296 FREQUENT RECTAL PALPATIONS FOLLOWING SUPEROVULATORY TREATMENT AFFECT SEX RATIO OF EMBRYOS RECOVERED FROM HOLSTEIN HEIFERS

Abstract

Skewing the sex ratio of offspring towards males or females is very important for the livestock industry. Many factors, such as maternal stress, have been suggested to affect the sex ratio (Pratt NC et al. 1989 J. Reprod. Fertil. 87, 763–769). In a recent study (Ideta A et al. 2007 J. Reprod. Dev. doi:10.1262/JRD.19035), the proportion of female embryos recovered from superovulated heifers in which ovulation patterns were observed by repeated transrectal ultrasonography tended to be higher than the expected ratio of 50:50 (66.7%, 26/39). To investigate this phenomenon, we repeated the experiment using a larger number of Holstein heifers. The superovulatory treatment began in the midluteal phase of the estrous cycle (days 8 to 10) and consisted of eight decreasing doses of FSH i.m. (total of 28 Armour units, Antrin R-10, Kawasaki-Mitaka, Kanagawa, Japan) for 4 days with treatment twice daily. Doses of 5 mL and 3 mL of a PGF2α analogue (Veterinary Pronalgon F Injection containing 5 mg mL–1 Dinoprost, Pfizer Animal Health, Tokyo, Japan) were administered i.m. to the animals along with the seventh and eighth FSH treatment, respectively. The heifers were divided into two groups. One group, the rectal palpation (RP) group (n = 9), received transrectal ultrasonography with rectal palpation at 4-h intervals from 36 to 76 h after the first PGF2α treatment. The other group, the Control group (n = 8) received no treatment. The heifers were artificially inseminated at 56 and 72 h after the first PGF2α treatment using frozen–thawed semen from one bull. Seven-day embryos were recovered nonsurgically. Grade 1 to 3 embryos (IETS classification) were selected for this study. Male and female embryos were separated using the loop-mediated isothermal amplification procedure (Hirayama H et al. 2004 Theriogenology 62, 887–896). Data were analyzed using ANOVA and chi-square test. The mean number of recovered ova and embryos was 15.7 � 3.8 (RP) and 14.4 � 2.2 (Control). There was no significant difference in the percentages of unfertilized ova (RP; 14.9 %, 21/141 and Control; 11.3% 13/115, P > 0.05), grade 1 embryos (RP; 51.1%, 72/141 and Control; 54.8%, 63/115, P > 0.05) and grade 1 to 3 embryos (RP; 65.2%, 92/141 and Control; 69.6%, 80/115, P > 0.05) between the two groups. The proportion of female grade 1 embryos in the RP group (66.7%, 48/72) was significantly higher than the expected ratio of 50:50 (P < 0.01). The female ratio of grade 1 embryos in the Control group was 50.8% (32/63). Furthermore, the proportion of female grade 1 to 3 embryos in the RP groups (66.3%, 61/92) was significantly higher than the expected ratio of 50:50 (P < 0.005). The female ratio of grade 1 to 3 embryos in the Control group was 51.3% (41/80). Results indicate that frequent ultrasound examinations and rectal palpations following superovulatory treatment may skew the sex ratio of embryos towards females in Holstein heifers.