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Reproduction, Fertility and Development Reproduction, Fertility and Development Society
Vertebrate reproductive science and technology
RESEARCH ARTICLE

Semen collection, ejaculate characteristics and in vitro manipulation of spermatozoa from six species of captive flying-fox (Pteropus spp.)

D. F. Melville A , E. G. Crichton A and S. D. Johnston A B
+ Author Affiliations
- Author Affiliations

A Wildlife Biology Unit, School of Agriculture and Food Science, University of Queensland, Gatton, Qld 4343, Australia.

B Corresponding author. Email: s.johnston1@uq.edu.au

Reproduction, Fertility and Development 27(8) 1233-1241 https://doi.org/10.1071/RD13391
Submitted: 18 November 2013  Accepted: 8 May 2014   Published: 8 July 2014

Abstract

Seminal characteristics are described in six Pteropus species including the critically endangered P. rodricensis. Spermic ejaculates (~40 μL) were collected using electro-ejaculation on 406 of 413 attempts. All flying-fox species had mean percentages of acrosome- and plasma-membrane (PM)-intact spermatozoa of >66% and >73%, respectively; the predominant sperm abnormalities found across all species were damaged, folded or missing acrosomes, bent midpieces and coiled tails. Seminal pH ranged from a low of 7.5 in P. giganteus to a high of 8.2 in P. alecto with the other species in between. Electro-ejaculates recovered in short succession from P. alecto revealed no differences in sperm quality, allowing spermatozoa to be utilised for multi-treatment experiments that evaluated the effects of transportation, incubation temperature and in vitro physico-chemical environments on acrosome and PM integrity. Pteropus alecto spermatozoa were successfully held at ~27°C and 37°C for up to 6 h before a reduction in PM integrity (P = 0.003) was observed. Acrosome and PM integrity decreased (P < 0.000) when P. alecto spermatozoa were incubated at 37°C for 30 min in a Tris–citrate buffer of pH 9.0 but remained stable at pH 5.0 to 8.0. Pteropus alecto mean (± s.e.m.) seminal osmolality was 307.0 ± 2.5 mOsm kg–1; nevertheless, spermatozoa were tolerant of media ranging from 160 to 1190 mOsm kg–1 but exposure to media of ≤160 mOsm kg–1 resulted in increased acrosome damage (P < 0.000).

Additional keywords: acrosome, Chiroptera, fruit bat, osmolality, pH, sperm preservation.


References

Barth, A. D. (1995). Evaluation of frozen bovine semen by the veterinary practitioner. In ‘Proceedings of Bovine Short Course’. pp. 105–110. (American College of Theriogenologists and Society for Theriogenology: Hastings, USA.)

Barth, A. D., and Oko, R. J. (1989). ‘Abnormal Morphology of Bovine Spermatozoa’. (Iowa State University Press: Ames, IA, USA.)

Crichton, E. G. (2000). Sperm storage and fertilization. In ‘Reproductive Biology of Bats’. (Eds E. G. Crichton and P. J. Krutzsch.) pp. 295–320. (Academic Press: San Diego.)

Crichton, E. G., Hinton, B. T., Pallone, T. L., and Hammerstedt, R. H. (1994). Hyperosmolality and sperm storage in hibernating bats: prolongation of sperm life by dehydration. Am. J. Physiol. 267, R1363–R1370.
| 1:CAS:528:DyaK2MXitVKqsbk%3D&md5=38803b12ae3159f6065d00c118b09baaCAS | 7977866PubMed |

de Jong, C. E., Jonsson, N. N., Field, H., Smith, C., Crichton, E. G., and Johnston, S. D. (2005). Collection, seminal characteristics and chilled storage of spermatozoa from three species of free-range flying fox (Pteropus spp.). Theriogenology 64, 1072–1089.
Collection, seminal characteristics and chilled storage of spermatozoa from three species of free-range flying fox (Pteropus spp.).Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD2MvlvF2mtw%3D%3D&md5=7958ab353753bc61916849773a23999bCAS | 15913752PubMed |

Forman, G. L., Smith, J. D., and Hood, C. S. (1989). Exceptional size and unusual morphology of spermatozoa in Noctilio albiventris (Noctilionidae). J. Mammal. 70, 179–184.
Exceptional size and unusual morphology of spermatozoa in Noctilio albiventris (Noctilionidae).Crossref | GoogleScholarGoogle Scholar |

Hall, L., and Richards, G. 2000. ‘Flying Foxes: Fruit and Blossom Bats of Australia’. (University of New South Wales Press Ltd.: Sydney.)

