A COMPARISON BETWEEN INVITRO FERTILIZATION AND MICROINJECTION OF IMMOBILIZED SPERMATOZOA FROM BULLS PRODUCING SPERMATOZOA WITH DEFECTS

The objectives of this study were to compare the fertilization rate of bovine in vitro matured oocytes by in vitro fertilization (IVF) and by microinjection of a single spermatozoon (M1) and to relate these rates with fertility reported for these bulls in artificial breeding. Bull A (Holstein) had a nonreturn rate of 75%. Semen from this bull is routinely used in our standard IVF procedure. Bull B (Ayrshire), used regularly in artificial breeding and related to bull D, had a nonreturn rate of 69.2%. Bull C (Brown Swiss), with a chromosomal translocation and trisomy, achieved a nonreturn rate of 42%. Bull D (Ayrshire) produced nonmotile spermatozoa (SPZ) and had an abnormality described as "tail stump defect." No pregnancies sired by bull D have been reported. Oocytes were either fertilized in vitro by capacitated SPZ or by microinjection of a single immobilized SPZ into the ooplasma. SPZ were treated with 0.1 muM A23187 and used for IVF. For microinjection SPZ were cocultured for 5 h with bovine oviduct epithelial cells (BOEC),and then immobilized by freezing and thawing twice without cryoprotectant. A single batch of killed SPZ (stored at -25-degrees-C) was used for all microinjections. All oocytes were cultured in Medium 199 for 22 h at 39-degrees-C and subsequently fixed, stained, and examined for evidence of fertilization (i.e., female and male pronucleus formation, SPZ decondensation). Fertilization rates following IVF with semen from bulls A, B, C, and D were 80%, 54%, 1%, and 2%, and following microinjection were 39%, 22%, 21%, and 34%, respectively.