A Case Report Concerning Male Gametes Rescued from a Siamese Eld’s Deer (<i>Rucervus eldii siamensis</i>): Post-Thawed Testicular and Epididymal Sperm Quality and Heterologous Zona Pellucida Binding Ability

Abstract

ABSTRACT. In the present study, the quality of frozen-thawed epididymal and testicular sperm recovered from a Siamese Eld's deer was examined. The epididymal sperm quality was assessed in fresh, cold-stored at 4°C and frozen-thawed samples. Zona binding ability of the frozen-thawed epididymal samples with Burmese Eld's deer oocytes was also evaluated. Testicular sperm extracted from tissues frozen at −80 or −196°C for one month were examined for membrane and DNA integrity. Epididymal sperm retained their quality for up to 24 hr of cold storage at 4°C. The percentages of sperm motility, intact membrane, intact acrosome and intact DNA were 30, 46.5, 27 and 89.5% in the frozen and thawed epididymal sperm, and the average ability to bind with oocytes was 92.5 ± 64 sperm/oocytes. Around 70% of the sperm extracted from testicular tissues cryopreserved at −196 and −80°C for one month showed an intact membrane. In conclusion, epididymal and testicular sperm survived for more than 13 hr post-mortem. Furthermore, cold storage at 4°C and cryopreservation at −196 and −80°C maintain the quality of epididymal and testicular sperm. This study represents a model for male gamete rescue in endangered Eld's deer. KEY WORDS: Siamese Eld's deer, sperm collection post mortem, sperm cryopreservation.doi: 10.1292/jvms.11-0491; J. Vet. Med. Sci. 75(1): 123-125, 2013 Cryotechnology of genetic resources is notable as a potential tool to conserve germplasm of endangered wild species [5]. While ejaculated sperm is recovered from live animals, epididymides and testes are potential major sources of male gametes in animals that die unexpectedly [8]. Live births have been produced using cryopreserved epididymal sperm in many species [10,15]. In addition, intracytoplasmic sperm injection (ICSI) with testicular sperm was successful in the human [6]. Siamese Eld's deer (Rucervus eldii siamensis) are native to Thailand but have been extirpated in the wild and are found only in captivity [12]. Although production of offspring after intrauterine insemination with frozen-thawed ejaculated Burmese Eld's deer sperm was reported [9], there are no studies of Siamese Eld's deer reproductive biology. The aim of this study was to evaluate and cryopreserve male gametes recovered from the epididymis and testes of a Siamese Eld's deer that died in captivity.A two-year-old male Siamese Eld's deer maintained at Dusit Zoo, Bangkok, Thailand, unexpectedly died of an unknown cause at 11:00 pm on January 18, 2009, and was kept in a refrigerator until the next day (10:00 am). The right testis with epididymis was removed, stored in phosphate buffered saline at 4°C and transported to a laboratory within 2 hr. Epididymal sperm was collected by placing the caudal epididymis into 10 ml pre-warmed phosphate buffer saline (PBS), cut into small pieces and incubated at 37°C for 10 min. After epididymal tissues were removed, sperm samples were evaluated for sperm concentration, total number, motility (20× magnification with a phase contrast microscope), morphology (William's...