352 IMPROVING THE EFFICIENCY OF LAPAROSCOPIC INTRAOVIDUCTAL INSEMINATION WITH SEX-SORTED BOAR SPERMATOZOA

Abstract

The insemination of a low number of sex-sorted spermatozoa is a critical issue that must be solved in order to enable the commercial application of this technology in pigs. A new procedure for laparoscopic intraoviductal insemination in sows has recently been reported (Vazquez et al. 2005 Reprod. Dom. Animals 40, 375 abst.). To improve the efficiency of this technique, this experiment was designed to determine the influence of insemination time, relative to the time of ovulation, on the number and quality of zygotes recovered after laparoscopic insemination of sows with sex-sorted spermatozoa. Spermatozoa were stained with Hoechst 33342 and sexed using the EPICS Altra flow sorter (Coulter Corporation, Miami, FL, USA) modified to operate at 42 psi for spermatozoa. Sorted spermatozoa were collected in tubes containing 1.5 mL of TEST-yolk (2%)-seminal plasma (10%). Post-weaning crossbred sows (n = 212; parity 2-4) were hormonally treated with eCG (Folligon; Intervet, Boxmeer, The Netherlands) and hCG (Veterin Corion, Divasa, Spain) and their ovaries were examined using transrectal ultrasonography at intervals of 4 h from 30 h after hCG injection to the laparoscopic insemination. Sows were allotted into three groups according to their ovarian status at insemination: preovulatory (P; n = 131), ovulating (O; n = 43), and ovulated (CL; n = 38) follicles. Follicle status was reconfirmed at insemination by direct observation using the laparoscope. Sows were inseminated in both oviducts with 0.3 million sex-sorted spermatozoa in 0.1 mL of extender. Eighteen hours later, putative zygotes were collected by washing the oviducts after laparotomy, fixed, stained with lacmoid, and examined by phase-contrast microscopy. Penetration rates were evaluated as numbers of monospermic and polyspermic oocytes per oocytes collected. Monospermic rates were evaluated as numbers of monospermic oocytes per oocytes penetrated. Data were analyzed by ANOVA. The number of putative zygotes collected were 2825, 957, and 736 for P, O, and CL groups, respectively. Penetration rates were not different (P > 0.05) among groups (90.4%, 94.5%, and 93.7% for P, O, and CL, respectively). However, the monospermic rate was significantly higher (P < 0.05) in the P group (97.4%) when compared to the O or CL groups (66.7% and 5.1% for O and CL, respectively). Moreover, percentages of sows with six or more zygotes, potentially able to carry the pregnancy to term, were 90.8% and 46.5% for P and O groups, respectively (P < 0.05). No sow of the CL group presented six or more zygotes. In conclusion, laparoscopic insemination should be performed only in sows with preovulatory follicles when sex sorted spermatozoa are inseminated using this technology.

This work was supported by CDTI and Fundacion Seneca.