Which insemination results in fertilization when several are performed before ovulation?

The aim of the present study was to determine which artificial insemination results in fertilization when mares are inseminated several times before ovulation. Mares in oestrus were inseminated over 62 cycles with fresh semen at 48 h intervals from when a follicle > or =30 mm in diameter was detected until ovulation. The number of inseminations was limited to three. Three fertile stallions were used and a different stallion was used for each artificial insemination. The order of the three stallions was changed for each cycle. Embryos were collected between day 10 and day 12 after ovulation and paternity was checked using DNA typing. When two inseminations were performed per cycle, 14 of 17 embryos were the result of the insemination performed on days 2-4 before ovulation and three embryos were the result of the insemination performed on days 0-2. When three inseminations were performed, 1 of 6, 2 of 6 and 3 of 6 embryos resulted from the inseminations performed 4-6, 2-4 and 0-2 days before ovulation, respectively. Thus, 17 of 23 (74%) oocytes were not fertilized as a result of the insemination performed 0-2 days before ovulation. The mean interval between fertilization and ovulation in the mares from which embryos were recovered and tested (n=23) was 2.6 +/- 1.0 days. These results indicate that spermatozoa can remain viable in the genital tract of mares for at least 2.6 days.