The first equine ICSI pregnancy was reported in 1996 [1] and since then the popularity of this advanced reproductive technology has been spreading to different laboratories in countries all over the world [2], and embryonic technologies are currently used in 28.7% of the world's countries [3]. This is associated with various aspects, including the potential to produce foals from infertile, old, or even deceased mares [4]. It also applies to rare semen samples with lower sperm viability. For example, based on our experience, a single semen straw can contribute to 10-15 ICSI sessions. In addition, equine ICSI-derived embryos can be effectively cryopreserved and stored [5], facilitating recipient management, embryo transfer and embryo commercialization.

Owing to these advantages in comparison to conventional reproductive techniques, horse breeders are increasingly using ICSI for embryo production. Different from commercial ICSI programs in the United States, the OPU-ICSI approach in Brazil is based on sending OPU teams to aspirate donor mares on-site and transporting the aspirated oocytes to a central laboratory. In Europe, veterinarians aspirate mares in breeding centers and send oocytes to an ICSI clinic or mares that have OPU and ICSI performed on the same facility. While the OPU-ICSI program reported in this manuscript enables bringing this advanced technology to many farms without the need of moving mares to breeding centers, having numerous Veterinarians performing OPU on different farm settings, and transporting the oocytes to an ICSI facility may introduce some factors that affect the success of embryo production.

Several factors have been identified to interfere with the results of OPU-ICSI embryo production, such as donor mare characteristics, the number of recovered oocytes [6] and different times of the estrus cycle [7]. In addition, old mares were reported to have less follicles and less oocytes recovered, which reduces the chance of producing ICSI embryos compared to young mares [6], [8], [9]. Season and breed of the oocyte donor mare have also been reported to affect the success of producing equine ICSI. For example, Cuervo Arango and coworkers [8] did not find a significant effect on oocyte developmental competence, but achieved higher oocyte recovery rate out of the breeding season in the northern hemisphere than in the breeding season. Although several published studies have reviewed the factors affecting the production of equine ICSI embryos in Europe and North America [6], [10], [9], [8], [11], [12], [13], [14]. Similar studies based on a large number of ICSI embryos have not been conducted in South America. Therefore, this study evaluated factors that affected the production of equine ICSI embryos under conditions in Brazil.