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Recent advances in nanotechnology have tremendously expanded its possible applications in biomedicine. Although, the effects of nanoparticles (NPs) at cellular and tissue levels have not been fully understood, some of these biological effects might be employed in assisted reproduction to improve male fertility particularly by enhancing sperm cell quality either in vivo or in vitro . This review summarises the available literature regarding the potential applications of nanomaterials in farm animal reproduction, with a specific focus on the male gamete and on different strategies to improve breeding performances, transgenesis and targeted delivery of substances to a sperm cell. Antioxidant, antimicrobial properties and special surface binding ligand functionalization and their applications for sperm processing and cryopreservation have been reviewed. In addition, nanotoxicity and detrimental effects of NPs on sperm cells are also discussed due to the increasing concerns regarding the environmental impact of the expanding use of nanotechnologies on reproduction.

Dietary polyunsaturated fatty acids (PUFAs) can influence fertility in farm animals. Some evidence in mice and sheep have suggested that PUFAs may influence offspring sex ratio, which may have significant value for cattle production. To test this hypothesis, three groups of Holstein cows were supplemented with either 0%, 3% or 5% protected fat (PF) in the form of calcium salt of fatty acids (rich in omega-6) from 14-21 days pre-partum until conception. Proven-fertile frozen semen from the same ejaculate was used for insemination. Calf sex recorded at birth was 8/19 (42.1%) male offspring in the control group, increasing to 14/20 (70%, P  > 0.05) and 17/20 (85%, P  < 0.05) in 3% and 5% PF, respectively. To test if this effect was caused by a direct influence on the oocyte, we supplemented bovine cumulus oocyte complexes during in vitro maturation with either omega-3 alpha-linolenic acid (ALA), omega-6 linoleic acid (LA) or trans-10, cis-12 conjugated linoleic acid (CLA). Sex ratio of the produced transferable embryos was determined using PCR of SRY gene. Similar to the in vivo results, sex ratio was skewed to the male side in the embryos derived from LA- and CLA-treated oocytes (79% and 71%) compared to control and ALA-treated oocytes (44% and 54%, respectively). These results indicate that both dietary and in vitro supplementation of omega-6 PUFAs can skew the sex ratio towards the male side in cattle. Further experiments are required to confirm this effect on a larger scale and to study the mechanisms of action that might be involved.

Semen cryopreservation is a well-established procedure used in veterinary assisted reproduction technology applications. We investigated damaging effects of cryopreservation on the structural and ultrastructural characteristics of bull sperm induced at different temperatures and steps during standard cryopreservation procedure using transmission (TEM) and scanning electron microscopy. We also examined the effect of cryopreservation on sperm DNA and chromatin integrity. Five healthy, fertile Friesian bulls were used, and the ejaculates were obtained using an artificial vagina method. The semen samples were pooled and diluted in a tris-yolk fructose (TYF) for a final concentration of 80 × 10 6  spermatozoa/ml. The semen samples were packed in straws (0.25 ml), and stored in liquid nitrogen (-196°C). Samples were evaluated before dilution, just after dilution (at 37°C), at 2 h and 4 h during equilibration, and after thawing (37°C for 30 s in water bath). In association with step-wise decline in motility and viability, our results showed that the plasma membrane surrounding the sperm head was the most vulnerable structure to cryo-damage with various degrees of swelling, undulation, or loss affecting about 50% of the total sperm population after equilibration and freezing. Typical acrosome reaction was limited to 10% of the spermatozoa after freezing. We also observed increased number of mitochondria with distorted cristae (15%). Chromatin damage was significantly increased by cryopreservation as evident by TEM (9%). This was mainly due to DNA breaks as confirmed by Sperm Chromatin Structure Assay (SCSA) (8.4%) whereas the chromatin structure was less affected as evaluated microscopically by toluidine blue staining. We concluded that, using standard cryopreservation protocol, the most pronounced damage induced by cryopreservation is observed in the plasma membrane. Further improvement of cryopreservation protocols should thus be targeted at reducing plasma membrane damage. Acrosomal, mitochondrial and chromatin damage are also evident but appear to be within acceptable limits as discussed.