Howard, J. G., Wildt, D. E., Chakraborty, P. K., and Bush, M. (1983). Reproductive traits including seasonal observations on semen quality and serum hormone concentrations in the Dorcas gazelle. Theriogenology 20, 221–234.
Reproductive traits including seasonal observations on semen quality and serum hormone concentrations in the Dorcas gazelle.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaL3sXmtVKnsr8%3D&md5=8c6578a60f9d344b2b75c79c82bd96deCAS |

Hutchins, M., Kleiman, D. G., Geist, V., and McDade, M. C. (2003) ‘Grzimek’s Animal Life Encyclopedia. Vol. 12–16, Mammals I–V’, 2nd edn. (Gale Group: Farmington Hills, MI, USA.)

IUCN 2013. IUCN Red List of Threatened Species. Version 2013.1. Available at www.iucnredlist.org [Verified 13 November 2013].

Jackson, S. (2003). ‘Australian Mammal: Biology and Captive Management’. (CSIRO Publishing: Collingwood, Vic., Australia).

Jonsson, N. N., Johnston, S. D., Field, H., de Jong, C., and Smith, C. (2004). Field anaesthesia of three Australian species of flying fox. Vet. Rec. 154, 664.
Field anaesthesia of three Australian species of flying fox.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD2c3pslWntA%3D%3D&md5=182a7f59831c6ce72cd575dd092837c8CAS | 15198316PubMed |

Koehler, J. K. (1977). Fine structure of spermatozoa of the Asiatic musk shrew, Suncus murinus. Am. J. Anat. 149, 135–151.
Fine structure of spermatozoa of the Asiatic musk shrew, Suncus murinus.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DyaE2s3hvFSjtw%3D%3D&md5=e5fb1aee8ee4d69d890c3ab5f08e72adCAS | 879041PubMed |

Kunz, T. H., and Jones, D. P. (2000). Pteropus vampyrus. Mamm. Species 642, 1–6.
Pteropus vampyrus.Crossref | GoogleScholarGoogle Scholar |

Kusunoki, H., Okuda, K., Ueda, K., Ooe, T., Hayashi, T., Ito, S., Kawakami, S., Saitou, E., Fukuoka, T., Hase, T., Sato, T., and Doi, O. (2006). Evaluation of semen quality of captive-living cheetahs (Acinonyx jubatus) in Japan. Jpn. J. Zool. Wildl. Med. 11, 25–30.

Melville, D. F., Crichton, E. G., Paterson-Wimberley, T., and Johnston, S. D. (2008). Collection of semen by manual stimulation and ejaculate characteristics of the Black flying fox (Pteropus alecto). Zoo Biol. 27, 159–164.
Collection of semen by manual stimulation and ejaculate characteristics of the Black flying fox (Pteropus alecto).Crossref | GoogleScholarGoogle Scholar | 19360614PubMed |

Melville, D. F., O’Brien, G. M., and Johnston, S. D. (2011). The novel reproductive biology of the female flying fox and its implications for the successful development of an artificial insemination programme. In ‘The Biology and Conservation of Australasian Bats’. (Eds B. Law, P. Eby, D. Lunney and L. Lumsden.) pp. 128–135. (Royal Zoological Society of NSW: Mosman, NSW, Australia.)

Melville, D. F., O’Brien, G. M., Crichton, E. G., Theilemann, P., McKinnon, A., and Johnston, S. D. (2012a). Reproductive seasonality and the effect of the GnRH agonist deslorelin as a contraceptive in captive male Black flying foxes (Pteropus alecto). Theriogenology 77, 652–661.
Reproductive seasonality and the effect of the GnRH agonist deslorelin as a contraceptive in captive male Black flying foxes (Pteropus alecto).Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC38XptVakuw%3D%3D&md5=1729d805fee9bb34846075a4eee9a696CAS | 22115809PubMed |

Melville, D. F., Johnston, S. D., and Miller, R. R. (2012b). Flying-fox (Pteropus spp.) sperm-membrane fatty-acid composition, its relationship to cold-shock injury and implications for cryopreservation success. Cryobiology 65, 224–229.
Flying-fox (Pteropus spp.) sperm-membrane fatty-acid composition, its relationship to cold-shock injury and implications for cryopreservation success.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC38XhtV2nu7fN&md5=a1582c11bc7f1ce7d66fe9ce9700c305CAS | 22771758PubMed |

