37 EFFECT OF EMBRYO STAGE ON PREGNANCY RATE FOLLOWING DIRECT TRANSFER OF BOVINE EMBRYOS FROZEN IN ETHYLENE GLYCOL

Abstract

Annually, more than 400 000 in vivo-recovered bovine embryos are officially reported by members of the Canadian and American Embryo Transfer Associations. Between 65 and 70% of these embryos are cryopreserved and more than 95% are frozen in ethylene glycol (EG). Statistics on factors affecting embryo freezing are difficult to obtain because many cattle breeders/farmers no longer report pregnancy rates back to embryo transfer (ET) practitioners. Concerns are often expressed as to the optimal stage at which to freeze bovine in vivo-derived embryos. This is a retrospective analysis of results from 5 commercial ET programs (1 in the United States, 3 in Canada and 1 in the Netherlands) for which pregnancy data relative to embryo stage at freezing were made available. Embryos representing 4 stages of development, as defined by the IETS (4 = late morula, 5 = early blastocyst, 6 = mid blastocyst and 7 = expanded blastocyst) are included in the data. The number of embryos thawed and transferred ranged from 3954 to 24 827 for the 5 programs, with a total of 72 828. Embryos were frozen in either 1.5 M EG or 1.5 M EG + 0.1 M sucrose and exposure time to cryoprotectant before cooling ranged from 4 to 40 min. Pregnancy rates are shown in Table 1. Although the pregnancy rate for stage 6 embryos was only 2.6 and 3.2 percentage points lower than stages 4 and 5, respectively, these differences were highly significant and pregnancy rates for stage 6 embryos were lower than those for stages 4 and 5 in 4 of the 5 ET programs. The small decreased survival of stage 6 embryos is probably only moderately important in a commercial context. However, the pregnancy rate of stage 7 embryos was lower than all other stages for the combined dataset as well as in all 5 ET programs, with the difference between stages 5 and 7 ranging from 6.5 to 16.4 percentage points. Clearly, stage 7 embryos survive freezing at a significantly lower rate than stages 4, 5 and 6 and neither time of exposure to EG nor inclusion of sucrose in the freezing medium provided an obvious improvement. Although bovine ET practitioners routinely attempt to collect embryos on day 7 post-oestrus, recovery of stage 7 embryos cannot always be avoided. Further investigation into factors contributing to the decreased survival of stage 7 embryos is warranted.

Table 1.  Effect of embryo stage on pregnancy rate of bovine embryos frozen in EG