On-farm lambing outcomes after transfer of vitrified and slow frozen embryos

https://doi.org/10.1016/j.anireprosci.2020.106467Get rights and content

Highlights

  • A frozen-thawed embryo transfer policy is suggested in small-scale sheep farms.

  • Different embryonic stages and freezing methods gave similar pregnancy outcomes.

  • In the field, embryos are practical to be collected at morula/blastocyst stages.

  • Vitrification method is considerably recommended in field conditions.

Abstract

The aim of the present study was to determine the most suitable embryonic stage and embryo freezing technique for commercial implementation of frozen embryo trading by small-scale sheep producers. There was a 2 × 2 factorial design utilized for conducting the study consisting of two embryo stages (2−8 cells or morula/blastocyst) and two cryopreservation protocols (vitrification or slow-freezing). For the in vivo produced embryos, there were treatments of crossbred donor ewes to induce superovulation. Embryos were recovered surgically on either Day 2 or 5.5 after estrous onset. The embryos were cryopreserved using either a vitrification or slow-freezing method before there was transfer to recipients. Ovarian response, embryo survival and lambing outcomes were analyzed. There were no differences in number of recovered and fertilized embryos at the two embryonic developmental stages. There were no effects of embryonic stages and cryopreservation methods on pregnancy rate, twinning rate, fetal birth weights and lamb weight at 1 month of age. When there was use of vitrified embryos for transfers, there was a greater lamb weight at 2 months of age (8.38 ± 0.20 compared with 7.78 ± 0.21 kg; P =  0.044) than when there was transfer of embryos cryopreserved using slow freezing procedures. Considering economic and practical benefits to small-scale sheep farms, morula/blastocyst stage-embryo collection and transfer into the uterus is more efficacious than transferring 2−8 cells embryos into the oviduct. Results of this study may contribute to the genetic improvement in the flocks of small-scale sheep producers.

Introduction

The biotechnology known as “assisted reproductive technologies (ART)” can be effectively used for improving genetic and reproductive performance in livestock production enterprises (Choudhary et al., 2016). Of the ART procedures, artificial insemination (AI) has been nationally implemented in Thailand’s livestock industries while embryo transfer (ET) is only used for research, especially when considering small ruminants, although it has been well developed in some parts of the world including Australia, South Africa, Mexico and Argentina for use in livestock enterprises (Moore and Hasler, 2017; Paramio and Izquierdo, 2014). In Thailand, the sheep and goat industry is continually growing, with a population of 0.9 million in 2019 (90 % goat and 10 % sheep), concomitant with the increase in farmer numbers every year (DLD, 2019). These operations are considered small scale (approximately 10–20 animals per farm). To utilize the advanced technology, (i.e. ET at small holder farms) the acceptance of farmers of this technology is essential.

It, however, is not clear whether the use of embryos at different developmental stages and/or different cryopreservation procedures will be more effective for ET by small ruminant producers. Pregnancy rates after ET and embryo survival rates were not different when there was transfer of either fresh or vitrified blastocysts (Shelton, 1992; Baril et al., 2001). Likewise, these rates were not affected by the quality and stage of embryo development (Spella et al., 2001). Mangalraj et al. (2009) and Kaur et al. (2014) have reported that pregnancy rates were greater with blastocyst transfer compared with the transfer of cleavage stage embryos. Results from another study indicated the transfer of embryos that had been vitrified at the morula stage resulted in a greater pregnancy rate than transfer of morulae-stage embryos that had been cryopreserved using slow freezing techniques (Green et al., 2009). Conversely, cryopreservation of embryos that were derived using in vitro fertilization (IVF) procedures using the slow freezing technique resulted in greater embryo development (cleavage and blastocyst rates) than with transfer of embryos cryopreserved using vitrification procedures (Bhat et al., 2014).

In a previous study, results indicated that use of an average ratio of 10–200 females per one male over 5 years results in an inbreeding rate that is undesirable (Techakumphu et al., 2018). Recently, there was development of a business model for small holder sheep farms, where ET was included to improve the genetic diversity in the flocks and herds of small producers (Khunmanee et al., 2019). This will help to solve the problems of inbreeding due to the availability of few rams in herds/flocks and the lack of breeders focused on genetic improvement in small holder farms. There, however, is no study of the application of ET in farms of small holder producers. The aim of the present study, therefore, was to determine the optimal stage of embryo collection and embryo freezing technique for application of ET by small holder sheep farmers in Thailand.

Section snippets

Materials and methods

The present study was conducted from September 2016 to March 2018 on a farm located at the Center of Learning Network for the Region (CLNR), Chulalongkorn University, Saraburi province at 14°31’59.99” N, 100°55’0.01” E, Thailand. All procedures in this study were consistent with the ethical standards issued by the National Research Council and were approved by Ethics Committee for Human and/or Animal Experimentation in meeting No.1531062, Faculty of Veterinary Science, Chulalongkorn University.

Embryo retrieval and cryopreservation

Regarding superovulation, all donor ewes (n = 44) responded to the superovulation treatment regimen as indicated by the presence of CL on Days 2 (6.70 ± 1.00) and 6 (6.77 ± 0.94) after insemination. The mean number of recovered embryos was 4.50 ± 1.01 (67.1 %) and 3.73 ± 1.09 (55.0 %) on Days 2 and 6 after insemination, respectively. The average number of embryos was 3.72 ± 1.00 (82.8 %) and 3.55 ± 1.02 (95.1 %) at Days 2 and 6 after insemination, respectively (Table 1). There were no

Discussion

To apply the business model using ET technology to small holder sheep farms, it is important to know the suitable stage and embryo freezing technique for application. For the success of an ET business, there are five important factors: local and international market, technology that is available, type of livestock production enterprise, government policy and investment priorities (Poapongsakorn, 2012). Apparently, there is considerable demand for goat and sheep at markets, increasing at an

Declaration of Competing Interest

The authors declare no conflict of interest.

Acknowledgements

This research was funded by the Agricultural Research Development Agency (Public Organization), ARDA, 2015, Thailand (grant number: 5807010560) and Research Assistant Scholarships, Graduate School, Chulalongkorn University. Sarawanee Khunmanee was PhD candidate in the program of the National Research Council of Thailand and the Industries (RRI) Ph.D. program (grant number: PHD58I0089). Special thanks to Andrew Lobb from Faculty of Arts, Chulalongkorn University for English proof reading.

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