Assisted Reproduction Techniques to Improve Reproduction in a Non-Model Species: The Case of the Arabian Bustard (Ardeotis arabs) Conservation Breeding Program.

Artificial reproductive technologies are highly valuable for ex situ conservation. While Arabian bustard populations are declining and extinct in some parts of the range, the International Fund for Houbara Conservation in the United Arab Emirates implemented a conservation breeding program. Since 2012, a total of 1253 eggs were laid through natural reproduction, 1090 were incubated and 379 of these were fertile (fertility rate of 34.8%), leading to the production of 251 chicks. To improve fertility and acquire crucial knowledge for other endangered large birds, artificial reproduction was implemented in 2018 using fresh, refrigerated, and frozen sperm. A total of 720 ejaculates were collected from 12 birds. We analysed these samples for concentration, volume, motility score (0 to 5), viability (eosin/nigrosine), length, and morphology. The first age at collection was 35.7 ± 18.8 months, mean volume was 89.2 ± 65.3 µL, mean concentration was 928 ± 731 sptz/mL and mean motility score was 2.61 ± 0.95. Morphology analyses revealed a bimodal distribution of sperm length. Five hundred and thirty-five ejaculates were cryopreserved and the initial motility score was 3.4 ± 0.7 and 2.0 ± 0.6 after thawing, while the percentage of normal and intact membrane sperm cells decreased from 88.8 ± 7.5% to 52.9 ± 1%. Sixty-five artificial inseminations were performed, leading to a global fertility rate of 84.3%-more precisely, 85.2% and 83.3%, respectively, for fresh and cryopreserved semen. All methods successfully produced fertile eggs, indicating that artificial insemination is an efficient tool for the conservation and genetic management of the species.

Evaluation of sperm head dimensions and chromatin integrity of epididymal sperm from domestic cats using the toluidine blue technique.

When using assisted reproductive technologies, there is seldom an evaluation of DNA integrity during sperm analysis, which is an important variable for proper embryo development. The toluidine blue staining technique allows the simultaneous evaluation of sperm chromatin and sperm head dimensions. The objectives of this study were to evaluate the applicability of the toluidine blue staining method for analyzing DNA abnormalities in epididymal sperm (from the caput, corpus, and cauda) of cats and to investigate the correlations among DNA condensation, morphology, and sperm head dimensions. The DNA alteration indexes were obtained using the toluidine blue and acridine orange techniques, and comparisons of these indexes indicated there was a 65.4% (r = 0.654; P < 0.001) correlation. The sperm from the cauda had greater chromatin stability (97.9%) than the sperm from the epididymal head (92.1%; P = 0.0023). There, however, was no difference in chromatin stability between sperm obtained from the corpus and cauda regions, indicating that these sperm were already mature. The sperm head dimension was correlated with chromatin decondensation, and the sperm head size decreased as the sperm were transported through the three epididymal regions (P < 0.0001). In addition, the percentage of sperm that were deficient in chromatin condensation decreased as the sperm were transported through the epididymal caput, corpus and cauda (26.4, 15.7, and 3.4%, respectively; P < 0.0001). Thus, the sperm head size predicts the quality of chromatin condensation in sperm cells.

Does lipid peroxidation and oxidative DNA damage differ in cryopreserved semen samples from young, adult and aged Nellore bulls?

The aims of this study were to evaluate cryopreserved semen of Nellore bulls of different ages and verify whether sperm quality declines with advancing age and whether lipid peroxidation and DNA damage are involved in this process. For this purpose, 40 Nellore bulls were divided into three age groups: Young, aged 1.8-2 years (n = 9); Adult, aged 3.5-7.0 years (n = 19); and Seniors, aged 8.0-14.3 years (n = 12). Three ejaculates were collected from each bull, cryopreserved and evaluated for various parameters including membrane integrity, mitochondrial potential (FITC-PSA and JC1), lipid peroxidation (C-11BODIPY 581 / 591) and oxidative DNA damage (8OHdG) using flow cytometry. The thawed semen of senior bulls was characterized by a low percentage of motile sperm (33.7 ±â€¯6.1%), higher damage to the plasma and acrosomal membrane (37.5 ±â€¯9.8%), and low mitochondrial potential (29.1 ±â€¯13.8%), as well as higher percentages of peroxidated cells (53.6 ±â€¯12.2%) and DNA damage (44.1 ±â€¯11.0%; P < 0.05). Lipid peroxidation was negatively correlated with motility (r = -0.35, P < 0.0002), average mitochondrial potential (r = -0.42; P < 0.0001) and showed a positive correlation with membrane injury and oxidative DNA damage (r = 039; P = 0.0003). Young bulls presented superior thawed sperm quality, possibly due to greater resistance to oxidative stress and, consequently, to cryopreservation. In conclusion, the sperm quality of bull semen declines with advancing age and is strongly associated with increased oxidative damage to both the plasma membrane and DNA.