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RESEARCH ARTICLE
Contents Vol 20(7)

Semen collection, characterisation and artificial insemination in the beluga (Delphinapterus leucas) using liquid-stored spermatozoa

J. K. O’Brien A B D , K. J. Steinman A , T. Schmitt C and T. R. Robeck A
+ Author Affiliations
- Author Affiliations

A SeaWorld and Busch Gardens Reproductive Research Center, 2595 Ingraham St, San Diego, CA 92109, USA.

B Centre for Advanced Technologies in Animal Genetics and Reproduction, Faculty of Veterinary Science, University of Sydney, NSW 2006, Australia.

C SeaWorld California, 500 SeaWorld Dr, San Diego, CA 92109, USA.

D Corresponding author. Email: justineo@vetsci.usyd.edu.au

Reproduction, Fertility and Development 20(7) 770-783 https://doi.org/10.1071/RD08031
Submitted: 17 February 2008  Accepted: 4 June 2008   Published: 1 August 2008

Abstract

Ejaculates were collected from a beluga (Delphinapterus leucas) to gain an understanding of sperm biology and develop a short-term sperm preservation method for use in artificial insemination (AI). Ejaculate parameters and biochemistry, semen production and serum testosterone concentrations of an adult male were characterised for 21 months. Sperm viability, acrosome integrity and morphology did not change (P > 0.05) but ejaculate volume, sperm concentration and total spermatozoa per ejaculate were higher (P < 0.05) from January to June than from July to December. Peak testosterone concentrations (P < 0.05) were observed from October to April (8.0 ± 1.6 ng mL–1). The effects of hyaluronic acid (HA), antioxidants, storage temperature and time on in vitro sperm characteristics were examined. Motility parameters and viability were improved (P < 0.05) when semen was stored at 5°C compared with 21°C. During the first 24 h of storage sperm agglutination was absent only at 5°C in the presence of HA. A nulliparous 28-year-old female was inseminated endoscopically with liquid-stored semen. A pregnancy and birth of a calf was achieved following AI for the first time in this species, thereby validating both the AI technique and the fertility of beluga spermatozoa after chilled storage in a specialised diluent.

Additional keywords: cetacean, hyaluronan, intrauterine insemination, semen storage.


Acknowledgements

Animal care, animal training, curatorial and veterinary staff at SeaWorld California and Texas are thanked for assistance with animal handling and sample collection. Mrs Mitzi Synnott and Mrs Nicole Grovhoug are particularly thanked for the initial training which led to successful collection of beluga semen, and we are grateful to all Wild Arctic Staff for ongoing collections and assistance with AI procedures. Animal Care Laboratory staff (SeaWorld California) including Ms Whitney Coleman, Mrs Leigh Ann Smith, Mrs Pam Thomas and Mrs Melinda Tucker are thanked for technical support, as is Ms Sherry Dickerson, Dr Steve Osborn and Mr Chris White (SeaWorld Texas) for sample collection and preparation. Dr Robert Anderson (Obstetrical and Gynaecology Research, Rush Medical Center, Chicago) performed the seminal plasma fructose analyses, for which we are grateful. Mr Brad Andrews is also thanked for invaluable support of this research. This project was funded by SeaWorld Corporation and is a SeaWorld Technical Contribution no. 2008–01-T.


References

Aires, V. A. , Hinsch, K. D. , Muller-Schlosser, F. , Bogner, K. , Muller-Schlosser, S. , and Hinsch, E. (2003). In vitro and in vivo comparison of egg yolk-based and soybean lecithin-based extenders for cryopreservation of bovine semen. Theriogenology 60, 269–279.
| Crossref | GoogleScholarGoogle Scholar | PubMed | Andrews B. (2007). ‘Beluga North American Studbook.’ Marine Mammal Taxon Advisory Group, American Zoo and Aquarium Association (AZA). (AZA: Silver Spring: MD, USA.)

