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P Imrat,
P Suthanmapinanth, K Saikhun,
S Mahasawangkul,
E Sostaric,
P Sombutputorn,
S Jansittiwate,
N Thongtip,
A Pinyopummin,
B Colenbrander,
W V Holt,
T A E Stout
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ABSTRACT: Semen cryopreservation and artificial insemination (AI) are potentially valuable methods for supporting the breeding management of endangered species like the Asian elephant. Cryopreservation of Asian elephant semen has however proven problematic with respect to maintenance of both adequate semen quality and fertility post-thaw. In this study, nine ejaculates from three adult bulls were used to compare the influence of extender (TEST versus INRA96(®)) and penetrating cryoprotectants (3% glycerol, 5% glycerol and 4% methylformamide) on post-thaw semen quality. We demonstrate that not only the freezing process, but also the quality of the semen before freezing, significantly influences the freezability of Asian elephant semen. Pre-freeze motility, viability, semen volume, semen pH, sperm concentration and the incidence of sperm mid-piece and tail abnormalities all significantly (p<0.05) affected post-thaw semen quality. While extender and cryoprotectant did not significantly affect any of the above semen quality parameters post-thaw, the skim-milk based extender (INRA96(®)) preserved DNA integrity better (p<0.05) than the egg yolk extender (TEST). Considerable between-ejaculate variation in all post-thaw semen quality parameters was also noted. It is concluded that strict criteria for semen quality is essential for the selection of Asian elephant bull ejaculates suitable for cryopreservation; stricter initial selection should improve the mean post-thaw quality.
Cryobiology 11/2012; · 2.06 Impact Factor
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ABSTRACT: This study was examined whether the species of felid affects synchronization accuracy at the G0/G1 stage of the cell cycle and the occurrence of apoptosis by different protocols, such as serum starvation, confluent and roscovitine treatment. Skin fibroblast cells were obtained from the Asian golden cat, marbled cat, leopard and Siamese cat. The cells from each animal were treated with either serum starvation for 1-5 days, cell confluency-contact inhibition for 5 days or roscovitine at various concentrations (7.5-30 μm). Flow cytometric analysis revealed that serum starvation for 3 days provided the highest cell population arrested at the G0/G1 stage, irrespective of the felid species. In all species, 100% confluency gave a significantly higher percentage of cells arrested at the G0/G1 stage compared with the non-treated control cells. The effects of roscovitine treatment and the appropriate concentration on the rates of G0/G1 cells differed among the felid species. Serum starvation for more than 4 days in the marbled cat and Siamese cat and roscovitine treatment with 30 μm in the Asian golden cat and leopard increased the rates of apoptosis. In conclusion, different felid species responded to different methods of cell cycle synchronization. Asian golden cat and Siamese cat fibroblast cells were successfully synchronized to G0/G1 stage using the serum starvation and roscovitine treatment, whereas only confluency-contact inhibition treatment induced cell synchronization in the leopard. Moreover, these three methods did not successfully induce cell synchronization of the marbled cat. These findings may be valuable for preparing their donor cells for somatic cell nuclear transfer in the future.
Reproduction in Domestic Animals 07/2012; · 1.36 Impact Factor
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ABSTRACT: Knowledge about the acrosomal status of Asian elephant (Elephas maximus) sperm is extremely limited. The objective of this study was to evaluate the viability and acrosomal status of Asian elephant sperm following induction by calcium ionophore and heparin using propidium iodide (PI) and fluorescein isothiocyanate conjugated peanut agglutinin (FITC-PNA). Semen samples were collected from elephant bulls by manual stimulation. Semen was diluted with extender, cooled to 4 degrees C and transported to a laboratory for the experiment. Sperm cells were incubated in modified Tyrode's medium containing either 1 mM calcium ionophore or 10 mg/ml heparin for 5 h at 39 degrees C. Sperm recovered at the onset (0 h), 1, 2, 3, 4 and 5 h of incubation were simultaneously assessed for the viability and acrosomal status using dual staining of FITC-PNA and PI. Results were confirmed by transmission electron microscopy. A progressive increase in the proportion of live-acrosome reacted sperm was observed within 3 h of incubation in both treatment groups which slightly decreased at 4 to 5 h of incubation. At 1 to 3 h of incubation, the percentage of live-acrosome reacted sperm induced by calcium ionophore was higher (P < 0.05) than those induced by heparin and the control. However, there were no statistical differences at 4 to 5 h of incubation. A progressive reduction of the percentage of motile sperm was observed in the control as well as both treatment groups. Sperm motility decreased sharply when they were incubated in calcium ionophore compared with incubation in heparin and control groups. These results indicate that the occurrence of live-acrosome reacted sperm in the Asian elephant was induced by calcium ionophore at a rate higher than that induced by heparin.
Journal of the South African Veterinary Association 09/2009; 80(3):146-50.
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ABSTRACT: The serow (Capricornis sumatraensis) is a critically endangered species. The objectives of this study were to evaluate ejaculate quality in captive males, and to investigate and characterize sperm morphology. Semen was collected using electroejaculation. Mean (+/-S.D.) seminal characteristics were: semen volume 2.3+/-0.8 mL, pH 7.8+/-0.4, and osmolality 329.9+/-32.9mOsmol/kg; sperm concentration 515.8+/-263.1 x 10(6) cells/mL; wave motion score (1-5) 3.9+/-0.4; motile sperm 60.5+/-22%; viable sperm 68.3+/-9.4%; morphologically normal sperm 70.8+/-19.3%; and an opacity that was yellowish to milky-white. Sperm head length, width, degree of elongation, area, and perimeter were 6.0+/-0.6 microm, 4.3+/-0.3 microm, 71.7+/-8.6%, 19.8+/-2.5 microm(2), and 17.9+/-2.1 microm. Based on these measurements, we categorized sperm head morphometry as small, medium, or large. In addition, sperm morphology was examined by light and scanning electron microscopy; overall, morphologically normal and abnormal sperm were similar to those reported for other bovidae. In summary, this study provided baseline data regarding semen characteristics of C. sumatraensis, which should be of value in the preservation of this endangered species.
Theriogenology 11/2008; 71(4):576-85. · 1.96 Impact Factor
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ABSTRACT: This study was conducted to investigate the effects of capacitating agents added at in vitro fertilization (IVF) and antioxidants supplemented during in vitro culture (IVC) on the development of buffalo embryos. In experiment I, in vitro embryo development of buffalo embryos was compared when the IVF medium was supplemented with heparin, caffeine and calcium ionophore A23187 either alone or in combination. There was no significant difference (P > 0.05) in the cleavage rates of oocytes among the treatment groups but the development rate to the blastocyst stage and the cell numbers of blastocyst in the heparin-treated group were significantly higher (P < 0.05) than that of other treatments. In experiment II, in vitro embryo development of buffalo embryos was compared when IVC medium was supplemented with either α-tocopherol (250 and 500 μM) or l-ascorbic acid (250 and 500 μM). The rate of development to the blastocyst stage of embryos cultured in medium supplemented with 250 μM α-tocopherol (33%, 41/123) and 250 μM l-ascorbic acid (31%, 38/123) was significantly higher (P < 0.05) than that of those cultured in medium alone (19%, 20/108) but not significantly different (P < 0.05) from medium supplemented with either 500 μM α-tocopherol (24%, 30/123) or 500 μM l-ascorbic acid (25%, 33/133). These results suggest that buffalo spermatozoa treated with heparin were suitable for IVF and that α-tocopherol and l-ascorbic acid added during IVC increased the rate of buffalo embryo development.
animal 10/2008; 2(10):1486-90. · 1.74 Impact Factor
