Post-thaw survival of ram spermatozoa and fertility after insemination asaffected by prefreezing sperm concentration and extender composition

摘要

A study was conducted to investigate the effects of prefreezing sperm concentration using two extenders on post-thaw survival and acrosomal status of ram spermatozoa (Experiment 1) and fertility after intrauterine insemination with differing doses of semen (Experiment 2). In autumn (Northern hemisphere), semen was collected by artificial vagina from 8 adult Leccese rams and ejaculates of good quality semen were pooled. Two extender systems for cryopreservation were considered, one based on milk-lactose egg yolk (Milk-LY) and the other based on tris-fructose egg yolk (Tris-FY). Experiment 1 (2 x 6 factorial scheme) examined the in vitro characteristics of spermatozoa in relation to the Milk-LY and Tris-FY extenders and six prefreezing sperm concentrations (50, 100, 200, 400, 500 and 800 x 10(6) spermatozoa/mL). Experiment 2 (2 x 4 factorial) evaluated the influence of the Milk-LY vs Tris-FY extenders and four doses (20; 40, 80 and 160 x 10(6) spermatozoa/0.25 mL) corresponding to prefreezing spermatozoa concentrations of 100, 200, 400 and 800 x 106 spermatozoa/mL, on fertility of ewes inseminated in uterus by laparoscope. Prefreezing sperm concentration influenced (P <0.01) freezability of spermatozoa and affected negatively all the in vitro parameters at 800 x 106 spermatozoa/mL. Overall, Milk-LY tended to ensure higher viability and acrosomal integrity of spermatozoa after thawing at the intermediate sperm densities (range 100 to 500 x 106 spermatozoa/mL). At 500 x 106 spermatozoa/mL concentration corresponded the best condition for survival of spermatozoa (71.2%), acrosome integrity (71.5%) and acrosomal loss (6.0%). At the lowest sperm concentration (50 x 106 spermatozoa/mL), Tris-FY resulted in a higher survival rate than Milk-LY (61.3%, P < 0.05) and lower acrosomal loss (9.7%, P < 0.05). Milk-LY supported spermatozoa motility better than Tris-FY after incubation at sperm concentration between 50 and 400 x 106 spermatozoa/mL (0.05 > P < 0.01). Semen doses of 20 to 40 x 106 spermatozoa/ewe provided satisfactory fertility rates (64 to 81%). The increase of inseminate doses to 160 x 106 spermatozoa/ewe failed to improve fertility, actually tending to decrease lambing rates.