Factors Affecting the Intrafollicular Transfer of Immature Oocytes in Beef Heifers: Does the Injection Matter?

This study aimed to evaluate whether the quality of injection and number of oocytes injected during Intrafollicular Transfer of Immature Oocytes (IFIOT) affect ovulation timing, follicle dynamics, oocyte recovery, nuclear maturation and in vitro embryo production in beef heifers. In three experiments, ovulator cows were synchronised and subjected to IFIOT, followed by the evaluation of follicular regression, recovery of cumulus-oocyte complexes (COCs), oocyte nuclear maturation, embryo development and pregnancy outcomes after embryo transfer. Data were analysed using the chi-square test and Proc Glimmix (SAS Institute). The injection procedure did not affect ovulation timing but induced follicular regression (p < 0.05). Injection quality and number of oocytes injected affected COC recovery (p < 0.05), with higher recovery rates observed following good-quality injections and lower oocyte numbers (48.90% vs. 24.49%). Although nuclear maturation rates and embryo production were similar (p > 0.05) across the groups, embryos produced via IFIOT exhibited a higher (p < 0.05) proportion of blastocysts classified as suitable for cryopreservation (51.72% vs. 33.81%). Furthermore, after transfer, the pregnancy rates of cryopreserved IFIOT-derived blastocysts were comparable to those of embryos produced using conventional in vitro maturation. These findings indicate that although IFIOT did not increase the total embryo yield, it improved oocyte recovery and significantly enhanced the production of high-quality cryotolerant blastocysts. Therefore, improving the injection technique is critical for enhancing IFIOT efficiency and embryo developmental competence.