Blom, E. (1973). The ultrastructure of some characteristic sperm defects and a proposal of the bull spermiogram. Nord. Vet. Med. 25, 383–387.
| PubMed | Calle P. P., Cook R. A., McClave C., and Palma S. (1993). Circulating gestational progesterone and estradiol concentrations, parturition, and placental descriptions of two beluga whales (Delphinapterus leucas). In ‘Proceedings of the International Association of Aquatic Animal Medicine’. Volume 24, p. 61. [Abstract]

Calle P. P., Cook R. A., Robeck T. R., Young S. J. F., and Jones M. H. (1996). Circulating gestational progesterone and oestradiol concentrations in Beluga whales (Delphinapterus leucas). In ‘Proceedings of the American Association of Zoo Vets’. pp. 340–342. [Abstract]

Calle P. P., Monfort S. L., Dunn J. L., Jensen E., Boehm R., Young S. J. F., and Robeck  T. R. (2000). Seasonal testosterone secretion in male white whales (Delphinapterus leucas). In ‘Joint Proceedings of American Association of Zoo Veterinarians and International Association of Aquatic Animal Medicine, New Orleans, Louisiana, September 17–21’. pp. 49–50. [Abstract]

Eppig, J. J. (1979). FSH stimulates hyaluronic acid synthesis by oocyte–cumulus cell complexes from mouse preovulatory follicles. Nature 281, 483–484.
| Crossref | GoogleScholarGoogle Scholar | PubMed | Holt W. V., Abaigar T., Watson P. F., and Wildt D. E. (2003). Genetic resource banks for species conservation. In ‘Reproductive Science and Integrated Conservation’. (Eds W. V. Holt, A. R. Pickard, J. C. Rodger and D. E. Wildt.) pp. 267–280. (Cambridge University Press: Cambridge.)

Holt, W. V. , O’Brien, J. K. , and Abaigar, T. (2007). Applications and interpretation of computer-assisted sperm analyses and sperm sorting methods in assisted breeding and comparative research. Reprod. Fertil. Dev. 19, 709–718.
| Crossref | GoogleScholarGoogle Scholar | PubMed | IUCN (1996) ‘The Red List of Threatened Animals.’ (IUCN Publications Service: Cambridge.)

IUCN (2006) ‘The Red List of Threatened Animals.’ (IUCN Publications Service: Cambridge.)

Jakab, A. , Sakkas, D. , Delpiano, E. , Cayli, S. , Kovanci, E. , Ward, D. , Revelli, A. , and Huszar, G. (2005). Intracytoplasmic sperm injection: a novel selection method for sperm with normal frequency of chromosomal aneuploidies. Fertil. Steril. 84, 1665–1673.
| Crossref | GoogleScholarGoogle Scholar | PubMed | Kenney R. M., Bergman R. V., Cooper W. L., and Morse G. W. (1975). Minimal contamination techniques for breeding mares: techniques and preliminary findings. In ‘Proceedings of the 21st Annual Convention of the American Association of Equine Practitioners’. pp. 327–336.

Kornovski, B. S. , McCoshen, J. M. , Kredentser, J. , and Turley, E. (1994). The regulation of sperm motility by a novel hyaluronan receptor. Fertil. Steril. 61, 935–940.
| PubMed | Mann T. (1964). ‘The Biochemistry of Semen and of the Male Reproductive Tract.’ (Methuen and Co.: London.)

Maxwell, W. M. C. , and Stojanov, T. (1996). Liquid storage of ram semen in the absence or presence of some antioxidants. Reprod. Fertil. Dev. 8, 1013–1020.
| Crossref | GoogleScholarGoogle Scholar | PubMed | NAMMCO (2000). Report of the NAMMCO scientific committee working group on the population status of Beluga and Narwhal in the North Atlantic. Annual Report of the North Atlantic Marine Mammal Commission. Tromsø, Norway 1999. pp. 153–188.

