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

BACKGROUND: While fast in vitro embryo development has been associated with improved outcomes, the relationships of slow in vitro embryo development and vitrified versus fresh in vitro produced (IVP) embryo transfer to foaling percentage and foal sex are less explored. OBJECTIVES: To determine the relationships of (1) day of blastocyst formation (D7-11 after intracytoplasmic sperm injection (ICSI)) and (2) vitrification and warming method (one- and three-step) versus fresh transfer with pregnancy, early pregnancy loss (EPL), foaling percentage and foal sex. STUDY DESIGN: Retrospective clinical study. METHODS: Blastocysts (n = 201) were collected on days 7-11 after ICSI of in vitro matured oocytes from Warmblood mares and transferred either fresh or vitrified-warmed into recipient mares on day 4 after ovulation. Pregnancy, EPL, foaling percentage, and foal sex were compared using multivariable generalised linear mixed-effects logistic regression models. RESULTS: Day of blastocyst formation was significantly associated with pregnancy outcome at D14 (p = 0.048), D42 (p = 0.046) and foaling percentage (p < 0.001). The odds of foaling decreased with developmental age, with significantly lower odds after D11 blastocyst transfer (OR = 0.0045, 95% CI 0.00034-0.061; p < 0.0001). Slow embryo development was associated with a significantly higher proportion of female offspring. The odds of obtaining a filly versus a colt were 5.6-fold higher for D10 blastocysts than D7 (95% CI 1.14-25.0; p = 0.034). Transfer of fresh versus vitrified-warmed blastocysts, using a one- or three-step protocol, did not significantly affect pregnancy or foaling outcome. MAIN LIMITATIONS: Small sample size per day of blastocyst formation. CONCLUSIONS: Slow in vitro embryo development is associated with decreased pregnancy and foaling outcomes following transfer of D11 IVP blastocysts. Transfer of D10 IVP blastocysts yields acceptable foaling percentages and is associated with a higher proportion of female offspring. Vitrification is effective for preserving equine embryos and allows the use of a simplified one step warming protocol.