8 First twin lambs produced from demi-embryos in the high Andes of Peru

The aim of this study was to assess the feasibility of producing live lambs after transfer of sheep demi-embryos at high-altitude conditions. The study was conducted at the Casaracra Experimental Center of the Daniel Alcides Carrión National University (Paccha, Yauli province, Junin region, Peru) in the Central Andes Mountains at an elevation of 3,921 m above sea level. Nine sheep were superovulated (Vivanco 2013 Proc. 29th Ann. Mtg. AETE, 43–74); embryos were collected surgically at 7 days post-mating and were handled and graded as per IETS recommendations. Most harvested embryos were allocated to an embryo cryopreservation study, but eight early blastocysts and blastocysts were washed three times in 100 µL of fresh flushing medium (EMCARE™, ICPbio Reproduction) before being transferred individually into a 25-µL drop of splitting medium (ABT 360) devoid of any supplements; drops of splitting medium were placed in a 90 mm tissue culture-treated Petri dish. A microsurgical blade (AB Technology Inc.) mounted on a micromanipulator (Narishige Group) fixed to an inverted phase-contrast microscope (Zeiss) was used to bisect by using only the right arm control attaching the holder of the microblade having the sharp edge parallel to the splitting dish. When embryos were transferred to drops, they were aligned so that the inner cell mass (ICM) was on the left side of the drop to facilitate bisection from left to right with a firm, one-step cut (Vivanco et al. 1991 Theriogenology 35, 292). Care was taken to split the ICM and trophoblast cells into approximately equal halves. After each embryo was bisected, 200 µL of embryo-holding medium was added to the splitting medium to aid in retrieval of the demi-embryos. All embryo handling was performed at 25°C. After bisection, each demi-embryo was transferred individually into 200 µL of embryo-holding medium until being loaded into an IVF catheter for surgical laparotomy transfer into a synchronous recipient. Both halves generated from one original embryo were transferred into a single recipient. A total of 16 demi-embryos were transferred into eight recipients (two demi-embryos per recipient). Two recipients were impregnated and gave birth to a couple of identical twin lambs, each one resulting in 25% survival to lambing of the demi-embryos, equivalent to 50% survival to lambing of the original embryos. From the first pregnant recipient, two male lambs were born with 2.3 and 2.5 kg bodyweight at birth after a gestation of 147 days. From the second pregnant recipient, two male lambs were born with 2.0 and 2.1 kg bodyweight after a gestation of 148 days. The lambs were of the Corriedale breed. Results indicate that it is feasible to produce healthy lambs after transfer of fresh demi-embryos at high-altitude conditions in the Central Andes Mountains. The lambs produced in this study are the first sheep lambs born from demi-embryos in Peru.

This research was supported by Universidad Nacional Daniel Alcides Carrión, Pasco, Peru.