Reproduction in Domestic Animals - Facultad de Ciencias Veterinarias

16 t h International Congress on Animal Reproduction
174 Poster Abstracts
P444
Effective low dose insemination with sex-sorted ram
spermatozoa
Beilby, K*; Grupen, C; Maxwell, WMC; Evans, G
The Faculty of Veterinary Science, The University of Sydney, Sydney,
Australia
Sex-sorted ram sperm is an attractive breeding management tool in
commercial production systems. However, the time required and cost
associated with sorting large commercial sperm doses limits the use of
this technology. Previous research has shown that insemination with
sex-sorted ram sperm resulted in equal, if not superior, fertility
compared to non-sorted controls at dose rates of 1, 5 and 15 million
motile sperm 1 . Pregnancy rates were similar for sex-sorted sperm at
these dose rates. Therefore, the current study aimed to determine the
minimum dose that could be inseminated before fertility was
significantly reduced. Semen was collected from 3 merino rams. Half
of each ejaculate was frozen immediately (non-sorted, control sperm)
and the remaining half was sex-sorted via modified flow cytometry
and then frozen (sorted sperm). Oestrus was synchronised in ewes
(N=138) using progestagen pessaries (12 days) and PMSG at pessary
removal (PR). Ovulation was further controlled using GnRH given
36h after PR. All animals were inseminated by laparoscopy 58h after
PR with either 5 x 10 5 (n = 60), 1 x 10 6 (n = 39) or 15 x 10 6 (n = 39)
sorted or non-sorted, frozen-thawed motile spermatozoa. Pregnancy
rates were assessed at day 60 after insemination using cutaneous realtime
ultrasound. There was no difference in pregnancy rate for groups
inseminated with sorted or non-sorted spermatozoa across all dose
rates. Furthermore, there was no difference between doses of 1 and 15
million spermatozoa. However, the pregnancy rates were lower (P <
0.05) for the 5 x 10 5 sperm dose (15 and 13%, for non-sorted and
sorted, respectively) compared with doses of 1 (49 and 41%) and 15 x
10 6 spermatozoa (44 and 49%). More than 90% of lambs born were of
the predicted sex. This study confirmed that a dose of 1 x 10 6 sexsorted
frozen-thawed motile sperm is the minimum that can be used
for laparoscopic insemination of ewes before fertility is reduced. This
is much lower than the current industry standard of 20-40 x 10 6
frozen-thawed sperm/inseminate. These results provide the sheep
industry with a promising assisted reproductive management strategy
that is not only effective, but practically applicable.
P445
Advantages of the association of glutamine and LDL (Low
Density Lipoprotein) for freezing canine sperm
Bencharif, D 1 *, Tainturier, D 1 , Pascal, O 1 , Larrat, M 1 , Langlois, ML 2 , Barrière,
JP 2 , Amirat-Briand, L 1
1Department of Biotechnologies and Reproductive Pathology, Nantes
Veterinary College, Nantes, France; 2 Department of Reproductive Pathology,
Mother and Child, CHU Hôtel Dieux, Nantes, France
Introduction Various studies have demonstrated the cryoprotective
action of glutamine (Glut). The latter improves spermatozoa motility
and provides superior protection for the cytoplasmic membrane. This
preliminary study aims to determine the ideal concentration of
glutamine when combined with 6% LDL to improve the
cryopreservation of canine semen.
Materials and method Exp n°1: 20 ejaculates were collected from 6
dogs (Beagle, Golden Retriever) aged from 3 to 6 years. Semen with a
motility of between 2 and 5 were frozen in different media: Basic
Medium (BM)+20% egg yolk (e.y), BM+6% LDL, and BM+6%
LDL+60, 70, 80, 90, or 100 mmol of Glut. Following collection, the
spermatic and prostatic fractions were mixed and diluted in the
different freezing media at +37°C to obtain a final concentration of
100x10 6 spermatozoa/ml. Exp n°2: In the same way as for Exp n°1,
the ejaculates were frozen in different media: BM+20% e.y, BM+6%
LDL, and BM+6% LDL+10, 20, 30, 40, or 50 mmol of Glut. Exp n°3:
10 ejaculates were collected from these same 5 dogs, and frozen in:
BM+20% e.y, 6% LDL milieu (e.y extract), and 6 % LDL
medium+20 mmol Glut. The semen was cooled to +4°C for 1 hour
then aspirated into 0.25 ml straws and stored for 30 min at +4°C. The
straws were placed in the liquid nitrogen vapours at -110°C for 10
minutes, before being plunged into the liquid nitrogen. The straws
were thawed in a water bath at +37°C for 30 seconds. The semen was
assessed 10 minutes after thawing with a HAMILTON THORN
CERROS 12 image analyser. For Exp 3, spermatozoa integrity was
analysed using Acridine orange, HOS, PSA-FITC, and Spermac ®
tests.
Results Exp n°1: the motility results were superior in the 6% LDL
media than the other media (BM+20% (e.y), BM+6% LDL+60, 70,
80, 90, 100 mmol of Glut) (43,13% vs 27,3, 28,7, 30,9, 24,1, 31,1,
27,8% respectively) (p