In Vitro Fertilization as a tool for the Genetics Improvement. History and Future Prospects.

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1 In Vitro Fertilization as a tool for the Genetics Improvement. History and Future Prospects. Ovum pick up followed by embryo production procedure (OPU-IVP) is strongly driven by the need of the breeding industry to enhance genetic improvement in dairy or beef cattle specially after the advent of sexed semen. This has now become even more important with the introduction of Genomic Selection (GS) with the possibility to aspirate very young heifers and collect the biopsy direct from the embryo. So an extensive use and successful OPU-IVP program is desired in order to increase the number of embryos and offspring per donor which will subsequently allow an enhanced selection intensity for the next generation. In the last decade, fertilization emerged as an alternative to superovulation and has become the technique of choice for bovine embryo production, especially in zebu breeds (Viana et al., 2012). Overall, there was a 7% reduction in the number of flushes reported worldwide from 111,806 in 2008 to 104,282 in However, the number of IVP embryos was up by 47,000 (12%) from a year ago. Brazil was responsible for 68% of the IVP in this period. The number of IVP embryos transferred also showed a healthy 17% increase from 254,000 in 2008 to 307,000 in One factor that contributed to a decrease in the activity of cattle worldwide is the increase of embryo production, especially in South America according to IETS Statistics and Data Retrieval Committee Report In 2011 global in vivo bovine embryo transfers remain stable, while in vitro transfers increased for the sixth consecutive year according to IETS Statistics and Data Retrieval Committee Report The Brazilian IVEP industry Three different periods were characterized for the use of IVF technologies in the Brazilian embryo industry: 1) the early years ( ), when IVF growth was driven by the growing demand from the embryo market, although the technology was still labeled as elitist; 2) a period of exponential growth ( ), when IVF overcame conventional ET as the technique of choice for embryo production; and 3) a later period, when total numbers tended to stabilize but IVF started to increase in importance in dairy breeds. The whole picture shows IVF as an interesting example of innovation, since the development of these new embryo technologies provided new products, processes and possibilities to satisfy demands and remarkably change the scenario of the Brazilian embryo industry (Viana et al., 2012).

2 To investigate why the preferred means to produce bovine embryos in Brazil has changed from in vivo to in vitro, we compared these two approaches in the same Nelore cows (n = 30) and assessed total embryo production and pregnancy rates. Without a specific schedule, all cows were subjected to ultrasound-guided ovum pick up (OPU)/in vitro production (IVP) and MOET. Embryos obtained from both methods were transferred to crossbred heifers. We concluded that in Nelore cows, with an interval of 15 d between OPU/IVP procedures, it was possible to produce more embryos and pregnancies compared to conventional MOET (Pontes et al., 2009). Figure 1. Production of bovine embryos in Brazil, according to the technique employed, during the period of 1995 to ET: embryos produced by superovulation (conventional embryo transfer); : embryos produced in a (laboratory; IVEP). Despite the importance of Nelore as a beef breed well adapted to tropical areas, there is also a growing interest for Zebu dairy breeds, such as Gir considering their adaptability to produce large amounts of milk under stressful conditions, e.g. high temperature, parasites, and on poor pasture. These characteristics are maintained in Gir-Holstein crossbred animals, usually named Girolanda, which are popular dairy cattle in Central and South America, and potentially other tropical areas. Recently, the method for embryo production has been considered for Girolanda donors, due to the growing efficiency of using sexed sperm in IVF, which facilitates production of a large number of females for the dairy industry. We develop a study to investigate oocyte yield using OPU and embryo production with sexed sperm, and to determine pregnancy rates following extended transportation of embryos from

3 (Holstein), (Gir), and (Holstein " Gir) breeds. All results were obtained from In Vitro Brasil, a large commercial IVF production. A total of 90,086 oocytes were collected in 5,407 OPU sessions and a total of 64,826 oocytes were used for IVF, producing 16,924 embryos (Pontes et al., 2010). To our knowledge, this is the largest program of embryo production in dairy cattle using sexed sperm. It is expected that this information can be used to facilitate the expansion of OPU/IVF programs in dairy Herds in Brazil and in others countries. Another reason for the success of IVEP in Brazil is the scale effect. IVEP activity has high fixed-costs, but allows for optimization in the use of high-cost semen straws and sexed semen, and also an improvement in the logistics of recipients synchronization and management, mostly due to a better predictability of oocyte yield per donor. The gain in predictability is more evident if compared to embryo production, a technique that often shows coefficients of variation greater than 100%. Consequently, when used on a large scale, the cost per pregnancy from IVEP can be lower than from conventional ET ( Viana et al., 2012). In Vitro Brasil Data in different countries As ripple effect, the very same model has been successfully benchmarked to a few different countries. In Uruguay (a relevant player in the global beef industry and the

