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Biological molecules isolated from organisms that live under subzero conditions could be used to protect oocytes from cryoinjuries suffered during cryopreservation. This study examined the cryoprotectant role of exopolysaccharides of Pseudomonas sp. ID1 (EPS ID1) in the vitrification of prepubertal and adult cow oocytes. IVM oocytes were vitrified and warmed in media supplemented with 0, 1, 10, 100 or 1000 µg mL−1 EPS ID1. After warming, oocytes were fertilised and embryo development, spindle morphology and the expression of several genes in Day 8 blastocysts were assessed. Vitrification led to significantly lower proportion of prepubertal oocytes exhibiting a normal spindle configuration. In fresh control oocytes and most groups of vitrified adult oocytes, similar percentages of oocytes with a normal spindle configuration were observed. Percentages of Day 8 blastocysts were similar for prepubertal oocytes vitrified in the absence or presence of 1 or 10 µg mL−1 EPS ID1 and for adult oocytes vitrified in the presence of 10 µg mL−1 EPS ID1 compared with non-vitrified oocytes. EPS ID1 supplementation had no effect on solute carrier family 2 member 3 (SLC2A3), ubiquitin-conjugating enzyme E2A (UBE2A) and histone deacetylase 1 (HDAC1) expression in Day 8 blastocysts form adult oocytes. However, supplementation with 10 and 100 µg mL−1 EPS ID1 led to increased expression of genes involved in epigenetic modifications (DNA methyltransferase 3 alpha (DNMT3A) and K (lysine) acetyltransferase 2A (KAT2A)) and apoptosis (BCL2 associated X apoptosis regulator (BAX) and BCL2-like 1 (BCL2L1)). The lowest BAX : BCL2L1 ratio was found in the 10 µg mL−1 EPS ID1-supplemented group. The results suggest that 10 µg mL−1 EPS ID1 added to vitrification and warming media may help protect bovine oocytes against cryodamage. Bovine oocyte cryopreservation remains ineffective due to the low ability of cryopreserved oocytes to undergo proper embryo development. This study examined whether the addition of the exopolysaccharide (EPS) ID1, synthesised by Pseudomonas sp. ID1, to vitrification and warming solutions could improve oocyte tolerance to vitrification. The findings show that 10 µg mL−1 EPS ID1 may help stabilise spindle morphology and improve embryo development in vitrified–warmed adult cow oocytes. [ABSTRACT FROM AUTHOR]

The combined use of reproductive technologies, such as transvaginal ovum-pick up and in vitro embryo production followed by direct transfer of cryopreserved embryos, has great potential for enhancing genetic selection and optimising cross-breeding schemes in beef and dairy cattle production systems. This, along with an effective cryopreservation procedure for cow oocytes, will enable the long-term conservation of female genetic traits and the advance of embryo biotechnology in this species. However, the low fertilisation rates and developmental competence of cryopreserved oocytes still need to be improved. Over the past two decades, many research efforts tried to overcome individual features of the bovine oocyte that make it notoriously difficult to cryopreserve. In addition, pregnancy rates associated with in vitro -produced (IVP) embryos remain lower than those obtained using in vivo counterparts. This, together with a lack of a standard methodology for IVP embryo cryopreservation that provides easier and more practical logistics for the transfer of IVP embryos on farms, has hindered international genetic trade and the management of embryo banks. This review updates developments in oocyte and IVP embryo vitrification strategies targeting high production efficiency and better outcomes. [ABSTRACT FROM AUTHOR]

In the present study we examined whether exposure to high concentrations of NaCl or sucrose before vitrification improves the cryotolerance of in vitro-matured bovine oocytes. In Experiment 1, oocytes were exposed to different concentrations of NaCl (375-1517 mOsm) or sucrose (375-812 mOsm) for 1 h. On the basis of the results of this experiment, in Experiment 2 oocytes were exposed to 0.25% NaCl (375 mOsmol) or 2.77% sucrose (375 mOsmol) solution, vitrified and warmed. Microtubule and chromosome configurations were examined by immunocytochemistry. In Experiment 3, in vitro embryo development was assessed after vitrification of oocytes with or without 2.77% sucrose (375 mOsmol) pretreatment. There was a similar percentage of oocytes showing normal spindle configurations in the sucrosepretreated and control groups. Higher rates of abnormal spindles were found in groups treated with NaCl or sucrose solutions with .375 mOsmol. After vitrification and warming, a significantly higher percentage of oocytes with normal chromosome configurations was recorded for oocytes exposed to 375 mOsmol sucrose solution before vitrification compared with the control vitrified oocytes. However, these percentages were significantly lower than those recorded in untreated controls. Cleavage and blastocyst rates were higher in non-vitrified than vitrified oocytes. In conclusion, pretreatment with 375 mOsmol NaCl or sucrose solution had no adverse effects on the spindle status of vitrified-warmed cow oocytes. However, sucrose pretreatment offered no benefits for embryo development. [ABSTRACT FROM AUTHOR]

The low number of oocytes collected from unstimulated donors by ovum pick-up means that embryos produced from each individual female have to be cultured individually or in very small groups. However, it has been demonstrated that single-embryo culture is less efficient than embryo culture in groups. To overcome this limitation, we developed a direct embryo-tagging system, which allows the collective culture of embryos from different origins whilst preserving their pedigree. Presumptive bovine zygotes were tagged with eight wheat-germ agglutinin biofunctionalised polysilicon barcodes attached to the outer surface of the zona pellucida (ZP). Four different barcodes were used to encode groups of 20-25 embryos, which were then cultured in the same drop. Cleavage, Day-7 and Day-8 blastocysts and barcode retention rates were assessed. In addition, Day-7 blastocysts were vitrified and warmed. Barcode attachment to the ZP of bovine embryos affected neither in vitro embryo development nor post-warming survival of the tagged embryos. All the embryos maintained barcodes attached until Day 8 of culture (3.63 ± 0.37 barcodes per embryo) and could be identified. In conclusion, identification of embryos by barcodes attached to the ZP is feasible and will allow the culture of embryos from different donors in the same drop. A novel direct bovine embryo-tagging system has been developed using polysilicon barcodes. This system allows the collective culture of embryos from different origins whilst preserving their pedigree, and facilitates the production of embryos from live animals of high genetic merit. [ABSTRACT FROM AUTHOR]