Embryo biopsy and vitrification applied to a commercial embryo transfer program in Argentina

Abstract

Use of embryo vitrification (EV) would have multiple advantages in a commercial embryo transfer (ET) program. EV would allow embryos to be transferred so that foals were born in early spring, which is preferred in Argentina due to health problems in foals that are born in late spring-summer. Currently, many high-level Polo mares compete in the spring, and the potential for embryo production from these mares is thus lost. EV would allow embryo storage against future loss or sale of a mare and allow owners to control which embryos were transferred in any given season. EV would also help the ET Center, by spreading work out through the breeding season and simplifying recipient synchronization. To explore the use of EV in our program, we compared pregnancy rates after transfer of vitrified-thawed vs. fresh embryos. Embryos were recovered on Day 7 to 8 post-ovulation from donor mares in our clinical program at Doña Pilar in September 2021 to May 2022 (Season 1) and then vitrified, and in September 2022 to January 2023 (Season 2) and transferred fresh. All embryos were transferred during Season 2. Fresh embryos were either biopsied via micromanipulation for sexing by PCR (BF, n=136) or were not biopsied (client did not request sexing orthe embryo was considered too small to biopsy, <∼300 µm diameter; F, n=62). Vitrified embryos were vitrified using a commercial human kit (CryoTech) after collapse of the blastocoele by micromanipulation, with biopsy (BV, n = 69) or without biopsy (MV, n = 9), or were vitrified without micromanipulation (<∼300 µm diameter; V). To date, the pregnancy rates per transferred embryo were BF 77/109 (70.6%); F 47/59 (79.6%); BV 24/36 (66.7%); MV 4/4 (100%) and V 9/14 (64.2%). There were no significant differences between treatments (Fishers exact test, p > 0.2). We conclude that embryo biopsy did not affect viability of fresh embryos, and that notably, the pregnancy rate was equivalent between fresh biopsied embryos (71%) and vitrified biopsied embryos (67%). The numerically lowest pregnancy rate resulted from embryos vitrified without manipulation. To the best of our knowledge, this is the first report on pregnancy rates of equine embryos vitrified after biopsy. These results indicate that a biopsy and vitrification program is feasible in a commercial embryo transfer center in Argentina. We foresee that in the near future most embryos collected after December 31st in our program will be vitrified and transferred between September and October of the following year.