Assisted Reproduction

Artificial insemination with fresh semen is a valuable management tool for the captive breeding of giant pandas due to the high incidence of sexual incompatibility. It is common practice in China to combine natural breeding and AI. Generally, the one available 'breeding' male is used to copulate with the female after which she is immediately inseminated with semen from one or more males in an attempt to ensure pregnancy and promote genetic diversity. Paternity analysis has revealed that siring success is heavily skewed to the initial mating male rather than to AI (see Chapter 10). This finding is not an indictment of AI efficiency, but more likely a reflection of optimal timing for sperm deposition - the copulating male benefits from peak oestrus and the earlier deposit of large sperm numbers compared to the later AI. Furthermore, AI when used alone has been shown to be as effective at achieving pregnancy as the usual natural mating plus AI approach (Liu, 1981; Feng & Zhang, 1988; Huang et al., 2002; see also Chapter 20). Pregnancies strictly through AI also have been achieved in Spain, Japan and the USA (Moore et al., 1984; Masui et al., 1989; Durrant et al., 2003).

The Biomedical Survey, which produced multiple semen samples, allowed us to conduct further studies which will eventually improve AI efficiency. For example, most current Chinese protocols that rely on fresh sperm mandate that a male be anaesthetised daily for semen collection. So a logical question is - would it be possible to cold-store giant panda sperm over the course of several days without losing functionality? In fact, others have already proven that cooled giant panda sperm from a single ejaculate can be used for AI over two to three days to produce offspring (in Spain: Moore et al., 1984; China, Wolong Nature Reserve: Huang et al., 2002; USA: Durrant et al., 2003). The ramifications are profound because this means that a single semen sample can be collected and used over the two- to three-day oestrus avoiding repeated re-anaesthetisation of the male. An additional benefit is steering clear of sperm cryodamage that occurs at lower temperatures.

Using semen collected during the Biomedical Survey, we further examined the sensitivity of giant panda sperm to cold storage by comparing the influence of two egg-yolk diluents (TEST vs. SFS) and glycerol (0 vs. 5%) on sperm motility and acrosomal integrity over 48 hours at 4°C (Olson et al, 2003; Table 7.5). Percentage sperm motility and normal acrosomes declined over time (p < 0.05). The type of egg-yolk diluent affected sperm motility (p < 0.05) but not acrosomes. After 48 hours of storage at 4°C, motility was lower (P < 0.05) in the SFS (mean range 33-36%) compared to the TEST (52-65%) diluents (see Table 7.5). Glycerol had no influence (p > 0.05) on acrosomal integrity over time in either diluent. However, adding glycerol to TEST reduced (p < 0.05) sperm motility at 48 hours (52%) compared to TEST without glycerol (65%), and both were superior to SFS with or without glycerol. Overall, TEST with 0% glycerol was the most effective diluent for maintaining the

Table 7.5. Influence of refrigeration (4°C) of giant panda semen diluted in TEST or SFS diluent containing 0% or 5% glycerol (n = 6 males) on sperm motility and acrosomal integritya

TEST 0%

TEST 5%

Table 7.5. Influence of refrigeration (4°C) of giant panda semen diluted in TEST or SFS diluent containing 0% or 5% glycerol (n = 6 males) on sperm motility and acrosomal integritya

TEST 0%

TEST 5%

Sperm motility (%)

0 0

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