Artificial insemination (AI) is a potentially useful tool for breeding captive elephants because it facilitates efforts to minimise inbreeding. However, cooled storage of elephant semen markedly reduces fertility. This study compared the effects on semen-quality parameters, including sperm DNA fragmentation, of storing elephant semen at 4°C or 15°C in a commonly-used diluent (TEST) or a diluent developed to protect against sperm DNA damage (BullMax). Storing elephant semen for >24 h in either extender at either temperature resulted in decreases in sperm motility, viability, acrosome integrity and DNA integrity (P < 0.05); the decrease in motility was especially rapid. A subjective impression of circular sperm movement in TEST was confirmed by a higher curvilinear velocity and amplitude of lateral head displacement, but lower straight-line velocity and linearity than in BullMax. Initial percentages of spermatozoa with fragmented DNA (%SDF) did not differ between extenders or temperatures, but the rate of increase in %SDF during a 48-h incubation at 37°C was higher in TEST than in BullMax (P < 0.05). In conclusion, BullMax allows more linear movement and better preserves DNA stability of stored elephant spermatozoa than TEST. Sperm DNA stability during incubation at 37°C is a promising, discriminative parameter for selecting semen storage conditions of bulls for elephant AI.

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 postthaw 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. Prefreeze 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 postthaw 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.

In captivity, male Asian elephants often yield poor quality semen after transrectal manually assisted semen collection; however, the reasons for the disappointing semen quality are not clear. Here we test the hypothesis that accumulation of senescent spermatozoa is a contributory factor, and that semen quality can therefore be improved by more frequent ejaculation. To this end we investigated the effect of collecting semen five times on alternate days, after a long period of sexual rest, on semen quality in Asian elephants known to deliver poor semen during infrequent single collections. All eight bulls initially displayed a high incidence of detached sperm heads and low percentages of motile (close to 0%) spermatozoa. After semen collection on alternate days, the percentages of detached sperm heads, and head and mid-piece abnormalities, were reduced significantly (p<0.05). In particular, one bull showed markedly improved sperm motility (increased from 0% to 60%) and membrane integrity (increased from 5% to 75%). In addition, advancing age significantly (p<0.01) correlated with lower percentages of sperm with intact membranes and a higher frequency of detached sperm heads. In contrast to sperm accumulation problems in other species, a small ampullary diameter correlated significantly (p<0.05) with reduced semen quality.