Morais, R. N., Mucciolo, R. G., Gomes, M. L. F., Lacerda, O., Moraes, W., Graham, L. H., Swanson, W. F., and Brown, J. L. (2002). Seasonal analysis of semen characteristics, serum testosterone and faecal androgens in the ocelot (Leopardus pardalis), margay (L. wiedii) and tigrina (L. tigrinus). Theriogenology 57, 2027–2041.
Seasonal analysis of semen characteristics, serum testosterone and faecal androgens in the ocelot (Leopardus pardalis), margay (L. wiedii) and tigrina (L. tigrinus).Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38XltFansLg%3D&md5=ebdbafa7072d94aa9b73fc7cd41c41b1CAS | 12066863PubMed |

Moreland, R. B., Richardson, M., Lamberski, N., and Long, J. (2001). Characterising the reproductive physiology of the male Southern black howler monkey, Alouatta caraya. J. Androl. 22, 395–403.
| 1:STN:280:DC%2BD3MrgsVOltQ%3D%3D&md5=6cb9227b483e115092d4e8e326ec2c3eCAS | 11330639PubMed |

Mortimer, D., and Mortimer, S. T. (2013). Computer-aided sperm analysis (CASA) of sperm motility and hyperactivation. Methods Mol. Biol. 927, 77–87.
Computer-aided sperm analysis (CASA) of sperm motility and hyperactivation.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3sXit1Snu74%3D&md5=959a257431346c122a7cebb16cf8a428CAS | 22992905PubMed |

Pfeiffer, B., and Mayer, F. (2013). Spermatogenesis, sperm storage and reproductive timing in bats. J. Zool. (Lond.) 289, 77–85.
Spermatogenesis, sperm storage and reproductive timing in bats.Crossref | GoogleScholarGoogle Scholar |

Phillips, D. M., Rasweiler, J. J., and Muradakli, F. (1997). Giant, accordioned sperm acrosomes of the Greater bulldog bat, Noctilio leporinus. Mol. Reprod. Dev. 48, 90–94.
Giant, accordioned sperm acrosomes of the Greater bulldog bat, Noctilio leporinus.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK2sXlt1Sis7s%3D&md5=5d84debd2cb90eb9c58337f213b9e09dCAS | 9266765PubMed |

Setchell, B. P. (1991). Male reproductive organs and semen. In ‘Reproduction in Domestic Animals’, 4th edn. (Ed. P. T. Cupps.) pp. 221–249. (Academic Press Inc.: Sydney.)

Songsasen, N., Yu, I., Murton, S., Paccamonti, D. L., Eilts, B. E., Godke, R. A., and Leibo, S. P. (2002). Osmotic sensitivity of canine spermatozoa. Cryobiology 44, 79–90.
Osmotic sensitivity of canine spermatozoa.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38XltlSqtrc%3D&md5=f14290eb132e62180d45167d7058f93cCAS | 12061850PubMed |

Thiangtum, K., Swanson, W. F., Howard, J., Tunwattana, W., Tongthainan, D., Wichasilpa, W., Patumrattanathan, P., and Pinyopoommintr, T. (2006). Assessment of basic seminal characteristics, sperm cryopreservation and heterologous in vitro fertilisation in the fishing cat (Prionailurus viverrinus). Reprod. Fertil. Dev. 18, 373–382.
Assessment of basic seminal characteristics, sperm cryopreservation and heterologous in vitro fertilisation in the fishing cat (Prionailurus viverrinus).Crossref | GoogleScholarGoogle Scholar | 16554013PubMed |

Valle, R. R., Guimaraes, M. A. B. V., Muniz, J. A. P. C., Barnabe, R. C., and Vale, W. G. (2004). Collection and evaluation of semen from captive howler monkeys (Alouatta caraya). Theriogenology 62, 131–138.
Collection and evaluation of semen from captive howler monkeys (Alouatta caraya).Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD2c3mtFSqsg%3D%3D&md5=9eb577ddf64cdf7c0f0b529fbdfddd25CAS | 15159108PubMed |

Watson, P. F., and Fuller, B. J. (2001). Principles of cryopreservation of gametes and embryos. In ‘Cryobanking the Genetic Resource: Wildlife Conservation for the Future?’. (Eds P. F. Watson and W. V. Holt.) pp. 23–46. (Taylor and Francis: London.)

Wildt, D. E. (1997). Male reproduction: assessment, management and control of fertility. In ‘Wild Mammals in Captivity: Principles and Techniques’. (Eds D. G. Kleiman, M. E. Allen, K. V. Thompson and S. Lumpkin.) pp. 429–550. (The University of Chicago Press: Chicago.)

Zee, Y. P., Holt, W. V., Nicolson, V., Pyne, M., and Johnston, S. D. (2009). Individual variability in post-thaw sperm survival in a captive koala population. Cryobiology 59, 69–74.
Individual variability in post-thaw sperm survival in a captive koala population.Crossref | GoogleScholarGoogle Scholar | 19426724PubMed |