Japanese Black beef cattle breeds (Wagyu) are well-known for producing meat with high-quality marbling that contains large amounts of oleic acids (omega-3 and omega-6) [1]. Full-blood Wagyu breeds in China are mostly imported from the USA and Australia, and multiple ovulation and embryo transfer technology (MOET) is routinely performed when breeding large numbers of Wagyu cattle [2]. However, ovum pickup combined with in vitro fertilization (OPU-IVF) has great potential for use in cattle reproduction and breeding, mainly due to a fast donor turnaround, the female age range available for OPU (i.e., calves, prepuberal females, heifers to older cows), and because it can be performed regardless of reproductive condition (open or pregnant) [[3], [4], [5]]. Thus, the importance of successful adoption of OPU-IVF in cattle breeding has become increasingly apparent, not only for traditional IVF initiatives but also for the production of large numbers of embryos from elite donors to enhance breeding capability [6].

OPU-IVF in Japanese Black cattle was first reported by Konishi et al. in 1996 using inhibin-immunized multiparous cows [7], and this technique has subsequently been used for Wagyu in vitro embryo production and breeding [[7], [8], [9], [10], [11]]. One prior study assessed the effect of suckling on the production of embryos following repeated OPU-IVF before and after timed artificial insemination (AI) in postpartum Japanese Black cows [12], and another examined the effects of season and reproductive phase on OPU oocyte developmental competence in these animals [13]. Currently, measurement of plasma anti-Müllerian hormone concentrations is used to select OPU heifer donors in breeding programs [6]. However, knowledge of donor effects on OPU-IVF outcomes and the developmental potential of embryos is limited. Thus, studies aimed at determining the effects of donor age range and multiparity on OPU and IVF outcomes are needed to expand reproductive breeding programs. Several previous investigations have shown that synchronization of follicle wave emergence before follicle-stimulating hormone (FSH) superstimulation and OPU improves in vitro embryo production in pregnant Holstein heifers [3,[14], [15], [16]]. However, it is unclear whether repeated FSH stimulation and consecutive OPU sessions affect blastocyst development of aspirated oocytes in Japanese Black cattle. Additionally, some MOET donors have an inefficient response to hormone superovulation and lower in vivo embryo yields [[17], [18], [19]]. However, it is unclear whether elite Japanese Black cattle with lower embryo yields from MOET can be used for OPU-IVF to produce more embryos for breeding.

The objectives of this study were to i) determine the effects of donor age and multiparity on OPU-IVF outcomes, ii) assess whether the use of oocytes from FSH-superstimulated donors subjected to five consecutive OPU aspiration sessions affects blastocyst development, and iii) evaluate whether elite Japanese Black beef donors with low embryo yields from MOET can be used to efficiently generate large numbers of embryos via OPU-IVF for herd breeding and expansion.