Abstract
M.Sc. (Zoology)
The reproductive biology of the South African leopard, Panthera pardus has not been studied in detail. In South Africa little is known about the population numbers of leopards due to their solitary and nocturnal nature and currently the conservation and management of leopard populations relies mainly on the contributions of non-governmental organisations, academic institutions and private individuals.
The aim of this study was to provide baseline information for the development of in-situ and ex-situ reproductive conservation methods for the leopard. In order to meet this aim, the following objectives were established: 1) determine the degree of relatedness of the leopards sampled, 2) establish baseline parameter values of a whole blood count and describe the ultrastructure of the blood cells, 3) obtain semen by means of electroejaculation and determine the efficiency of a previously described cryopreservation protocol for leopard spermatozoa, 4) describe the morphology and ultrastructure of the leopard spermatozoa using florescence and electron microscopy, 5) describe the histology and ultrastructure of the leopard testes and the events of spermatogenesis using light and electron microscopy.
Between January 2011 and February 2013, blood and semen samples were obtained from eleven leopards after being sedated with a combination of Medetomidine and Ketamine. The DNA was extracted from the blood (ARC Genetics Department) and analysed (Onderstepoort Veterinary Genetics Laboratory). The blood was also used for the analysis of the baseline blood parameter values (Lancet Laboratories). Whole blood was fixed in 2.5% phosphate buffered gluteraldehyde and prepared for transmission and scanning electron microscopy to describe the ultrastructure of the cells. Techniques to examine sperm morphology included florescence and electron
xviii
microscopy. The semen was fixed in 2.5% gluteraldehyde and phosphate buffer for the ultrastructural assessment. Testes samples obtained from a leopard that died during transportation were fixed in Bouin’s fixative and a phosphate buffered 2.5% gluteraldehyde solution for light and electron microscopy respectively. The testes samples were prepared using standard techniques and stained with Hemotoxylin and Eosin for light microscopy and uranyl acetate and lead citrate for electron microscopy.
The DNA analysis revealed that two pairs of leopards were related on a half-sibling level. The mean parameter values of the whole blood count of P. pardus were similar to the values recorded for Asian leopards, P. pardus African lions, Panthera leo and bobcats, Lynx rufus and fell within the normal ranges for the domestic cat, Felis catus. The ultrastructural assessment of the blood cells was comparable with those that have been described for the Asian leopard as well as most other mammalian species. A small volume of semen (≤0.5 ml) could be obtained from five out of nine male leopards that were sampled. The morphology and ultrastructure of the leopard spermatozoa conforms to the generalised structure of spermatozoa of most mammalian species. A large number of morphologically abnormal spermatozoa were noted. This has also been reported for many feline species, including the Indian leopards. Spermatozoa abnormalities identified included coiled tails, cytoplasmic droplets and knobbed acrosomes. The cryopreservation of the spermatozoa yielded a maximum post-thaw progressive motility of 24.4%. The histology and ultrastructural events of spermatogenesis in the leopard testes were compared to that of the domestic cat and some differences were observed between the domestic cat testes and leopard testes. The results of this study provide baseline information on the genetic diversity and reproductive biology of the leopards in South Africa. This can be used in the development of assisted reproductive techniques that may one day aid in conservation strategies for the leopards.