopreservation. To quantify the effect of these treatments, the parameters recorded included mass motility, progressive motility, viability, morphology, and acrosome integrity of spermatozoa subjected to the respective treatments. The optimal processing protocol of Southern white rhinoceros sperm samples using INRA, includes transporting sperm samples at 4°C for 3 hours, followed by equilibration of 0.5 hours, without any trehalose supplementation. Thawing at 35°C for 30 seconds induced less morphological damage than thawing at 45°C for 15 seconds. The potential of Biladyl to be used as a transport and cryodiluent of Southern white rhinoceros spermatozoa, was evaluated by transporting sperm samples at 4°C in Biladyl A for 3, 4, 6, 8, 10 and 12 hours respectively; followed by equilibration for 1, 2, 4, and 6 hours in Biladyl B. Spermatozoa cryopreserved using Biladyl, were thawed at 35°C for 30 seconds, and at 45°C for 15 seconds, respectively. Biladyl A was supplemented with 50mM trehalose prior to equilibration to determine the membrane-stabilizing potential of trehalose. The same sperm parameters recorded for INRA post-thaw samples, were also recorded for the post-thaw Biladyl post-thaw samples. The optimal processing protocol of Southern white rhinoceros semen using Biladyl includes transporting sperm samples at 4°C for 3 hours, followed by 1 hour equilibration in Biladyl B. Trehalose did not have a membrane-stabilizing effect when supplemented to Biladyl A for fresh, equilibrated, or post-thaw spermatozoa. When cryopreserving spermatozoa in Biladyl B, sperm samples should be thawed at 35°C for 30 seconds to minimize morphological damage, rather than thawing at 45°C for 15 seconds. AFRIKAANSE OPSOMMING: Die doel van hierdie studie was om 'n basis te vestig vir toekomstige studies om Suidelike witrenoster sperm monsters te karakteriseer. Die effek van ouderdom, paringsgeskiedenis en sosiale status van Suidelike witrenosterbulle op sperm parameters van monsters wat met elektro-ejakulasie versamel is, is beskryf. Die optimale tyd vir onderskeidelik die vervoer en ekwilibrasie, en die ontdooiingsprotokol is bepaal met INRA- en Biladyl as mediums. Die membraan-stabiliserende kapasiteit van trehalose tydens verkoeling en ekwilibrasie, om morfologiese skade te verminder, is getoets deur onderskeidelik INRA Freeze® en Biladyl A met 50mM trehalose aan te vul. Semen monsters is van agt vrylopende Suidelike witrenosterbulle versamel deur middel van elektro- ejakulasie. Die vars ejakulate is beskryf in terme van makroskopiese parameters deur monstervolume, kleur, viskositeit, massa motiliteit, progressiewe individuele motiliteit en spermkonsentrasie, aan te teken. Die mikroskopiese parameters wat gemeet is, sluit in sperm lewensvatbaarheid, -morfologie en -akrosoomintegriteit. Die mees optimale spermmonsters is versamel van 'n volwasse (ouer as 10 jaar) Suidelike witrenosterbul wat met veelvuldige koeie rondgeloop het en oorheersing oor ondergeskikte bulle kon uitoefen deur gereelde paring te beoefen. Verwerkingsprotokolle met behulp van INRA is getoets deur spermmonsters by 4°C in INRA96® vir onderskeidelik 3, 4, 5 en 6 uur te vervoer, gevolg deur ekwilibrasie periodes van 0.5, 1, 1.5 en 2 uur in INRA Freeze®. Die effek van twee verskillende ontdooiingstempo's is geëvalueer deur die gevriesde sperm monsters onderskeidelik by 35°C vir 30 sekondes en by 45°C vir 15 sekondes te ontdooi. INRA Freeze® is aangevul met 50mM trehalose om die beskermende kapasiteit daarvan tydens ekwilibrasie en bevriesing te bepaal. Om die effek van hierdie behandelings te kwantifiseer, het die parameters wat aangeteken is, massa motiliteit, progressiewe