Embryo developmental kinetics and vitrification in relation to pregnancy outcome, foaling percentage and foal sex.

Objective Overview

• This study investigated how the timing of embryo development and the method of embryo preservation affect pregnancy success, live birth rates, and the sex ratio of foals produced through in vitro fertilization in horses.

Background and Purpose

• Previous studies suggested that rapid embryo development in vitro correlates with better pregnancy outcomes.
• Less is known about how slower embryo development and the use of frozen (vitrified) versus fresh embryos influence pregnancy success, early pregnancy loss, foaling rates, and foal sex.
• The study sought to analyze:
• The relationship between the day embryos reached the blastocyst stage (days 7 to 11 after fertilization via ICSI).
• The effects of embryo preservation methods, comparing fresh embryos with those vitrified and then warmed using either a one-step or three-step warming process.

Methods

• This was a retrospective clinical study involving 201 equine blastocysts produced by intracytoplasmic sperm injection (ICSI) of in vitro matured oocytes from Warmblood mares.
• Embryos were collected on days 7 to 11 post-ICSI.
• Embryos were transferred to recipient mares four days after ovulation either fresh or after being vitrified and warmed.
• Pregnancy outcomes were measured at days 14 and 42 post-transfer.
• Outcomes assessed included pregnancy rate, early pregnancy loss (EPL), foaling percentage (successful live birth), and foal sex.
• Data analysis was performed using multivariable generalized linear mixed-effects logistic regression models to control for variables.

Key Findings

• Day of blastocyst formation:
• The timing of embryos reaching the blastocyst stage was significantly linked to pregnancy outcomes at days 14 and 42 and to foaling rates.
• Embryos that reached blastocyst stage later (especially at day 11) had markedly lower odds of resulting in live foals.
• For example, embryos transferred on day 11 had an odds ratio of 0.0045 for foaling compared to earlier-developed embryos, indicating minimal chance of live birth.
• Embryo development speed and foal sex:
• Slower developing embryos (e.g., day 10 blastocysts) were associated with a significantly higher likelihood of producing female foals (fillies).
• The odds of having a filly versus a colt were 5.6 times higher for day 10 blastocysts than for day 7 blastocysts.
• Vitrification versus fresh transfer:
• No significant differences in pregnancy rates or foaling percentages were found between fresh embryo transfers and those involving vitrified-warmed embryos.
• The vitrification warming method (one-step vs. three-step) did not impact these outcomes either, emphasizing the effectiveness and flexibility of the vitrification protocols.

Conclusions and Implications

• Slow in vitro embryo development, especially embryos reaching the blastocyst stage on day 11, correlates with lower pregnancy success and foaling rates.
• Embryos developing around day 10 still provide acceptable chances of live birth but tend to produce more female offspring.
• Vitrification is a reliable method for embryo preservation in horses and simplified one-step warming methods are as effective as more complex protocols.
• These findings have practical implications for optimizing embryo transfer timing and preservation strategies in equine assisted reproduction.
• The study notes limited sample sizes for each developmental day, suggesting cautious interpretation and a need for further research.