Successful Vitrification of In vivo Embryos Collected from Superovulated Japanese Black Cattle (Wagyu)

Abstract

The aim of this study was to determine whether vitrification is an effective method when used for Japanese Black Cattle (Wagyu) in vivo-derived embryos, collected following a superovulation treatment and embryo transfer (MOET) programme. In vivo-derived morula and blastocysts collected on day 7 after artificial insemination, were vitrified using a modified droplet vitrification (MDV) procedure and subsequently warmed for transfer (ET) into synchronized recipients. Fresh embryos, and embryos cryopreserved using a standardized slow freezing procedure (direct thaw/direct transfer, DT) served as ET controls. Two different follicle-stimulating hormone (FSH) sources, Folltropin(®) Canada (FSH BAH, 24 donors) and a brand prepared by the Chinese Academy of Science (FSH CAS, 16 donors), were compared in a series of superovulation outcomes following well-established FSH administration protocols. Following data analysis, the total number of ovulations recorded at the time of embryo flushing (10.5 vs 8.5; p = 0.28) and the total number of transferable embryos (6.2 vs 5.1; p = 0.52) were similar between the two FSH sources. ET for MDV (39.7%, n = 78), DT (35.2%, n = 71) and fresh controls (47.1%, n = 34) resulted in similar pregnancy rates (p > 0.05). When MDV was used, a higher pregnancy rate (42.6%) resulted from the transfer of vitrified morulae, when compared to the DT counterparts (24.3%), (p = 0.05). Transfer of vitrified morulae resulted also in higher pregnancy rate, when compared to the transfer of vitrified blastocysts (42.6% vs. 29.4%; p < 0.05). Transfer of DT blastocysts resulted in higher pregnancy rate than morulae, similarly cryopreserved (47.1% vs. 24.3%, p < 0.05). In conclusion, MDV is an effective alternative methodology for cryopreservation of in vivo-derived embryos. This study gives also indication that, compared to vitrified blastocysts, MDV of morula stage embryos results in higher pregnancy rates following warming and transfer into synchronized recipients.

Effects of service number on conception rate in Japanese Black cattle

“…The mean CR of the Japanese Black cattle in this study was 47.0 ± 0.5% for heifers and 47.8 ± 0.2% for cows, respectively. In previous studies, Japanese Black cattle have been reported to have CR of 51.7% after frozen‐thawed embryo transfer (Nishigai, Kamomae, Tanaka, & Kaneda, ) and 47.1% after fresh embryo transfer (An et al., ). Other beef breeds have been reported to have a 73% CR after embryo transfer (Spell, Beal, Corah, & Lamb, ), and local zebu cattle have a reported 57.7% CR after insemination with frozen semen (Mollah et al., ).…”

The Outcome and Economic Viability of Embryo Production Using IVF and SOV Techniques in the Wagyu Breed of Cattle

“…As described by An et al [ 20 ], the hormones administered in a superovulation protocol and embryo transfer protocol are part of the initial upfront costs involved for this technique, which may be either cost-prohibitive or hard to obtain in a developing country. Considering this, the use of the IVF technique is a great alternative to attained high pregnancy rates in animals that do not respond well to SOV protocols; at the same time, that improves the production and the genetic value of Wagyu herds.…”

“…Limited information is available regarding the response of the Wagyu cattle to SOV protocols. In China, the superovulation protocols with the use of 400 mg of Folltropin have produced an acceptable number of transferable embryos per collection per cow [ 20 ]. However, not much is known about the success of the reproductive characteristics of the Wagyu breed in Brazil, South America—a country with tropical and subtropical climates.…”

Significant heparin effect on bovine embryo development during sexed <i>in vitro</i> fertilization

“…The modified droplet vitrification method developed by our group, resulted in a satisfactory and reliable outcome with sexed IVF bovine embryos [8], in Wagyu beef embryos in vivo [8, 28], cloned bovine embryos [38], and mouse embryos [39]. In this study, the pregnancy/implantation rate of 39.1–40.3% derived from IVF with heparin at a concentration of 20–40 µg/ml was not different to that observed in the routine IVF controls (heparin 10 µg/ml), and was similar to that reported in our previous studies with embryos produced through sexed and non-sexed IVF [8].…”

“…Embryos were cryopreserved by modified droplet vitrification (Vitrification and Warming Kit, Renova Life, College Park, MD, USA) [8, 28]. Briefly, embryos were treated with 0.25% trypsin (Gibco, Grand Island, NY, USA) for 1 min.…”