beweeglikheid, lewensvatbaarheid, morfologie en akrosoomintegriteit van sperme wat aan die onderskeie behandelings onderwerp is, ingesluit. Die optimale verwerkingsprotokol van Suidelike witrenoster sperm monsters met die gebruik van INRA, sluit in die vervoer van sperm monsters by 4°C vir 3 uur, gevolg deur ekwilibrasie tydperk van 0.5 uur, sonder enige trehalose-aanvulling. Ontdooiing by 35°C vir 30 sekondes het minder morfologiese skade veroorsaak as ontdooiing by 45°C vir 15 sekondes. Die potensiaal van Biladyl om as 'n vervoer- en bevriesingsverdunningsmiddel van Suidelike witrenosters sperme gebruik te word, is geëvalueer deur spermmonsters by 4°C in Biladyl A vir onderskeidelik 3, 4, 6, 8, 10 en 12 uur te vervoer; gevolg deur ekwilibrasie vir 1, 2, 4 en 6 uur in Biladyl B. Sperme wat met Biladyl gevries is, is onderskeidelik by 35°C vir 30 sekondes en by 45°C vir 15 sekondes ontdooi. Biladyl A is aangevul met 50mM trehalose voor ekwilibrasie om die membraan- stabiliserende potensiaal van trehalose te bepaal. Dieselfde spermparameters wat vir INRA-monsters na ontdooiing aangeteken is, is ook aangeteken vir die monsters na die ontdooiing van Biladyl na die ontdooiing. Die optimale verwerkingsprotokol van Suidelike witrenoster sperme met behulp van Biladyl sluit in die vervoer van sperm monsters by 4°C vir 3 uur, gevolg deur 1 uur ekwilibrasie in Biladyl B. Trehalose het nie 'n membraan- stabiliserende effek gehad toe dit aangevul is met Biladyl A vir vars, geëkwilibreerde of bevrore-ontdooide sperme nie. Wanneer sperme in Biladyl B gevries word, moet spermmonsters vir 30 sekondes by 35 ° C ontdooi word om morfologiese skade te verminder, eerder as om vir 15 sekondes by 45 ° C te ontdooi. Masters 2025-06-04T12:39:17Z 2025-06-04T12:39:17Z 2025-03 Thesis https://scholar.sun.ac.za/handle/10019.1/132351 Stellenbosch University xix, 246, 6 unnumbered pages : illustrations application/pdf Stellenbosch : Stellenbosch University |
| spellingShingle | White rhinoceros -- Breeding Semen Semen -- Evaluation White rhinoceros -- Reproduction -- Endocrine aspects White rhinoceros -- Behavior Endangered species White rhinoceros -- Conservation -- Management Semen -- Transportation UCTD Ferreira, Jana Refinement of Southern white rhinoceros semen sample processing protocols to optimize post-thaw sperm viability and integrity |
| title | Refinement of Southern white rhinoceros semen sample processing protocols to optimize post-thaw sperm viability and integrity |
| title_full | Refinement of Southern white rhinoceros semen sample processing protocols to optimize post-thaw sperm viability and integrity |
| title_fullStr | Refinement of Southern white rhinoceros semen sample processing protocols to optimize post-thaw sperm viability and integrity |
| title_full_unstemmed | Refinement of Southern white rhinoceros semen sample processing protocols to optimize post-thaw sperm viability and integrity |
| title_short | Refinement of Southern white rhinoceros semen sample processing protocols to optimize post-thaw sperm viability and integrity |
| title_sort | refinement of southern white rhinoceros semen sample processing protocols to optimize post thaw sperm viability and integrity |
| topic | White rhinoceros -- Breeding Semen Semen -- Evaluation White rhinoceros -- Reproduction -- Endocrine aspects White rhinoceros -- Behavior Endangered species White rhinoceros -- Conservation -- Management Semen -- Transportation UCTD |
| url | https://scholar.sun.ac.za/handle/10019.1/132351 |
| work_keys_str_mv | AT ferreirajana refinementofsouthernwhiterhinocerossemensampleprocessingprotocolstooptimizepostthawspermviabilityandintegrity |