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Due to the lack of Rhinoceros and high poaching rates, zoological institutions and wildlife conservation need to enhance the Rhinoceros’ populations by performing Artificial Insemination (AI) in ex-situ conservation. Preserving genetic diversity by banking biological material such as sperm is essential during AI operation and wildlife ecosystem maintenance.
Semen sample quality needs to be consistent as sample exchanges are performed between institutions and fertility changes are tracked from the endangered species. Concentration, which is the number of sperm in a sample, and motility, known as the portion of sperm swimming, are crucial to determine the fertilization success rate.
How is the semen evaluation performed?
In this research, semen assessments are performed with both fresh and cryopreserved samples collected from Black Rhinoceros (Diceros bicornis), White Rhinoceros (Ceratotherium simum), and Greater One-Horned Rhinoceros (Rhinoceros unicornis) with the approval of the Cincinnati Zoo and Botanical Garden’s Institutional Animal Care and Use Committee by the zoological institutions.
The parameters of the iSperm software application were adjusted to determine the semen sample quality. Spermatozoa were considered motile with average path velocity (VAP) > 5 µm/s. And those with straight-line velocity (VSL) > 25 µm/s and straightness of sperm movement (calculated as VSL/VAP) > 75% were considered progressively motile sperm.
Then, manual assessments with a hemocytometer were done on a separate aliquot to validate the iSperm’s analysis. To determine the accuracy of sperm cell identification by the iSperm and the accordance of both concentration evaluations, the Pearson correlation (P-value ≤ 0.05 is significant) and Bland–Altman analyses (ratio = 1 indicates complete agreement) were performed.
Is the iSperm reliable and can be standardized for sperm assessment?
It was proven that the analysis performed by the iSperm had a significant correlation with the manually observed sperm evaluation with r = 0.9908, P < 0.0001 as shown in Fig. 1A. And the bias (mean ratio) between the iSperm and the observer was 0.97 in Fig. 1C, which was close to a complete agreement. Semen samples diluted with different extenders, for instance, egg- and milk-based formulations, were still evaluated accurately by the iSperm system.
The iSperm-derived values were remarkably in agreement with values obtained using a hemocytometer with r = 0.9847, P < 0.0001 in Fig. 1B. Altogether with a 1.03 bias (mean ratio) as shown in Fig. 1D, which shows the correlation between the iSperm count and hemocytometer count.
Excellent reliabilities of the iSperm’s concentration and total motility were obtained as the Intraclass Correlation Coefficient (ICC) is higher than 0.96, as shown in Table 1.
What does iSperm offer for the evaluation of sperm quality?
In summary, this study has demonstrated the effectiveness and reliability of the iSperm mCASA for assessing the quality of rhinoceros sperm. Through rigorous evaluation of fresh and cryopreserved samples from multiple rhinoceros species, we have established a significant correlation between the iSperm analysis and manual observation of sperm quality. The iSperm’s accuracy, indicated by the close agreement in results and the high Intraclass Correlation Coefficient (ICC), shows its potential as a valuable tool for ex-situ conservation.
With its portability and tablet-based interface, the iSperm provides a convenient means of evaluation in laboratory settings and a practical solution for fieldwork. This ability to quickly track and analyze moving sperm cells, particularly those displaying progressive motility, enhances assessments’ accuracy, compatibility, and repeatability.
This article is brought to you by iSperm mCASA, a mobile/mini computer-assisted semen analyzer with high accuracy and affordability for use anytime, anywhere by anyone. Check out www.isperm.co or Contact us to the iSperm can assist your breeding program.
- Rispoli, L.A. and Roth, T.L. (2023). Validation of the iSperm for assessing rhinoceros Sperm. Theriogenology Wild, 3. https://doi.org/10.1016/j.therwi.2023.100048.
- Rhino Calf Horns mother and - free photo on Pixabay - Pixabay. (n.d.). https://pixabay.com/photos/rhino-calf-horns-mother-and-child-6065480/