4 only live cattle exporter and dairy heifers/cows supplier to China on the Western part of the world), for instance, IVP, started from scratch, and has produced 6,904 IVF embryos in 2011 and 8,972 in As China regularly imports ship loads of Holstein heifers, the seedstock industry tailors its processes to meet all demands (large scale regular offers of pregnant Holstein IVF heifers, in most cases, bred from another Holstein IVF embryo). A very similar breakthrough scenario has been taking place in South Africa. Again, from roughly (not say literally) nothing, during a 120 days long breeding season, production developed from 2,858 IVF embryos in 2010, 5,028 in 2011 to 6,384 in As result, the Boran registered herd jumped from approximately 4,000 animals in the African continent to over 4,000 Boran IVF products (yearlings, calves and confirmed pregnancies) only in South Africa from 2010 to In a different point of this learning curve, Russia has definitely understood the role played by this powerful breeding tool. Throughout a very brief Spring season (60 days long), over 500 for OPUs produced 2,799 viable IVF embryos from a recently imported set of top Angus donors heifers and cows. Commercial first calf cows (also imported from the USA and AUS, less than 6 months ago, as bred heifers) were the only source of recipient groups (conceptions rates for those groups, under fixed time ET protocols, were close to 60%). Focused on reproducing/multiplying best performances in a herd scale, rather than one (or few) specific high-end elite animal, those countries aim for very efficient and consistent industry prospects. Final Remarks IVEP technologies are interesting examples of innovation in livestock production. Firstly, the development of reproductive protocols was based on the confluence of research efforts in correlated areas. Secondly, IVEP should not be considered merely as an improvement in the way bovine embryos are produced when compared to conventional ET, since it resulted in new products, procedures, applications, and, why not, a new market. Thirdly, the technology was quickly adopted by the private sector and replaced the preceding one ( embryo production) as the standard procedure in Brazil, South Africa, Uruguai and others countries. Finally, IVEP caused a significant change in the scenario of the world embryo industry, in beef and dairy herds, with predictable consequences for genetic progress and farm productivity. This facilitates a significant increase in the selection intensity and

5 shortening of the generation interval, finally leading to the major goal of the breeding industry: enhanced genetic improvement. References Pontes JH, Nonato-Junior I, Sanches BV, Ereno-Junior JC, Uvo S, Barreiros TR, Oliveira JA, Hasler JF, Seneda MM Comparison of embryo yield and pregnancy rate between in vivo and in vitro methods in the same Nelore ( ) donor cows., 71: J.H.M. Viana, L.G.B. Siqueira, M.P. Palhao, L.S.A. Camargo Features and perspectives of the Brazilian embryo industry.anim. Reprod., v.9, n.1, p.00-00, Jan./Mar J.H.F. Pontes, K.C.F. Silvaa, A.C. Bassob, A.G. Rigo, C.R. Ferreirab,G.M.G. Santosa, B.V. Sanches, J.P.F. Porcionato, P.H.S. Vieira, F.S. Faifer,F.A.M. Sterzaa, J.L. Schenkd, M.M. Seneda, Large-scale embryo production and pregnancy rates from,, and dairy cows using sexed sperm. 74 (2010) J.H.F. Pontes, F.A. Melo Sterzab, A.C. Basso, C.R. Ferreira, B.V. Sanches,K.C.P. Rubin, M.M. Seneda Ovum pick up, embryo production, and pregnancy rates from a large-scale commercial program using Nelore cattle( ) donors. 75 (2011) B.V. Sanches, L.S.R. Marinho, B.D.O. Filho, J.H.F. Pontes, A.C. Basso,.L.G.Meirinhos,K.C. Silva-Santos, C.R. Ferreira, M.M. Seneda Cryosurvival and pregnancy rates after exposure of IVF-derived Bos indicus embryos to forskolin before vitrification. 80 (2013)