Effects of Individual Stallions and Mare Breeds on Assisted Fertilization In Vitro

Abstract

Comparative experiments were conducted to assess the effect o f individual stallions on the fertilizing ability o f the spermatozoa in in-vitro-matured and partially zona-removed (PZR) equine oocytes (Experiment 1), as well as the effect o f mare breeds for ovaries from which oocytes were collected (Experiment 2). Experiment 1: Spermatozoa from 3 fertile stallions (A: Thoroughbred, B: Crossbred, and C: Palomino) were separately frozen-thawed in 5% glycerol and washed by a 4530% discontinuous percoll-gradient. Aliquots of the sperm were incubated for 3 h at 38.5°c in Brackett and Oliphant (BO) medium containing 2.5 mM caffeine and then treated for 60 sec with 0.1μM calcium ionophore A23187. The sperm suspension was introduced into BO medium supplemented with 0.3% BSA where 6 to 10 PZR oocytes after 32 h maturation culture were placed, and the oocytes and spermatozoa were co-incubated for 20 h. There were significant differences among stallions in the fertilization rates with PZR oocytes (34.2, 72.2, and 100% in stallions A, B, and C, respectively; P<0.05). High frequencies of monospermy in the fertilized oocytes were observed in the stallions with low fertilization rates (76.9, 57. 7, and 22.0% in stallions A, B, and C, respectively), resulting in similar overall proportions o f monospermic fertilized zygotes (range 22.0 to 41.7%). Experiment 2: Using mixed semen from 2 stallions (Stallions A and B), spermatozoa were incubated with PZR oocytes which were derived from the mare ovaries o f di f ferent breeds (Draft and Light horse). No significant difference in fertilization rates was found between the mare breeds (Draft and Light horse, fertilization rates 50.0 and 52.6%; overall monospermic fertilization rates 40.5 and 42.1 %, respectively).