O’Brien J. K. and Robeck T. R. (2005). Assisted reproductive technology in the bottlenose dolphin (Tursiops truncatus): male. In ‘Proceedings of the Bottlenose Dolphin Reproductive Workshop. European Association of Aquatic Mammals, Paris’.

O’Brien, J. K. , and Robeck, T. R. (2006). Development of sperm sexing and associated assisted reproductive technology for sex pre-selection of captive bottlenose dolphins (Tursiops truncatus). Reprod. Fertil. Dev. 18, 319–329.
| Crossref | GoogleScholarGoogle Scholar | PubMed | Ognetev G. N. (1981). Studies on the ecology and taxonomy of the white whale (Delphinapterus leucas Pall., 1776) inhabiting the Soviet Arctic. Report on the International Whaling Commission 31, pp. 515–520.

Pagl, R. , Aurich, J. E. , Müller-Schlösser, F. , Kankofer, M. , and Aurich, C. (2006). Comparison of an extender containing defined milk protein fractions with a skim milk-based extender for storage of equine semen at 5 degrees C. Theriogenology 66, 1115–1122.
| Crossref | GoogleScholarGoogle Scholar | PubMed | Robeck T. R., and O’Brien J. K. (2005). ‘Development and Application of Assisted Reproductive Technologies in Cetaceans. World Association of Zoos and Aquariums. Marine Conservation Issues. Number 7.’ (Ed. P. Dollinger.) pp. 8–10. (WAZA Liebefeld-Bern: Switzerland.)

Robeck T. R., Greenwell M., Boehm J. R., Yoshioka M., Tobayama T., Steinman K. J., and Monfort S. T. (2003). Artificial insemination using frozen–thawed semen in the Pacific white-sided dolphin (Lagenorhynchus obliquidens). In ‘Proceedings for the 34th Annual Conference of the International Association for Aquatic Animal Medicine, Waikoloa, Hawaii’. pp. 50–52. [Abstract]

Robeck, T. R. , Steinman, K. J. , Gearhart, S. , Reidarson, T. R. , McBain, J. F. , and Monfort, S. L. (2004). Reproductive physiology and development of artificial insemination technology in killer whales (Orcinus orca). Biol. Reprod. 71, 650–660.
| Crossref | GoogleScholarGoogle Scholar | PubMed | Rodriguez-Martinez H., Larsson B., Pertoft H., and Kjelle L. (1998). GAGs and spermatozoon competence in vivo and in vitro. In ‘Gametes: Development and Function’. (Eds A. Lauria, F. Gandol, G. Enne and L. Gianaroli.) pp. 239–274. (Serono Symposia: Italy.)

Rodriguez-Martinez, H. , Tienthai, P. , Suzuki, K. , Funahashi, H. , Ekwall, H. , and Johannisson, A. (2001). Involvement of oviduct in sperm capacitacion and oocyte development in pigs. Reprod. Suppl. 58, 129–145.
| PubMed | Steinman K. J., O’Brien J. K., and Robeck T. R. (2007). Characterisation of reproductive cycles and the development of an ovulation induction method in the beluga (Delphinapterus leucus). In ‘Proceedings for the 38th Annual Conference of the International Association for Aquatic Animal Medicine, Bahamas’. 38, pp. 162–164. [Abstract]

Tienthai, P. , Kjellén, L. , Pertoft, H. , Suzuki, K. , and Rodriguez-Martinez, H. (2000). Localisation and quantitation of hyaluronan and sulfated glycosaminoglycans in the tissues and intraluminal fluid of the pig oviduct. Reprod. Fertil. Dev. 12, 173–182.
| Crossref | GoogleScholarGoogle Scholar | PubMed | Vishwanath R., Munday R., and Shannon P. (1994). Field trials on the efficacy of Desferal – a chelating agent – in extending fertilising ability of bull sperm diluted in ambient temperature diluent. In ‘Proceedings of the New Zealand Embryo Transfer Workshop’. pp. 72–73. (Immuno-Chemical Products: Auckland.)