First Evaluations and Cryopreservation of Semen Samples from Sunda Clouded Leopards (Neofelis diardi)

Abstract

<jats:p>A better understanding of semen characteristics and resilience to freezing temperatures is necessary before developing assisted reproductive techniques and systematic biobanking for the Sunda clouded leopard. The objective of this study was to evaluate for the first time the semen and sperm quality (in fresh and frozen samples) of two captive Sunda clouded leopards in Malaysia. A total of 17 examinations of the reproductive tract (using ultrasonography) and electro-ejaculations were performed on the two leopards over a 2-year period. Samples obtained from Leopard 1 (8 years old) varied in terms of volume (402 ± 92 µL), pH (7.9 ± 0.9), sperm motility (54.5 ± 24.2%), sperm concentration (122.4 ± 84.7 × 106 sperm/mL), normal morphology (23.9 ± 12.3%), and viability (55.2 ± 18.2%). Midpiece defects represented the most common structural abnormality followed by abnormal tail and head defects. Samples from Leopard 2 (11 year old with abnormal testicular tissue) were of lesser quality. Two frozen semen samples from Leopard 1 were thawed and examined for acrosome integrity. Post-thawed samples contained &lt;10% of motile spermatozoa but almost 50% of abnormal acrosomes. The present results emphasized the high incidence of structurally-abnormal spermatozoa, similar to the mainland clouded leopard. Post-thaw evaluations showed that the few surviving spermatozoa could potentially be used for in vitro fertilization or sperm injection. However, more individuals must be studied to validate those first findings that are exciting but still preliminary.</jats:p>

The Comparison of Semen Collection in Electroejaculation, Rectal Massage and Combination of Both Methods in the Critically Endangered Malayan Pangolin, Manis javanica

“…The pangolins are closely related phylogenetically with the order of Carnivora [ 35 ], prompting a potential comparison across the order due to the lack of available data. Although some semen characteristics in pangolins were similar when compared to the wild felid [ 36 ], the semen volume, concentration and viability appeared to be different. In contrast, the semen characteristics in brown bears seemed to be superior in terms of total motility [ 37 ].…”

A preliminary study on semen collection, its evaluation, and testicular and sperm morphometries in the wild proboscis monkey (Nasalis larvatus)

“…The semen pH was measured using pH indicator strips (Whatman, Cytiva, Global Life Science Solutions, Marlborough, MA, USA). The semen motility was assessed using a CX43-32P01 phase contrast microscope (Olympus, Tokyo, Japan) based on semen criteria previously detailed ( 30 ). In brief, a volume of 5 μL of each ejaculate was placed on a 37°C prewarmed glass slide with a coverslip and 200 spermatozoa cells were counted.…”

“…For morphological examination, thin smears on glass slides were prepared from droplets of diluted semen. If the sperm concentration was moderate or below, a dilution ratio of 1 : 2 (aliquot of semen : stain) was used ( 30 ) and if the sperm concentration was high, a dilution ratio of 1 : 3 (aliquot of semen : stain) was used ( 30 ). Two types of stains were utilised: Diff-Quik for observation of abnormalities and eosin-nigrosin for evaluation of live or dead status of spermatozoa.…”

Establishment of semen collection technique using electroejaculator and semen cryopreservation of Javan leopard (Panthera pardus melas Cuvier, 1809)

“…The stimulation protocol was done by modifying the technique performed in the Sumatran tiger [23] and clouded leopard [11], which was conducted in as many as five stimuli at each voltage starting at 4.0, 4.5, 5.0, 5.5, and 6 V. Each trigger is given 5 s with a pause between impulses of 4.0-5.0 V of 5 s each, while 5.0-5.5 V with a pause of 10 s, and 5.5-6 V with a pause of 5 s. Erection and ejaculation reaction data were tabulated and descriptively presented.…”

“…Studies on the reproductive biology of the Javan leopard have never been reported, unlike in several other wild cats, including the Indian [5], Arabian (P. pardus nimr) [6], and the Indochina (P. pardus delacouri) leopard [7]. Semen cryopreservation efforts in wild cats such as the jaguar (Panthera onca) [8], clouded leopard (Neofelis nebulosa) [9], Prionailurus bengalensis euptilurus [10], and the Sunda clouded leopard (Neofelis diardi) [11] have been reported. A sperm bank/genome bank is a vital step and effort for longterm wildlife conservation [12].…”