 Reference Base  Collection and analysis of semen from a black rhinoceros |
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| Location: |
World |
| Subject: |
Reproduction - Management methods |
| Species: |
Black Rhino |
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Numerous factors, including poaching and human encroachment, have contributed to declining numbers of the black rhinoceros ( Diceros bicornis) in the wild. As a result, the black rhinoceros has been classed as an endangered species.
Many zoos, wild animal game parks, and preserves have attempted to establish breeding pairs, but with mixed success. One of the main problems has been incompatibility. Thus, we saw the need to develop a technique for obtaining semen from the rhinoceros. A secondary reason was to determine the ability of rhinoceros spermatozoa to withstand freeze preservation techniques, which we have applied to other mammalian species. The capability of successfully freezing rhinoceros sperm would be of obvious advantage to zoos and captive collections in this and other countries desiring to propagate the species without the risks of shipping such large and unpredictable animals. It is a simple matter to ship stored semen to those interested in artificial insemination of the species.
A black rhinoceros at the National Zoo had sired offspring in 1967, 1970, and 1978. Later, a female rhinoceros died of tuberculosis. Results of subsequent intradermal tuberculin testing included a positive response in this male. It was treated with isoniazid for 6 months, after which time it was decided o euthanize it because of declining health and risk of spreading the disease to the rest of the herd.
The animal was given 0.6 mg carfentanil citrate (ZR 33799; Janssen Pharmaceutica, Beerse, Belgium) by pole syringe. In 11 minutes, immobilization was adequate for electroejaculation.
Electroejaculation involved administering a 110-V, 60-Hz. sine wave stimulus from a stimulator capable of an output of 0 to 60 V continuous range and 0 to 1 A.
Initially, rectal probe stimulation was combined with direct penile stimulation, by attaching one lead from the stimulator to the ventral electrode lead of the rectal probe and one lead to a 5-cm-wide aluminium foil electrode wrapped around the rhinoceros' penis. Neither erection nor ejaculation were obtained. Therefore, the penis was extended manually and two aluminium foil electrodes were wrapped around the penis approximately 17 cm apart (Fig. 1). Physiologic saline solution was the used to wet the area of the electrodes. The electric leads were attached to these electrode strips, and stimulus was applied. By the eighth stimulus, using 22V and 200 mA, the penis became fully erect and an ejaculate was obtained (Fig. 2). Electrical stimulation was continued, using a range of 10 to 29 V and 150 to 210 mA. The stimulus pattern was approximately 2 seconds from 0 to peak voltage, 3 seconds at peak voltage, and 2 seconds from peak to 0 voltage. After 3 seconds at 0 voltage, the pattern was repeated. Ejaculation continued through a total of 62 cycles, at which point the rhinoceros was rested for 45 minutes.
The ejaculate was analyzed, using previously reported procedures. Mean (n=10) dimensions of the spermatozoa were:
head length, 6.0 m; head with, 3.5 m; midpiece-neck length, 7.8 m; midpiece width, 0.84 m; tail length, 36.8 m; and total length, 50.6 m.
Table 1 contains the results of semen analysis and freezing. The sperm appeared undamaged after thawing from -196? C. On the basis of morphological features and motility of the motile sperm, which we used for making an in vitro estimate of fertilizing ability, the sperm appeared capable of fertilization. Sperm motility was rapid in both ejaculates, before and after freezing. Table 2 lists the two voltage, amperage, and stimulus patterns used to obtain the ejaculates. Both patterns seemed to elicit similar ejaculatory response, although the second collection contained fewer sperm, most likely because of sperm depletion in the reproductive tract.
Table 1. Results of semen analysis and freezing in the rhinoceros
Collection Volume Total sperm Post-collection Post-freeze
l count motility (%)* motility (%)
1 44.6 4.4 x 106 60 20
2 18.8 1.4 x 106 40 15
* Percentage motile sperm observed at x 100
Table 2. Electroejaculation patterns used in the black rhinoceros.
Collection Voltage Amperage Stimulus pattern volts
range range (mA) (no. stimulations)
1 10-29 150-210 29 (14); 10 (3); 15 (5); 22 (20); 17 (20)
2 8-25 60-200 8 (4); 10 (5); 15 (5); 20 (5); 10 (3); 15 (5);
20 (10); 25 (8); 10 (4); 15 (5); 10 (5);
15 (5); 20 (6)
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